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EDITORIALOsteoporosis and steroid antagonists of theWnt wayOlmos JM, Hernández JL

ORIGINAL ARTICLESBlood sclerostin and Dkk-1 in patients whostart treatment with glucocorticoids.Preliminary resultsGifre L, Ruiz‐Gaspà S, Monegal A, Nomdedeu B,Guañabens N, Peris P

Response of osteoblasts to compounds ofstrontium or calcium: proliferation, differen-tiation, mineralisation and whole genomeresponseFernández‐Murga ML, Serna E, Sanz‐Salvador L,Hervás‐Lorente A, Portero J, Cano A

The reality of osteoporosis in patients hospi-talized in Internal MedicineNeila Calvo S, Nan Nan D, García Ibarbia C, OlmosMartínez JM, González Macías J, HernándezHernández JL

CLINICAL NOTEMultiple skeletal-related events in a patientwith breast cancerHerrero Vicent C, Pascual Pla FJ, Samper Hiraldo JM,Gavilá Gregori J

SPECIAL ARTICLEThe history of SEIOMM (1987-2013)Roig Escofet D, García Borrás J.With the collaboration of: del Pino Montes J, SosaHenríquez M, González Macías J, Moro Álvarez MJ,Herrero L, Calvo Catalá J, Giménez Úbeda E,Quesada Gómez M, Díaz Curiel M, LluchMesquida P, Gómez Alonso C, Olmos Martínez JM,Torrijos Eslava A, Guañabens Gay N, NoguésSolán X

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SUMMARY Vol. 5 - Nº 4 - Noember-December 2013

Our coverFat vesicles stained with oilred resulting in culturedadipocytes differentiationmesenchymal stromal cells

Authors:Antonio Casado Díaz,Raquel Santiago Mora yJosé Manuel QuesadaGómez

Sociedad Española de Investigación Ósea y del Metabolismo Mineral (SEIOMM)

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METHODOLOGY AND DESIGN OF DATA

Pedro Saavedra SantanaJosé María Limiñana Cañal

Committee of expertsEditorial Committee

Teresita Bellido. Ph.D.Department of Medicine, Division of Endocrinology. IndianaUniversity School of Medicine. Indianapolis, Indiana. EstadosUnidos

Ernesto Canalis. MD, PhDSt Francis Hospital and Medical Center, Hartford,Connecticut. University of Connecticut. School of Medicine,Farmington, Connecticut. Estados Unidos

Patricia Clark Peralta. MD, PhDFacultad de Medicina, UNAM. Unidad ClínicaEpidemiológica. Hospital Infantil Federico Gómez. MéxicoDF. México

Dr. Javier del Pino MontesDepartamento de Medicina. Universidad de Salamanca.Sección de Reumatología. Hospital Universitario deSalamanca. Salamanca. España

Dr. Manuel Díaz CurielUniversidad Autónoma de Madrid. Unidad de MetabolismoÓseo. Hospital Fundación Jiménez Díaz. Instituto deInvestigación FJD. Fundación Hispana de Osteoporosis yMetabolismo Mineral (FHOEMO). Madrid. España

Dr. Adolfo Díez PérezUniversidad de Barcelona. Servicio de Medicina Interna.Instituto Municipal de Investigación Médica. (IMIM). Hospitaldel Mar. Barcelona. España

Dr. Oswaldo Daniel MessinaFacultad de Medicina. Universidad de Buenos Aires. HospitalCosme Argerich. Buenos Aires. Argentina

Dra. María Jesús Gómez de Tejada Romero (Redactora Jefe)Universidad de Sevilla. Departamento de Medicina. Sevilla.España

Dr. Manuel Sosa Henríquez (Director)Universidad de Las Palmas de Gran Canaria. Grupo deInvestigación en Osteoporosis y Metabolismo Mineral.Hospital Universitario Insular. Servicio de Medicina Interna.Unidad Metabólica Ósea. Las Palmas de Gran Canaria.España

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Reviewers Volume 5 (2013)

Olmos JM, Hernández JLDepartamento de Medicina Interna - Hospital Universitario Marqués de Valdecilla-IFIMAV - Universidad de Cantabria - RETICEF - Santander

Osteoporosis and steroid antagonistsof the Wnt way

he association between an excessof glucocorticoids and osteoporosiswas indicated more than 80 yearsago by Harvey Cushing when des-cribing the disease which takes hisname. Subsequently, after theintroduction of the glucocorticoidsas anti-inflammatory drugs, it was

confirmed that exogenous hypercortisolism wasalso detrimental to the skeleton, in such a waythat steroidal osteoporosis is now considered tobe the most common form of secondary osteopo-rosis in our ambit1. Osteopenia induced by corti-coids affects predominantly trabecular bone and ismost intense during the first months of treatment,when more than 10% of bone mass may be lost2.In addition to inducing the loss of bone, the glu-cocorticoids alter its quality, which would explainthe notable increase in fractures (nearly 75%)during the first three months of treatment, evenbefore bone mineral density falls.The mechanisms which are involved in the reduc-tion in the quantity and quality of bone tissue arevarious. The glucocorticoids act directly on theosteoblasts, inhibiting their replication, differentia-tion and functional activity, and favouring boththeir apoptosis and that of the osteocytes3,4. Theyalso act on the osteoclasts, reducing their prolife-ration but prolonging their survival3,5. On the otherhand, the glucocorticoids exert an indirect effecton the generation of osteoblast cells by suppres-sing the expression of bone morphogenetic pro-teins (BMP), and of the Runx2 transcription factor,which is required to induce osteoblast differentia-tion from the mesenchymal mother cells6. In addi-tion, these drugs increase the expression ofPPARγ, which favours the differentiation mothercells into adipocytes and slows their differentia-tion into osteoblasts, contributing to an increase in

fat in the bone marrow at the expense of the oste-oblasts and of trabecular bone7. Finally, the corticoids may also intervene in theWnt (wingless) pathway, which acts to modulatethe differentiation and activity of bone cells. Thiscomplex signalling pathway is made up of a num-ber of components, including ligands, membranereceptors, intracellular and antagonist effectors8.The best known mechanisms for transmission ofthe signal of the Wnt ligands are those included inwhat is called the canonical pathway, in which β-catenin plays a central role, although there areother alternative or non-canonical pathwayswhich use different mediators9. The Wnt ligandsare glucoproteins capable of bonding with theirreceptor and initiating the activity of the pathway.The membrane receptors are made up of frizzledproteins and proteins related to the receptor oflow density lipoproteins type 5 and 6 (LRP5 andLRP6). Finally, various types of molecules havebeen described with an inhibitory action on theWnt pathway. In some cases these are moleculeswhich act as decoys which bond with to the Wntligands and thus compete with its bonding to thereceptor. This is the case for some soluble pro-teins of the frizzled type which are secreted intothe extracellular environment. Another inhibitormolecule is sclerostin, a glucoprotein of 190amino-acids coded by the SOST gene which isexpressed in the osteocytes and which bonds toLRP5/6, impeding the formation of the LRP 5/6-frizzled-Wnt complex. The sclerostin is releasedinto the blood circulation, making it possible todetermine its blood concentration10. There is astrong correlation between the content of scleros-tin in the bone and its level in the blood, whichwould indicate that the production of this proteinoccurs in the bone and that its measurement in theblood may reflect its activity in the tissue11.

T

125EDITORIAL / Rev Osteoporos Metab Miner 2013 5;4:125-126

Correspondence: José M. Olmos - Departamento de Medicina Interna - Hospital Universitario Marqués deValdecilla - Avda. Valdecilla s/n - 39008 Santander (Spain)e-mail: [email protected]

Other molecules capable of antagonising the Wntsignals by bonding themselves to the LRP 5/6 andKremen co-receptors are those of the dickkopffamily. There are at least four members of this familyof which the type 1 (Dkk-1) is especially significantin the bone9,12,13. Similarly to sclerostin, Dkk-1 mayalso be determined in the blood, with a higher con-centration of this antagonist having been describedin postmenopausal women or those with low bonemass. It has also been suggested that the reductionin the effect of teriparatide may be related to anincrease in the concentrations of this antagonist14.In this edition of the Review of Osteoporosis andMineral Metabolism Grifé et al.15 analysed thevalues of blood sclerostin and Dkk-1 in patientswho has started treatment with glucocorticoids,confirming that, contrary to what occurs in expe-rimental studies, steroid treatment is associatedwith a reduction in Dkk-1, while no changes areobserved in concentrations of sclerostin. As theauthors note, there are various reasons which mayjustify these findings. Sex, age, renal function,estrogenic status, the existence of associated dise-ases or the quantity of bone mass are determiningfactors in levels of sclerostin and probably also ofDkk-114,16,17. On the other hand, there is also thepossibility that the blood values of both antago-nists do not adequately reflect their expression intissue. Moreover, contrary to what might beexpected a priori, there was no relationship bet-ween the blood concentrations of the antagonistsof the Wnt pathway and those of the markers forbone remodelling. However, it should be notedthat the data published to date have been contra-dictory, with in some cases an inverse relationshipbetween levels of sclerostin and some markers forformation having been described17, while in otherstudies this relationship has not been able to beconfirmed16,18,19. In any case, the results of thisexcellent work invite further studies which analy-se the effect of glucocorticoids on the Wnt path-way antagonists, as well as their relationship withbone mass and markers for remodelling, whichwould without doubt help to clarify the role theWnt pathway antagonists play in the developmentof steroidal osteoporosis.

Bibliography

1. Canalis E, Mazziotti G, Giustina A, Bilezikian JP.Glucocorticoid-induced osteoporosis: Pathophysiologyand therapy. Osteoporos Int 2007;18:1319-28.

2. Weinstein RS. Glucocorticoid-induced bone disease.En: Rosen CJ. Primer on the Metabolic Bone Diseasesand Disorders of Mineral Metabolism. 8th Ed. Iowa:

Wiley & Sons Inc.; 2013. p.473-81.3. Weinstein RS. Glucocorticoid-induced osteoporosis

and osteonecrosis. Endocrinol Clin Metab North Am2012;41:595-611.

4. O’Brien CA, Jia D, Plotkin LI, et al. Glucocorticoids actdirectly on osteoblasts and osteocytes to induce theirapoptosis and reduce bone formation and strength.Endocrinol 2004;145:1835-41.

5. Jia D, O’Brien CA, Stewart SA, et al. Glucocorticoidsact directly on osteoclasts to increase their lifespan andreduce bone density. Endocrinol 2006;147:5592-9.

6. Karsenty G, Kronenberg HM, Settembre C. Geneticcontrol of bone formation. Annu Rev Cel Dev Biol2009;25:629-48.

7. Abdallah BM, Kassem M. New factors controlling thebalance between osteoblastogenesis and adipogenesis.Bone 2012;50:540-5.

8. Velasco J, Riancho JA. La vía Wnt y el hueso. Rev EspEnf Metab Oseas 2008;17:5-9.

9. Baron R, Kneissel M. Wnt signaling in bone homeosta-sis and disease: from human mutations to treatments.Nat Med 2013;19:179-92.

10. McNulty M, Singh RJ, Li X, Bergstralh EJ, Kumar R.Determination of serum and plasma sclerostin concen-trations by enzyme-linked immunoassays. J ClinEndocrinol Metab 2011;96:1156-62.

11. Alonso G, García-Martín A, Muñoz-Torres M. Vía Wnty esclerostina como nuevas dianas para la evaluacióny el tratamiento de la osteoporosis. Med Clin (Barc)2012;139:634-9.

12. Tian E, Zhan F, Walker R, Rasmussen E, Ma Y, BarlogieB, et al. The role of the Wnt-signaling antagonist DKK1in the development of osteolytic lesions in multiplemyeloma. N Engl J Med 2003;349:2483-94.

13. Morvan F, Bouloukos K, Clemen-Lacroix P, Roman S,Suc-Royer I, Vayssier B, et al. Deletion of single alleleof the Dk1 gene leads to increase in bone formationand bone mass. J Bone Miner Res 2006,21:934-45.

14. Gatti D, Viapiana O, Idolazzi L, Fracassi E, Rossini M,Adami S. The waning of teriparatide effect on boneformation markers in postmenopausal osteoporosis isassociated with increasing serum levels of DKK1. JClin Endocrinol Metab 2011;96:1555-9.

15. Grife L, Ruiz-Gaspa S, Monegal A, Nomdedeu B,Guañabens N, Peris P. Esclerostina y Dkk-1 séricos enpacientes que inician tratamiento con glucocorticoi-des. Resultados preliminares. Rev Osteoporos MetabMiner 2014;4:127-32.

16. García-Martín A, Reyes-García R, Rozas-Moreno P,Varsavsky M, Luque-Fernández I, Avilés-Pérez MD, etal. Variables que influyen en las concentraciones deesclerostina en los pacientes con diabetes mellitus tipo2 y su asociación con el metabolismo óseo. RevOsteoporos Metab Miner 2012;4:109-15.

17. Gaudio A, Pennisi P, Bratengier C, Torrisi V, Lidner B,Mangiafico RA, et al. Increased sclerostin levels asso-ciated with bone formation and resorption markers inpatrients with immobilization-induced bone loss. J ClinEndocrinol Metab 2010;95:2248-53.

18. Möder UI, Clowes JA, Hoey K, Peterson JM, McCreadyL, Ousler MJ, et al. Regulation of circulating sclerostinlevels by sex steroids in women and men. J BoneMiner Res 2011;26:27-34.

19. Mirza FS, Padhi ID, Raisz LG, Lorenzo JA. Serum scle-rostin levels negatively correlate with parathyroid hor-mone levels and free estrogen index in postmenopau-sal women. J Clin Endocrinol Metab 2010;95:1991-7.

126EDITORIAL / Rev Osteoporos Metab Miner 2013 5;4:125-126

ORIGINAL ARTICLES / Rev Osteoporos Metab Miner 2013 5;4:127-132127

Gifre L1, Ruiz-Gaspà S2, Monegal A1, Nomdedeu B3, Guañabens N1,2, Peris P1,2

1 Servicio de Reumatología - Unidad de Patología Metabólica Ósea - Hospital Clínico de Barcelona 2 CIBERehd - Hospital Clínico de Barcelona 3 Servicio de Hematología - Hospital Clínico de Barcelona

Blood sclerostin and Dkk-1 in patientswho start treatment with glucocorticoids.Preliminary results

Correspondence: Laia Gifre - Servicio de Reumatología - Hospital Clínic - c/Villarroel, 170 - 08036 Barcelona (Spain)e-mail: [email protected]

Date of receipt: 17/07/2013Date of acceptance: 19/11/2013

Scholarship Working from the SEIOMM to attend the 34 Congress of the ASBMR (Minneapolis, Minnesota 2012).

SummaryBackground and objectives: The Wnt pathway and its inhibitors (sclerostin and Dkk-1) have a primaryrole in the regulation of bone mass and osteoblastogenesis. The objective of this study was to analysethe effect of treatment with glucocorticoids (GCC) on the inhibitors of the Wnt pathway and their rela-tionship with bone mass and the parameters for bone turnover.Methods: A transverse study including 15 patients (9 women and 6 men) with an mean age of 51±21 yearsat the start of treatment with GCC (≥7.5 mg/day, ≤6 months). Levels of sclerostin, blood Dkk-1 and bloodmarkers for bone turnover (procollagen 1 N-terminal propeptide [P1NP], osteocalcin [OC], and carboxy-terminal telopeptide of collagen type 1 [CTX] ) were determined, and bone densitometry ( DXA) in thelumbar spine was carried out, in all patients. The results were compared with a control group.Results: The mean dose of glucocorticoids was 58±21 mg/day, in the majority of patients (73%) indicatedby idiopathic thrombocytopenic purpura. The patients treated with glucocorticoids had a reduction in theparameters for bone formation compared with a control group (OC: 7.4±2.8 vs 24.4±6.2 ng/ml, p<0.01)and a reduction in blood Dkk-1 (29.6±23.6 vs 48.3±15.6 pmol/L, p=0.02). No significant differences wereobserved in values for blood sclerostin, although this correlated positively with the dose of GCC recei-ved and lumbar bone mineral density.Conclusion: Contrary to what is seen in experimental studies, the start of treatment with glucocorticoids isassociated with a reduction in blood levels of Dkk-1. These results indicate the necessity of analysing theseinhibitors and their relationship with remodelling and bone mass during this process over the long term.

Key words: sclerostin, Dkk-1, glucocorticoids.


IntroductionTreatment with glucocorticoids (GCC) is associatedwith a marked loss of bone mass and the develop-ment of fractures in the initial phases of treatment,as well as being one of the most common causesof secondary osteoporosis1. The GCCs act espe-cially on the osteoblasts and osteocytes, reducingthe replication, differentiation and function of theosteoblasts and inducing apoptosis in the osteo-blasts and osteocytes. These changes lead overtime to a reduction in the formation and quality ofbone, which is the finding most characteristic ofsecondary osteoporosis induced by GCC2-4.

The Wnt pathway, a cell signalling pathway,has a fundamental role in the modulation of oste-oblast activity. This pathway is integrated throughvarious components which include ligands, mem-brane receptors, intracellular effectors and antago-nists. The Wnt pathway antagonists, notableamong which are sclerostin and Dkk-1, bond withthe membrane receptors (essentially LRP-5 and -6)and inhibit the activity of this pathway, and con-sequently osteoblast activity.

Recent experimental studies, both in vitro andin vivo, indicate that treatment with GCC reducesthe differentiation of the osteoblasts through theWnt pathway by means of an increase in its inhi-bitors, sclerostin and Dkk-15-7. However, currentlythere are hardly any clinical data on the effect oftreatment with such inhibitors in GCC. Thus theobjective of this study has been to analyse theblood levels of sclerostin and Dkk-1 in patientswho have recently begun treatment with GCC, andevaluate its relationship with the markers for boneremodelling and bone mineral density (BMD).

Patients and methodsStudy populationA transverse study which included patients whohad initiated (<3 months) treatment with dosesequal to, or greater than, 7.5 mg/day of predniso-ne or equivalent. The patients were proposed bythe haematology service of the Hospital ClinicBarcelona (August 2010 to 2012) and recruitedconsecutively.

All the patients complied with the followinginclusion criteria: aged over 18 and with normalvalues of creatinine, liver function, calcium andphosphorus. Excluded were patients who had follo-wed treatment with GCC for more than 6 months,those with diseases or processes which affectedbone metabolism (Paget’s disease, rheumatoid arth-ritis, hyperparathyroidism, hypercortisolism, malab-sorption syndrome, malignant tumours, transplant,pregnancy or recent breastfeeding) and/or thosewho followed treatment with drugs which interferedwith bone metabolism (bisphosphonates, strontiumranelate, selective estrogen receptor modulators,calcitonin, estrogen therapy, denosumab, osteofor-mers thiazides or anti-convulsives).

In all the patients the risk factors for osteoporo-sis were evaluated, including: family history offemoral fracture, personal history of fractures, tobac-co and alcohol consumption, age at menopause,

dietary intake of calcium (mg/day) and history ofrenal lithiasis. Recorded in addition was the cause oftreatment with GCC, the dose and treatment regime(accumulated dose [mg] and duration [days]).

The results were compared with a healthy con-trol group of similar age and sex.

The study was carried out with the approval ofthe ethics committee of the hospital and confor-med with the directives pertinent to research inhumans. All the patients signed their informedconsent to their inclusion.

Analytical testsBlood was taken from all patients at between 8and 10 am after a night of fasting. A biochemicalprofile was carried out which included calcium,phosphorus, creatinine, and total alkaline phos-phatase, determined by standard techniques.

The following biochemical markers for forma-tion were measured: osteocalcin (OC, radioimmu-noassay, Elsa-Osteo-Cis, Gif-sur-Yvette, France)and amino-terminal propeptide of procollagentype I (PINP, automated method Cobas e411,Roche); and of bone resorption: carboxy-terminaltelopeptide of collagen type I (CTX, automatedmethod Cobas e411, Roche).

Blood levels of sclerostin and Dkk-1 were mea-sured using ELISA (Biomedica, Austria), with acoefficient of intravariation of 4-6% and 7-8%, anda coefficient of intervariation of 5-7% and 9-12%,respectively.

Bone mineral densityThe BMD in the lumbar spine and femur were deter-mined in all patients using dual X-ray absorptiometry(DXA; Lunar Prodigy, Radiation CorporationMadison, WI, U.S.). The densitometric risk categories(normal BMD, osteopenia and/or osteoporosis)were defined according to WHO criteria8.

Statistical analysisThe results have been expressed as the mean ±standard deviation from the mean (SD). The diffe-rences between the means of the continuous varia-bles were analysed using the Mann-Whitney non-parametric U test, and the differences between pro-portions by means of the Fisher test. To evaluatethe association between variables the Pearson coef-ficient of correlation was used. Values p<0.05 wereconsidered statistically significant. The statisticalanalysis of the data was carried out using the SPSSsoftware programme (version 18.0, Chicago, U.S.).

ResultsThe clinical characteristics of the patients includedin the study are shown in Table 1.

15 patients were included (9 women [4 postme-nopausal] and 6 men), with an average age of 51±21years. The average dose of GCC used was 58±21mg/day (range 20-100 mg/day) and the averageduration of treatment was 42±24 days (range 4-90days). 73% of the patients had received treatment foridiopathic thrombocytopenic purpura, 20% for hae-molytic anaemia and 3% for both causes (Evans


syndrome). 27% had family his-tory of fracture of the femur and20% had densitometric osteopo-rosis. Dietary intake of calciumwas 593±305 mg/day and onlyone patient was an active smo-ker. The patients who had follo-wed treatment with GCC sho-wed a significant reduction inmarkers for formation with res-pect to the control group (PINP:18±9 vs 47±9 ng/ml, p<0.01; OC:7.4±2.8 vs 24.4±6.2 ng/ml,p<0.01). No significant differen-ces were reported in values ofmarkers for resorption with res-pect to the control group (CTX:0.60±0.27 vs 0.45±0.16 ng/ml,p=0.21).

Those patients treated withGCC had a reduction in bloodlevels of Dkk-1 compared withthe control group (29.6±23.6 vs48.3±15.6 pmol/L, p=0.02),while concentrations of scleros-tin were similar in both groups(39.6±23.3 vs 33.8±20.3, p=0.6)(Figure 1).

In the group of patients trea-ted with GCC, the values of scle-rostin were positively correlatedwith the accumulated dose ofGCC (r=0.573 p=0.026) and lum-bar BMD (r=0.550, p=0.034)(Figure 2). The values of Dkk-1were not related with any of theparameters analysed. No rela-tionship was observed betweenthe values for markers for boneremodelling and blood values ofsclerostin and/or Dkk-1. In thecontrol group, the values ofsclerostin were positively corre-lated with age (r=0.661, p=0.02)(Table 2).

DiscussionContrary to what occurs in experimental studies,the initiation of treatment with GCC in patientswith haematological processes is associated with areduction in blood levels of Dkk-1. This effect,however, differs as a function of the antagonistanalysed, since no significant changes were obser-ved in concentrations of sclerostin at the time ofthe evaluation.

The patients included in this study had lowblood levels of Dkk-1 after initiating treatmentwith GCC at medium-to-high doses. These fin-dings contrasted with the results of earlier experi-mental studies. Thus, in cultures of osteoblastsand osteocytes (MLO-Y4 cells), treatment withdexamethasone resulted in an increase in Dkk-19,which is associated with the dose and period overwhich treatment is received7. There are similar

results in animal experimentation models, inwhich have been observed an increase in theexpression of Dkk-1 in the bone tissue after initia-ting treatment with GCC7 and an attenuation of thedeleterious effect of GCC in the bone by blockingthe effect of Dkk-1 in mice5. Even though the cau-ses of these differences are not clear, the methodof treatment with GCC, including the dose andperiod of treatment, may explain, in part, theseresults6. Neither can we discount a counter-regula-tory effect of this Wnt pathway antagonist in situa-tions of prolonged exposure to GCCs, or thatthese blood levels may indicate not only the cellfunction of the osteoblasts and osteocytes but alsothe number of cells, which, as is well known,reduces (due to an increase in apoptosis) with tre-atment with corticoids.

Table 1. Clinical characteristics of patients treated with GCC

Patients with GCC(n=15)

Age (years) 51±21

Sex (female/male) 9/6

Risk factors for osteoporosis:

BMI (kg/m2) 25±5

Dietary calcium intake (mg/day) 593±305

History of kidney stones (%) 13

Active smoking (%) 7

Alcohol consumption habitual (%) 13

Family history:

Femoral fracture (%) 27

Treatment regimen with GCC:

Daily dose of GCC (mg/day) 58±21

Duration of treatment with GCC (days) 42±24

Cumulative dose of GCC (g) 2.5±1.3

BMD (g/cm2):

Lumbar 1.122±0.156

Femoral neck 0.927±0.113

Total femur 0.958±0.109

GCC: glucocorticoids; BMI: body mass index; BMD: bone mineral density.


However, it is worth commenting that therehave recently been preliminary indications of areduction in blood values of Dkk-1 at three monthsfrom the start of treatment with GCC10, and simi-larly, recent studies have described a paradoxicalresponse of Dkk-1 similar to that observed in ourstudy, in other clinical situations. Thus, contrary toexpectations, reduced values of Dkk-1 have beenobserved in immobilised patients11, and an increasein blood values of Dkk-1 in patients with primaryhyperparathyroidism12. Also it has been described aparadoxical response of Dkk-1 after treatment withteriparatide13 and denosumab14. These authors havesuggested that there is a relationship between boneremodelling and values of Dkk-1. In any case, it isimportant to remember that there may be factorswhich influence blood values of Dkk-1, such as anunderlying disease and concomitant treatment,among others, which should be taken into accountwhen analysing the concentrations of this antago-nist. In addition, the relationship between bloodlevels and expression in tissue is controversial7.

In our study, bloodvalues of sclerostin afterthe initiation of treatmentwith GCC were similar tothose in the control group.However, a positive corre-lation was observed bet-ween values of sclerostinand the accumulated doseof GCC, suggesting aGCC-dependent effect onthis Wnt pathway antago-nist. Studies in mice trea-ted with GCC have repor-ted an increase in theexpression of sclerostinafter treatment3. However,in patients who started tre-atment with GCC (in thefirst 96 hours) a reductionin values of sclerostin hasbeen reported15, a findingwhich has not been obser-ved in postmenopausalwomen treated withGCC10, nor in patients withhypercortisolism due toCushing’s syndrome16, inwhom there has beenobserved an increase invalues of sclerostin.

The circulating levelsof sclerostin in the gene-ral population has beenassociated with age, sex,estrogenic status (pos-tmenopause) and thetotal quantity of bonemass17-19, which meansthat these factors need tobe taken into accountwhen its concentration is

analysed. So, in the healthy subjects included inour study we observed a positive correlation oflevels of sclerostin with age, a finding which wasnot observed in the group treated with GCC, pos-sibly due to the direct effect of the GCC on theWnt pathway. In this group of patients it wasobserved, however, that there was a positivecorrelation between blood levels of sclerostin andlumbar BMD, a relationship which has also beenobserved in other studies and which has beenattributed to a higher production of sclerostin bythe osteocytes, due to the greater quantity ofbone18,20.

The relationship between the markers for boneremodelling and the Wnt pathway antagonists isuncertain and varies in different clinical situations.Thus, García-Martin et al.21,22 in a group of patientswith diabetes mellitus type 2, described an inver-se correlation between values of sclerostin andmarkers for bone formation (bone AP) and boneresorption (sCTX and TRAP 5b). Similar data havebeen reported in the general population18 and in

Figure 1 . Blood levels of sclerostin and Dkk-1 in patients in treatment withGCC (grey) in comparison with healthy controls (white)

Figure 2. Correlation between values of sclerostin, the accumulated dose ofGCC (mg) and lumbar bone mass (lumbar BMD)

* p=0.02

70

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kk-1

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r=0.573p=0.026

r=0.550p=0.034

Sclerostin (pmol/L)

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patients immobilised after avascular-cerebral accident23,although in other situations,such as in patients with chro-nic renal insufficiency, no sig-nificant correlations have beenobserved20,24,25. In our study,although significant reductionsin markers for bone formation(P1NP ad OC) were observedafter treatment with GCC,these were related with neitherDkk-1 nor sclerostin.

The main limitations of thisstudy were the reduced num-ber of patients included, andthe lack of follow up of thesepatients.

In conclusion, the effect oftreatment with GCC on bloodvalues of the Wnt pathwayantagonists differs as a func-tion of the antagonist beingevaluated. While levels ofDkk-1 diminished at the startof treatment, the values ofsclerostin showed no signifi-cant changes. All this suggeststhe necessity of carrying outprospective studies including agreater number of patientswith better follow up to analy-se the effect of the GCCs onthe Wnt pathway antagonists.

Conflict of interest: Theauthors declare that there is noconflict of interest.

This work has been funded bygrants from the Hospital ClinicBarcelona and the CatalanSociety of Rheumatology.

Bibliography

1. van Staa TP, Leufkens HG, Cooper C. The epidemio-logy of corticosteroid-induced osteoporosis: a meta-analysis. Osteoporos Int 2002;13:777-87.

2. Hernandez MV, Guanabens N, Alvarez L, Monegal A,Peris P, Riba J, et al. Immunocytochemical evidence onthe effects of glucocorticoids on type I collagensynthesis in human osteoblastic cells. Calcif Tissue Int2004;74:284-93.

3. Yao W, Cheng Z, Busse C, Pham A, Nakamura MC,Lane NE. Glucocorticoid excess in mice results in earlyactivation of osteoclastogenesis and adipogenesis andprolonged suppression of osteogenesis: a longitudinalstudy of gene expression in bone tissue from glucocor-ticoid-treated mice. Arthritis Rheum 2008;58:1674-86.

4. Canalis E, Mazziotti G, Giustina A, Bilezikian JP.Glucocorticoid-induced osteoporosis: pathophysiologyand therapy. Osteoporos Int 2007;18:1319-28.

5. Wang FS, Ko JY, Yeh DW, Ke HC, Wu HL. Modulationof Dickkopf-1 attenuates glucocorticoid induction ofosteoblast apoptosis, adipocytic differentiation, andbone mass loss. Endocrinology 2008;149:1793-801.

6. Mak W, Shao X, Dunstan CR, Seibel MJ, Zhou H.

Biphasic glucocorticoid-dependent regulation of Wntexpression and its inhibitors in mature osteoblasticcells. Calcif Tissue Int 2009;85:538-45.

7. Thiele S, Ziegler N, Tsourdi E, De Bosscher K,Tuckermann JP, Hofbauer LC, et al. Selective glucocor-ticoid receptor modulation maintains bone mineraldensity in mice. J Bone Miner Res 2012;27:2242-50.

8. Assessment of fracture risk and its application to scre-ening for postmenopausal osteoporosis. Report of aWHO Study Group. World Health Organ Tech Rep Ser1994;843:1-129.

9. Ohnaka K, Taniguchi H, Kawate H, Nawata H,Takayanagi R. Glucocorticoid enhances the expressionof dickkopf-1 in human osteoblasts: novel mechanismof glucocorticoid-induced osteoporosis. BiochemBiophys Res Commun 2004;318:259-64.

10. Gossiel F LN, Eastell R. The Effect of GlucocorticoidTherapy on Regulators of Bone Formation inPostmenopausal Women Treated with Teriparatide orAlendronate. ASBMR Annual Meeting 2011:S80.

11. Frings-Meuthen P, Boehme G, Liphardt AM, BaeckerN, Heer M, Rittweger J. Sclerostin and DKK1 levelsduring 14 and 21 days of bed rest in healthy youngmen. J Musculoskelet Neuronal Interact 2013;13:45-52.

12. Viapiana O, Fracassi E, Troplini S, Idolazzi L, RossiniM, Adami S, et al. Sclerostin and DKK1 in primaryhyperparathyroidism. Calcif Tissue Int 2013;92:324-9.

Table 2. Correlation between values of sclerostin and values of Dkk-1,markers for bone remodelling and the other parameters analysed(Pearson r with and without adjusting for age§)

GCC(n=15)

Controls(n=20)

Age (years) 0.03/- 0.661*/-

BMI (kg/m2) 0.254/0.267§ 0.320/-0.036§

Markers of bone remodeling:

PINP (ng/ml) -0.295/-0.375§ -0.12/0.131§

OC (ng/ml) -0.204/-0.212§ -0.593/0.134§

CTX (ng/ml) -0.249/-0.360§ 0.098/0.296§

Wnt pathway antagonists:

Dkk-1 0.01/0.01§ -0.205/0.309§

BMD (g/cm2):

Lumbar 0.537*/0.651§* -

Femoral neck 0.171/0.30§ -

Total femur 0.187/0.258§ -

GCC: glucocorticoids; BMI: body mass index; PINP: amino-terminal pro-peptide of pro-collagen type I; OC: osteocalcin; CTX: carboxy-terminaltelopeptide of collagen type I; BMD: bone mineral density.* p<0.05


13. Gatti D, Viapiana O, Idolazzi L, Fracassi E, Rossini M,Adami S. The waning of teriparatide effect on boneformation markers in postmenopausal osteoporosis isassociated with increasing serum levels of DKK1. JClin Endocrinol Metab 2011;96:1555-9.

14. Gatti D, Viapiana O, Fracassi E, Idolazzi L, Dartizio C,Povino MR, et al. Sclerostin and DKK1 in postmeno-pausal osteoporosis treated with denosumab. J BoneMiner Res 2012;27:2259-63.

15. Brabnikova Maresova K, Pavelka K, Stepan JJ. Acute effectsof glucocorticoids on serum markers of osteoclasts, osteo-blasts, and osteocytes. Calcif Tissue Int 2013;92:354-61.

16. Belaya ZE, Rozhinskaya LY, Melnichenko GA,Solodovnikov AG, Dragunova NV, Iljin AV, et al. Serumextracellular secreted antagonists of the canonicalWnt/beta-catenin signaling pathway in patients withCushing's syndrome. Osteoporos Int 2013;24:2191-9.

17. Modder UI, Clowes JA, Hoey K, Peterson JM,McCready L, Oursler MJ, et al. Regulation of circulatingsclerostin levels by sex steroids in women and in men.J Bone Miner Res 2010;26:27-34.

18. Modder UI, Hoey KA, Amin S, McCready LK,Achenbach SJ, Riggs BL, et al. Relation of age, gender,and bone mass to circulating sclerostin levels inwomen and men. J Bone Miner Res 2011;26:373-9.

19. Mirza FS, Padhi ID, Raisz LG, Lorenzo JA. Serum scle-rostin levels negatively correlate with parathyroid hor-mone levels and free estrogen index in postmenopau-sal women. J Clin Endocrinol Metab 2010;95:1991-7.

20. Polyzos SA, Anastasilakis AD, Bratengeier C,

Woloszczuk W, Papatheodorou A, Terpos E. Serumsclerostin levels positively correlate with lumbar spinalbone mineral density in postmenopausal women--thesix-month effect of risedronate and teriparatide.Osteoporos Int 2012;23:1171-6.

21. García-Martin A, Rozas-Moreno P, Reyes-Garcia R,Morales-Santana S, Garcia-Fontana B, Garcia-SalcedoJA, et al. Circulating levels of sclerostin are increasedin patients with type 2 diabetes mellitus. J ClinEndocrinol Metab 2012;97:234-41.

22. García-Martín A, Reyes-García R, Rozas-Moreno P,Varsavsky M, Luque-Fernández I, Avilés-Pérez M, et al.Variables que influyen en las concentraciones de escle-rostina en los pacientes con diabetes mellitus tipo 2 ysu asociación con el metabolismo óseo. RevOsteoporos Metab Miner 2012;4:109-115.

23. Gaudio A, Pennisi P, Bratengeier C, Torrisi V, LindnerB, Mangiafico RA, et al. Increased sclerostin serumlevels associated with bone formation and resorptionmarkers in patients with immobilization-induced boneloss. J Clin Endocrinol Metab 2010;95:2248-53.

24. Cejka D, Jager-Lansky A, Kieweg H, Weber M,Bieglmayer C, Haider DG, et al. Sclerostin serum levelscorrelate positively with bone mineral density andmicroarchitecture in haemodialysis patients. NephrolDial Transplant 2012;27:226-30.

25. van Lierop AH, Witteveen JE, Hamdy NA, PapapoulosSE. Patients with primary hyperparathyroidism havelower circulating sclerostin levels than euparathyroidcontrols. Eur J Endocrinol 2010;163:833-7.


Fernández-Murga ML1*, Serna E2, Sanz-Salvador L1, Hervás-Lorente A1, Portero J2, Cano A1,3

1 Laboratorio de Ecofisiología, Nutrición y Salud, IATA-CSIC - Fundación para el Fomento de la Investigación Sanitaria y Biomédica dela Comunidad Valenciana (FISABIO) - Hospital Universitario Dr. Peset - Valencia2 Unidad Central de Investigación en Medicina-INCLIVA - Universidad de Valencia - Valencia3 Departamento de Pediatría, Obstetricia y Ginecología - Universidad de Valencia y Servicio de Obstetricia y Ginecología - HospitalUniversitario Dr. Peset - Valencia

Response of osteoblasts to compounds of strontium or calcium: proliferation,differentiation, mineralisation andwhole genome response

Correspondence: María Leonor Fernández Murga - Laboratorio de Ecofisiología, Nutrición y Salud, IATA-CSIC -Hospital Universitario Dr. Peset - La Coma s/n - 46100 Paterna - Valencia (Spain)e-mail: [email protected]


Work awarded for SEIOMM in SEIOMM XVII Congress in La Coruña, 2011.

SummaryBackground: The mechanisms which trigger osteogenesis are not yet clear. The objective of this studywas to evaluate the role of strontium and calcium, provided in different molecular forms, as inductors ofdifferent mechanisms of osteoblast stimulus, including proliferation, differentiation and mineralisation ofpreosteoblast cells. The whole genomic response was also investigated using the microarray technique.Methods: An experimental study was designed with murine preosteoblast cells MC3T3-E1, which were sti-mulated for 3 hours and 7 days. Biochemical and genome gene expression studies of mouse (Affymetrix)were carried out.Results: Strontium bonded with ranelate (SrRn) was the most powerful inductor of the capacity of mine-ralisation in comparison with the other compounds used (2.55 times that of the control). The studies ofwhole gene expression showed that after 3 hours 2030 genes change, of which 1644 are specific to thisphase. On the other hand, after 7 days of treatment only 329 genes change, of which 147 are specific.The biological processes most enriched after 3 hours are those involved in the regulation of transcription(147 genes), metabolic processes (140 genes) and protein phosphorylation (44 genes) among others,while at 7 days these are changes relating to the cell cycle (18 genes) and carbohydrate metabolism ingeneral (12 genes).Conclusion: Strontium bonded with the ranelate anion performed as the most powerful inductor of oste-ogenesis compared with other anions such as chloride or the hydroxides. The stimulation for 3 hoursshowed greater changes in gene expression in comparison with 7 days. The biological processes affec-ted may be useful in speculating on the signalling cascades involved in the activation of the osteoblast,and on new molecular targets for therapeutic purposes.

Key words: osteoporosis, osteogenesis, strontium, calcium, gene microarray, gene expression.


IntroductionThe skeleton provides support and mineral equili-brium to the organism. Skeletal tissue is formedduring growth and is maintained through adult lifeby the continuous renewal of the bone matrix,through a process known as bone remodelling. Inthis process an important role is played by twotypes of cell, osteoclasts and osteoblasts. Duringgrowth bone formation exceeds resorption, resul-ting in a net gain. Conversely, during aging a dise-quilibrium occurs which results in a negative bonebalance1. Extrinsic mechanisms, such as changesin levels of hormones and growth factors, andintrinsic mechanisms associated with cell senes-cence, may cause the dysfunction of the osteo-blasts2,3. This causes an increase in the number ofbone remodelling units, which encourages trabe-cular perforation and a reduction in endocorticalbone which in turn cause reduced bone resistan-ce4,5.

With the aim of limiting the excess resorptionwhich follows the menopause, a series of anti-resorptive drugs has been developed, such as bis-phosphonates, monoclonal antibodies againstcytokines involved in osteoclastic differentiation,cathepsin K inhibitors, etc6,7. Their long-termeffects are, however, not known7. A significantchallenge in the treatment of osteoporosis is theidentification of strategies capable of reversing thedeterioration of bone formation linked to age. Todate, the number of anabolic agents which promo-te osteoblastogenesis is limited. The availability ofparathyroid hormone (PTH) was a significantadvance in the treatment of osteoporosis8. Theintermittent provision of PTH increases bone for-mation in patients with osteoporosis, which resultsin an increase in trabecular bone mass and corti-cal thickness8-10. However, this anabolic treatmenthas some limitations linked to the low half-life andhigh cost of this molecule.

Strontium (Sr), a cation close to calcium in theperiodic table has been shown to act pharmacolo-gically on bone metabolism11,12. In certain experi-mental models it appears to develop an anabolicaction, which has generated interest in associatedpathways capable of promoting bone formation.In ovariectomised rats (OVX), an animal model forpostmenopausal osteoporosis, it was observedthat, both short- and long-term, treatment with Srimpedes the loss of trabecular bone induced bydeficiency in estrogens13,14. Controlled clinical trialsin postmenopausal osteoporotic women showedthat treatment with Sr reduced the relative risk ofvertebral fracture in comparison with the placebogroup15, as well as the risk of all non-vertebralfractures and hip fracture, as was analysed in asubgroup of patients of advanced age16,17.

Various mechanisms may explain the reductionin risk of fracture induced by Sr in osteoporosis.One of the possible mechanisms is the increase inbone mineral density (BMD)18. Another wouldinvolve the effect of Sr on bone resorption andformation19. In support of this finding, it was foundthat 12 months of treatment with Sr causes an

increase in the number of osteoblasts, formationof the matrix and a reduction in the number ofosteoclasts in patients with osteoporosis20,21. Inaddition to the effects on bone cells and bonemicroarchitecture, Sr can increase bone resistanceby means of changes in the properties of the bonematrix. A small fraction (less than 10%) of stron-tium is incorporated in the bone22. Given this, thebeneficial effect of Sr on bone resistance cannotsolely result from its pharmacological effects onthe bone cells, bone remodelling and the microar-chitecture, but may also arise from its effects onthe properties of the bone matrix. Therefore, morebasic and clinical analyses are required to clarifythese effects at a molecular level.

In this work, we have studied the effects of dif-ferent compounds of Sr and a calcium salt on dif-ferent parameters related to osteogenesis usingpreosteoblast cultures as a model. In addition, wecarried out a whole genome study by means ofmicroarray with the aim of obtaining informationon the genes and signalling pathways involved inosteoformation. For this, we used a microarray ofmouse GeneChip Mouse Gene 1.0 ST, which con-tains more than 750,000 probes which represent28,000 genes. The genes were analysed at diffe-rent times, short (3 hours, acute phase) or long (7days, chronic phase). The changes in gene expres-sion were grouped according to the most notablemetabolic processes and those related to the pro-liferation, differentiation and activity of the osteo-blasts.

Materials and methodsCell culturesThe murine preosteoblast cell line MC3T3-E1(Sigma-Aldrich, St. Louis, US) was used. The cellswere kept in a culture medium composed of α-MEM (Invitrogen, California, US) supplementedwith 10% foetal bovine serum (FBS, Invitrogen,California, US), 2 mM L-glutamine, 100 u/ml peni-cillin and 100 µg streptomycin (Invitrogen,California, US) in an incubation cabinet at 37 ° Cin a humid atmosphere with 5% CO2. In order tocarry out the tests they were left to grow until 60-70% confluence.

Before any stimulation the cells were kept for24 hours in a medium with 2.5% FBS. After thisperiod the cells were treated with strontium rane-late (SrRn, AK Scientific Inc., US), strontium chlo-ride (SrCl2), calcium chloride (CaCl2) and stron-tium hydroxide ( [Sr(OH)2] (Sigma-Aldrich) at aconcentration of 2 mM for the periods specified ineach experiment . The treatments were carried outin a medium of α-MEM supplemented with 2.5%FBS, 2 mM L-glutamine, 100 u/ml penicillin and100 µg/ml streptomycin. For the prolonged treat-ments, the cultivation medium was refreshed by50% and the corresponding treatment added every3 days.

Cell proliferation test (XTT)The cells were seeded in a plate of 96 wells (den-sity 4x103 cells/well). They were incubated for 48


hours with the aforementioned stimuli. The cellproliferation was measured with the CellProliferation ELISA II test (XTT-assay, RocheApplied Science, Mannheim, Germany) followingthe manufacturer’s instructions. After the incuba-tion period, a measurement of absorbance at 450and 650 nm at 37° was taken in a Victor™X3 2030Multilabel Reader (Perkin Elmer, Massachusetts,US.) plate reader.

Measurement of alkaline phosphatase (ALP) activityThe MC3T3-E1 cells were seeded in plates of 96wells at a confluence of 60% and subjected to theaforementioned stimuli for 24 hours, 3 days and 7days. After the treatments the cells were washedwith PBS, harvested with Triton 100 0.1% PBS andsonicated at 4°C. After centrifuging at 4°C at20,000 g for 5 minutes, the supernatants were har-vested, from which the alkaline phosphatase acti-vity was determined using the ALP Reagent test(Thermo scientific, Massachusetts, U.S.). Theabsorbance at 405 and 660 nm (reference absor-bance) was determined at 37°C each minute for 10minutes in a Victor™X3 2030 Multilabel Reader(Perkin Elmer, U.S.) plate reader.

The concentration of protein was determinedusing the Bradford colorimetric method (BioRad,Hemel Hempstead, United Kingdom). The ALPactivity was expressed as µg of product formedper minute of reaction over the quantity of proteinpresent in the supernatant (U/mg).

Mineralisation testsTo quantify the mineralisation alizarin red wasused (ARS, Sigma-Aldritch, St. Lois, Missouri, U.S.)an organic colourant capable of bonding with thedeposits of calcium in the cells. It is used therefo-re as a marker for the capacity to form a calcifiedmatrix. For the experiments, the MC3T3-E1 cellswere seeded in plates and incubated with the dif-ferent treatments SrRn, SrCl2, CaCL2 and Sr(OH)2for 24 hours , 3 days and 7 days. Then the cellswere washed with PBS and fixed with formal-dehyde 10% for 15 min. After fixing, they were tre-ated with a solution of ARS (40 mM, pH4.2) for 20min at room temperature. To detect their capacityfor mineralisation the cells marked with ARS wereincubated with acetic acid 10% for 30 min at roomtemperature. Subsequently, the cell suspensionwas harvested and incubated at 85°C for 10 minfollowed by 5 mins on ice, to harvest the superna-tants after centrifuging at 20,000 g for 15 min.Potassium hydroxide (KOH) 10% was added toeach supernatant, and finally, the intensity offixing with ARS was determined by measuringabsorbance at 405 nm in a plate reader Victor™X32030 Multilabel Reader (Perkin Elmer, U.S.).

Extraction of total RNAThe cells were cultivated in triplicate at a concen-tration of 106 cells per sample. After this, theywere divided into an equal number of sampleswith or without treatment with 2 mM of SrRn for3 hours and for 7 days. After these periods of incu-

bation the culture medium was eliminated and theextraction of total RNA was carried using the trizolmethod according to the maker’s (Invitrogen,California, U.S.) specifications. The quantity andpurity obtained was confirmed by determiningabsorbance at 260 and 280 nm using NanodropND-1000 (Thermo Fisher Scientific, Wilmington,Delaware, U.S.).

Microarrays of gene expressionThe analysis of the expression of whole genomewas performed using the GeneChip Mouse Gene1.0 ST Array (Affymetrix, Santa Clara, California,U.S.) microarray which has more than 750,000probes, with a longitude of 25 oligonucleotideswhich represent 28,000 mouse genes. Biologicaltriplicates were obtained for each condition: con-trols, 3 hours and 7 days of treatment. We startedfrom 300 ng of total RNA, having previously analy-sed its integrity ((Bioanalizador, 2100 Agilent), inorder to synthesise the single-stranded cDNA. Itslater fragmentation and marking was carried outfollowing the manufacturer’s (Affymetrix, SantaClara, California, U.S.) indications. The hybridisa-tion took place over 17 hours at 45°C with rotationat 60 rpm in the oven recommended byAffymetrix: “GeneChip Hybridization Oven 640”(Affymetrix, Santa Clara, California, U.S.). Afterthis period, the microarrays were passed througha wash process and, finally, the marking wascarried out with streptavidin-phycoerythrin usingthe “GeneChip Fluidics Station 450” (Affymetrix,Santa Clara, California, U.S.). Subsequently, themicroarrays were scanned with “GeneChipScanner 3000 7G” (Affymetrix, Santa Clara,California, U.S.) and images of each of the sam-ples obtained. The quality controls of these ima-ges were performed using the programme“Affymetrix Genechip Command Console”.

Statistical analysis of the resultsThe normalised .CEL files of the images of thesamples from all the experiments were analysedwith the software Partek Genomics Suite version6.6 (Partek Inc., St. Louis, Missouri, U.S.). Themicroarrays were normalised using the RMA algo-rithm. The differential gene expression was iden-tified using ANOVA by means of a highly restricti-ve analysis using False Discovery Rate <0.05(FDR<0.05).

The expression of the baseline genes whichchanged over time between the control at 3 hourscompared with the control at 7 days were dis-counted from the analysis to enable the more spe-cific observation of those genes which changedafter the stimulation with SrRn.

The cell processes included the significantgenes, both for the acute and the chronic phases,were analysed using the software Pathway Studioversion 9.0, database ResNet9.0;2012Q3 (Mammal)(Ariadne Genomics, Rockville, Maryland, U.S.).

The biochemical and cell proliferation dataobtained were analysed with the programmeGraphPad Prism version 4 (GraphPad Software,


Inc., California U.S.). The comparative statisticalstudy was carried out using single factor analysisof variance (ANOVA) and the Bonferroni test forthe multiple comparatives. The level of statisticalsignificance was established at values p<0.05, forall variables analysed.

ResultsThe effect of different salts of Sr in MC3T3-E1 pre-osteoblast culturesCell proliferationFigure 1A presents the effect of different salts stu-died on cell proliferation determined by the XTTmethod. All the stimuli induced cell proliferationnegative with respect to the control after 48 hoursof treatment. CaCl2 induced the highest degree ofproliferation, with an increase of 2.13 times, whichwas highly significant (p<0.001). All the salts ofstrontium produced a slight increase in cellularproliferation (p<0.05) in the period studied withrespect to the controls. There were no significantdifferences between the salts studied.

Cell differentiation (alkaline phosphatase andmineralisation)ALP is a marker for early osteoblast differentiationand an increase in its activity is considered to bean indicator of an increase in the activity of themature osteoblasts. Figure 1B shows the activity ofALP with the different salts over 7 days.Measurements were also taken at shorter timeperiods (24 hours and 3 days) but no increase inALP activity was observed with any of the stimuli(data not shown), therefore, during this period oftime the cells still remained undifferentiated.However, the evaluation at 7 days produced anincrease in ALP activity, such that the strontium asa chloride or hydroxide salt achieved an increaseof 4.45 which was highly statistically significant(p<0.001). The salts CaCL2 or SrRn showed alower increase in the activity of ALP, of approxi-mately 2 times with respect to the control, of sta-tistical significance (p<0.01).

Since it is known that Sr is incorporated intothe bone matrix, the impact of the salts of Sr andof CaCl2 on the capacity of the mineralisation ofthe pre-osteoblasts MC3T3-E1 was evaluated.Similar to what was observed in the ALP tests nomineralisation was observed at 24 hours or 3 days(data not shown), but the test was positive at 7days after the stimulus (Figure 1C). The SrRnachieved an increase of 2.55 times in mineralisa-tion measured by ARS with respect to the controland 2.2 times if we compare it with other com-pounds of strontium (p<0.05), which also appearto induce mineralisation slightly when comparedto the control. The cells stimulated with SrRn sho-wed a greater capacity for mineralisation eventhan the CaCL2 salt studied, with a strong statisti-cal significance (p=0.06).

In conclusion, the Sr bonded with ranelateinduced slight cell proliferation, and an increase inALP activity sufficient to achieve the maximummineralising capacity of all the salts investigated.

With this knowledge, the investigation of the geneexpression mediated by this salt was begun.

Analysis of gene expression using microarrayThe effect of SrRn on the gene profile of MC3T3-E1 cells was also investigated over 3 and 7 days ofstimulus. These periods of study were selected inearlier studies by our group, where we detectedthe most intense changes in gene expression rela-ted to signalling cascades Wnt and NFAT, bothimportant routes in the process of osteogenesis23.The results obtained by microarray were made intriplicate and are represented in a principal com-ponents analysis (PCA) map (Figure 2). It is obser-ved that the samples from the same conditionresemble each other and therefore the ellipses aresmall. The fact that the ellipses do not overlapconfirms to us that there are genetic differenceswith the other conditions. The samples includedwithin the 3 hour stimulus were very different,especially in comparison with those not treated(control). After 7 days, the differences narrowed,which suggests that the impact on gene expres-sion reduces in relation to the acute phase.

A comparative statistical analysis was carriedout using single factor analysis of variance(ANOVA) with an FDR ≤0.05. We observed that inthe acute phase there are 2,030 genes whichmodify their expression, of which specific to thisphase are 1,644 genes which change significantlywhen the samples are treated with SrRn. On theother hand, after 7 days of stimulus, 329 geneschanges in a statistically significant way, with only147 of those specific to the chronic phase. Theremaining genes were common to the two phases.In this study those genes which changed theirexpression among the controls at 3 hours and 7days were discounted and we focussed on thegenes which changed specifically in each periodof time. Represented in Figures 3A and 3B are thebiological processes in which those genes withdifferential expression may be integrated. Therepresentation is made according to the number ofgenes which change, from higher to lower,although it should be underlined that all the chan-ges presented are statistically significant. Thecellular processes differed significantly accordingto the period of treatment, 3 hours or 7 days, withSrRn. In the acute phase the cellular processesassociated with those genes whose expressionchanged corresponded to the regulation of trans-cription (147 genes), to general metabolic proces-ses (140 genes), to transport (89 genes, of which52 are related to transport of proteins) and 44genes related to the phosphorylation of proteins.On the other hand, processes such as cell death,DNA repair or response to DNA damage were lessenhanced. In contrast, at 7 days of stimulation,chronic phase (Figure 3B), the changes were dif-ferent and the biological processes most enhancedwere those metabolic processes which are invol-ved above all in the metabolism of glucose (6genes) and of carbohydrates in general (6 genes),cell cycle and division (10 genes), cell prolifera-


tion (6 genes) and, to a lesser extent, thoserelated to hypoxia and lipopolysaccharidemetabolism.

DiscussionIn this work, we studied the effect of threecompounds of strontium on murine MC3T3-E1 pre-osteoblasts. We observed that Sr pro-motes cell proliferation and differentiation, aswell as mineralisation, but its potential chan-ges according to the anion to which it is bon-ded. If in the culture medium there is a sour-ce of organic phosphate, we observe discre-te zones of mineral deposits which containhydroxyapatite. Among the strontium com-pounds studied, the Sr bonded with ranelatewas the most powerful inductor of minerali-sation in comparison with chloride or hydro-xide (Figure 1C). This process was accompa-nied by a small increase in the activity of ALPassociated with the osteoblast phenotype.Conversely, the highest alkaline phosphataseactivity was observed with Sr bonded withboth chloride or hydroxide, demonstratingthe versatility of the action of the cationaccording to the molecular framework whichaccompanies it. The cells used in this studyhave been very well described and are there-fore an excellent model for studies of mine-ralisation, however they have a low expres-sion of ALP. Other subclones of the same cellline have been characterised which, in con-trast, have high ALP activity and do not havemineralisation capability24. It is known thatlittle ALP (0.05 U/mg) activity is required inorder for inorganic phosphorus to be obtai-ned in vitro25. In mesenchymal cells frombone medulla low ALP activity has also beenobserved, as well as their being capable ofmineralisation26. Therefore, the activity ofALP in the samples stimulated with SrRn isnecessary and sufficient to produce a mine-ralisation matrix.

The sequence of the induction of the for-mation of mineralised foci progressed in aregular order over the time periods studiedof 0-1-3-7 days. Experiments carried out withascorbic acid have shown that this process isprolonged to 2 to 3 weeks27, showing the effi-cacy of the action of SrRn. Presumably, suc-cessive waves of change in gene expressionoccur over the period of 0-7 days requiredfor the differentiation of the pre-osteoblaststo a mineralisation phenotype. We carriedout studies of up to 21 days of treatment withthe stimuli, observing that the most intensechanges in the cell, such as gene expressionand activation of signalling cascades relatedto osteoformation processes, occur veryshortly after the stimulus28.

Given the high degree of mineralisationobserved, and the fact that this process canonly be executed by mature osteoblasts,strontium bonded with ranelate (SrRn) is for

Figure 1. The effect of different compounds of Sr inMC3T3-E pre-osteoblast cultures. Cell cultures without sti-mulation and treated with CaCL2 were used as controls.(A) cell proliferation during 48 hours of stimulation; (Band C) alkaline phosphatase activity; ALP and the capabi-lity of mineralisation determined by the alizarin redmethod after 7 days of treatment, respectively. Theresults are represented as the mean ± standard deviationof triplicates, *p<0.05, **p<0.01, ***p<0.001, comparisonsmade with respect to the control (0)

**

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Stimulations (2 mM)


us an excellent stimulus for the study of the cellprocesses which occur when osteogenesis orbone formation take place in these cells. We haveused an expression matrix from the mouse whichcontains 28,000 genes. Earlier studies only used588 or 8,700 probes29,30. Our matrix allows theinvestigation of the genes of numerous growthfactors, cytokines, interleukins and their receptors,as well as key genes involved in different stages ofembryo development. MC3T3-E1 cells are a well-established osteogenic clonal cell line, which pro-vide an excellent model for the study of patternsof gene expression in osteoblast differentiation. Inour study, the SrRn induced in these cells a pro-cess of maturation which can be divided into twophases, which result in the formation of a minera-lised matrix (Figure 1C). Each of these phasesrequires strictly regulated expression of genes andtranscription factors.

The first phase of this process of maturation (3hours after stimulation, acute phase) is triggeredon contact with the stimulus, SrRn. During thisphase the MC3T3-E1 cells are not yet differentia-ted, and are not capable of producing a minerali-sation matrix. However, in these cells the expres-sion of many genes is activated, up to a total of1,644, in which the 10 most enriched cell proces-ses from our global gene expression study areshown in Figure 3A. In this stage many geneschange which are related to the regulation of

transcription, metabolic processes in general,molecular transport, etc. It is notable thatamong those general metabolic process enri-ched in this short period of time is proteinphosphorylation-dephosphorylation, a pro-cess which participates in a large number ofcell routes of great interest. Some of thesehave been shown to be involved in the matu-ration of the osteoblasts, such as the Wnt andNFAT signalling routes31,32. However, wecurrently have insufficient information regar-ding the cell pathways which are activatedduring the differentiation, proliferation andmaturation of osteoblasts, in which doubtlessthe processes of protein phosphorylation-dephosphorylation are crucial.

In accord with this pattern, in an earlierwork we confirmed that the Wnt/β-cateninand NFAT pathways, both powerful osteopro-genitors, are activated within only 15 minutesof stimulus with SrRn, which rapidly induceschanges in gene expression. In contrast, 21day cultures do not show activation of thesepathways (data not shown).

Once confluence is achieved, and coexis-tent with an increase in ALP activity andmineral matrix deposit (Figures 1B and 1C),the cells enter what is known as the differen-tiation phase (7 days). This is characterisedby an increase in the formation of the bonematrix, and is associated with the change inexpression of 147 genes. The cell processeswhich are increased in this phase are thoserelated to the energetic state of the cells: car-

bohydrate metabolism in general, metabolic pro-cess of glucose and other carbohydrates, responseto lipopolysaccharides or gluconeogenesis. Thismakes this cell line an excellent model for thestudy of the molecular events involved in the pro-cess of osteogenesis. The microarrays allow thesimultaneous monitoring of a great number ofgenes associated with the metabolism of bone,which enables the detection of potential targets fortherapeutic or diagnostic purposes. The use ofmicroarrays is becoming ever more accessible andthe data generated from this type of approach arelargely descriptive. However, the general informa-tion which is obtained is a very useful baseline formore analytical studies of functional routes whichcome into play during cell differentiation, an eventwhich is mediated by receptors, transcription fac-tors, proteins, enzymes, etc., which are in turnregulated by changes in gene expression.

The analysis here presented offers a dynamicvision of these events during the differentiation ofosteoblasts. The results of the expression matrixwhich is presented here complement earlier stu-dies29,30. However, many earlier gene studies inosteoblasts were analysed under different condi-tions, with different cell lines, or under inductionby different agents. The ability to compare datacollected from hundreds of genes under a combi-nation of conditions with data from other systemsstrengthens our general understanding of the

Figure 2. Representation of principal components analysis(PCA) map. All the samples are represented in 3D througha non-supervised analysis of principal components. Theellipses are drawn to include 61.3% of the genes in eachgroup


molecular basis of osteogenesis.These data are valuable not only fora better understanding of osteoge-nesis but also for comparison withother types of tissues. This informa-tion should help in the develop-ment of more efficacious treatmentfor bone disorders and in the pre-diction of secondary effects onbone metabolism of drugs whichare aimed at the same factors forintervention in other diseases.

Bibliography

1. Khosla S, Riggs BL. Pathophysiologyof age-related bone loss and osteopo-rosis. Endocrinol Metab Clin NorthAm 2005;34:1015-30.

2. Flynn JM, Spusta SC, Rosen CJ, MelovS. Single cell gene expression profi-ling of cortical osteoblast lineagecells. Bone 2013;53:174-81.

3. Khosla S, Oursler MJ, Monroe DG.Estrogen and the skeleton. TrendsEndocrinol Metab 2012;23:576-81.

4. Raisz LG. Pathogenesis of osteoporo-sis: concepts, conflicts, and prospects.J Clin Invest 2005;115:3318-25.

5. Martin TJ, Seeman E. Bone remode-lling: its local regulation and theemergence of bone fragility. BestPract Res Clin Endocrinol Metab2008;22:701-22.

6. Riggs BL, Parfitt AM. Drugs used totreat osteoporosis: the critical needfor a uniform nomenclature based ontheir action on bone remodeling. JBone Miner Res 2005;20:177-84.

7. Baron R. Osteoporosis therapy-dawnof the post-bisphosphonate era. NatRev Endocrinol 2011;8:76-8.

8. Neer RM, Arnaud CD, Zanchetta JR,Prince R, Gaich GA, Reginster JH, etal. Effect of parathyroid hormone (1-34) on fractures and bone mineraldensity in postmenopausal womenwith osteoporosis. N Engl J Med2001;344:1434-41.

9. Zhang L, Yang M, Liu D, Guo C, Li L,Yang G. The rhPTH (1-34), but notelcatonin, increases bone anabolicefficacy in postmenopausal women with osteoporosis.Exp Clin Endocrinol Diabetes 2012;120:361-6.

10. Yu EW, Neer RM, Lee H, Wyland JJ, de la Paz AV, DavisMC, et al. Time-dependent changes in skeletal respon-se to teriparatide: escalating vs. constant dose teripara-tide (PTH 1-34) in osteoporotic women. Bone2011;48:713-9.

11. Dahl SG, Allain P, Marie PJ, Mauras Y, Boivin G,Ammann P. Incorporation and distribution of strontiumin bone. Bone 2001;28:446-53.

12. Marie PJ. The calcium-sensing receptor in bone cells:a potential therapeutic target in osteoporosis. Bone2010;46:571-6.

13. Marie PJ, Hott M, Modrowski D, De Pollak, Guillemain,Deloffre P, et al. An uncoupling agent containing stron-tium prevents bone loss by depressing bone resorptionand maintaining bone formation in estrogen-deficientrats. J Bone Miner Res 1993;8:607-15.

14 Ammann P, Badoud I, Barraud S, Dayer R, Rizzoli R.Strontium ranelate treatment improves trabecular andcortical intrinsic bone tissue quality, a determinant of

bone strength. J Bone Miner Res 2007;22:1419-25.15. Meunier PJ, Roux C, Seeman E, Ortolani JE, Badurski S,

Spector TD, et al. The effects of strontium ranelate onthe risk of vertebral fracture in women with postmeno-pausal osteoporosis. N Engl J Med 2004;350:459-68.

16. Reginster JY, Kaufman JM, Goemaere S, Devogelaer JP,Benhamou CL, Felsenberg D, et al. Maintenance ofantifracture efficacy over 10 years with strontium rane-late in postmenopausal osteoporosis Osteoporos Int2012;23:1115-22.

17. Reginster JY, Seeman E, De Vernejoul MC, Adami S,Compston J, Phenekos C, et al. Strontium ranelatereduces the risk of nonvertebral fractures in postmeno-pausal women with osteoporosis: Treatment ofPeripheral Osteoporosis (TROPOS) study. J ClinEndocrinol Metab 2005;90:2816-22.

18. Bruyere O, Roux C, Detilleux J, Slosman DO, SpectorTD, Fardellone P, et al. Relationship between bonemineral density changes and fracture risk reduction inpatients treated with strontium ranelate. J ClinEndocrinol Metab 2007;92:3076-81.

19. Bruyere O, Collette J, Rizzoli R, Decock C, Ortolani S,

Figure 3. Representation of the most significant biological proces-ses of each phase in which are integrated those genes modulatedby the treatment. The representation is shown from higher to loweraccording to the number of genes which change. All are statisticallysignificant (p<0.001)

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Cormier C, et al. Relationship between 3-month chan-ges in biochemical markers of bone remodelling andchanges in bone mineral density and fracture inciden-ce in patients treated with strontium ranelate for 3years. Osteoporos Int 2010;21:1031-6.

20. Busse B, Jobke B, Hahn M, Priemel M, Niecke M, SeitzS, et al. Effects of strontium ranelate administration onbisphosphonate-altered hydroxyapatite: matrix incor-poration of strontium is accompanied by changes inmineralization and microstructure Acta Biomater2010;6:4513-21.

21 Braux J, Velard F, Guillaume C, Bouthors S, Jallot E,Nedelec JM, et al. A new insight into the dissociatingeffect of strontium on bone resorption and formation.Acta Biomater 2011;7:2593-603.

22. Boivin G, Deloffre P, Perrat B, Panczer G, BoudeulleM, Mauras Y, et al. Strontium distribution and interac-tions with bone mineral in monkey iliac bone afterstrontium salt (S 12911) administration. J Bone MinerRes 1996;11:1302-11.

23. Fernández-Murga Chavanne ML, Noguera R, Rubio E,García Pérez MA, Aliaga R, Cano Sánchez A. Las cas-cadas de señalización Wnt y su implicación en la oste-ogénesis. Rev Osteoporos Metab Miner 2012;4:13.

24. Wang D, Christensen K, Chawala K, Xiao G,Krebsbach P, Franceschi R. Isolation and characteriza-tion of MC3T3-E1 preosteoblast subclones with distinctin vitro and in vivo differentiation/mineralizationpotential. J Bone Miner Res 1999;893-903.

25. Bellows CG, Heersche JNM, Aubin JE. Inorganic phos-

phate added exogenously or released from beta-glyce-rophosphate initiates mineralization of osteoid nodulesin vitro. Bone Miner 1992;17:15-29.

26. Hoemann CD, Gabalaway H, McKee MD. In vitro oste-ogenesis assays: Influence of the primary cell sourceon alkaline phosphate activity and mineralization.Pathol Biol 2009;57:318-23.

27. Quarles, D Yohay DA, Lever LW, Caton R, Wenstrup RJ.Distinct proliferative and differentiated stages of muri-ne MC3T3-E1 cells in culture: an in vitro model of oste-oblast development. J Cell Biol 1992;96:683-92.

28. Fernández-Murga L, Rubio E, Calap E, Aliaga RM,García Pérez MA, Cano Sánchez A. Estudios in vitro desales de estroncio sobre la osteogénesis y su efecto enla vía de señalización Wnt/914 β-catenina. RevOsteoporos Metab Miner 2011;3:9.

29. Beck GR, Zerler B, Moran E. Gene array analysis ofosteoblast differentiation. Cell Growth Differ2001;12:61-83.

30. Raouf A. and Seth A. Discovery of Osteoblast-associatedgenes using cDNA microarrays. Bone 2002;30:463-71.

31. Fromigué O, Haÿ E, Barbara A, Marie PJ. Essential roleof nuclear factor of activated T cells (NFAT)-mediatedWnt signaling in osteoblast differentiation induced bystrontium ranelate. J Biol Chem 2010;285:25251-8.

32. Rybchyn MS, Slater M, Conigrave AD, MasonRS. AnAkt-dependent increase in canonical Wnt signalingand a decrease in sclerostin protein levels are involvedin strontium ranelate-induced osteogenic effects inhuman osteoblasts. J Biol Chem 2011;286:23771-92.


Neila Calvo S, Nan Nan D, García Ibarbia C, Olmos Martínez JM, González Macías J, Hernández Hernández JLUnidad de Metabolismo Óseo - Servicio de Medicina Interna - Hospital Marqués de Valdecilla-IFIMAV - Universidad de Cantabria - RETICEF - Santander

The reality of osteoporosis inpatients hospitalized in InternalMedicine

Correspondence: Sara Neila Calvo - Unidad de Metabolismo Óseo - Departamento de Medicina Interna - HospitalUniversitario Marqués de Valdecilla - Avda. Valdecilla, s/n - 39008 Santander (Spain)e-mail: [email protected]


SummaryPurpose: a) to know the prevalence of previous osteoporosis and vertebral fractures in patients admittedto an Internal Medicine department from a third-level hospital; b) to determinate the proportion ofpatients discharged with a diagnosis of osteoporosis, and the percentage of those receiving treatment; c)to quantify the risk of fracture by applying the FRAX calculation tool; and d) to know the serum levelsof 25OHD in these patients.Patients and methods: Retrospective study, based on the review of clinical charts of all the patients admit-ted to the Internal Medicine department of Marqués de Valdecilla University Hospital, during April 2012.The information was gathered by a standardized protocol, including demographic, clinical, radiologicaland laboratory variables.Results: Three hundred patients were studied (mean age, 80 years). Thirty-four (11.3%) had a previousdiagnosis of osteoporosis and 14 (4.8%) of them were, or had been, on treatment. A diagnosis of osteo-porosis, in the hospital discharge report, was noted in 14 patients. No treatment was prescribed in oneof them. According to the FRAX calculation tool, mean risk for major osteoporotic fracture was 10.5%,and mean risk for hip fracture was 5.4%. Mean serum 25OHD level was 16 ng/ml, and more than 80%of patients had values below 20 ng/ml.Conclusion: Osteoporosis is an underdiagnosed and undertreated disease, in patients admitted to an InternalMedicine Department, whatever the reason. Moreover, we have observed a high prevalence of 25OHD defi-ciency among these patients. Hospitalization can represent an excellent opportunity for the internists andother clinicians, to pay attention to the presence of osteoporosis and its related complications.

Key words: vertebral fracture, osteoporosis, diagnosis, chest-X-ray, 25OHD, Internal Medicine.


IntroductionOsteoporosis is a disease characterised by a reduc-tion in bone mass and by alterations in the micro-architecture of bone tissue, which leads to anincrease in its fragility and, consequently, a higherrisk of suffering fractures1. The prevalence of oste-oporosis increases with age, from 15% in womenbetween 50 and 59 years of age to more than 80%in those over 802.

The principal osteoporotic fractures are verte-bral, hip, wrist, humerus and pelvis. These fractu-res bear significant health and social implications.Furthermore, in a high percentage of patients theylead to significant morbidity, such as loss of mobi-lity, loss of ability to carry out independently basicactivities of daily living, chronic pain and evendepression3,4. Similarly, osteoporotic fractures,above all hip fractures, are associated with a sig-nificant increase in mortality5, this fact is especiallyimportant, since it has been estimated that theannual incidence worldwide of hip fractures inwomen will have increased by 3.5 times between1990 and 20506.

In the last few decades, in hospitals in general,and in general internal medicine services in parti-cular, an increase has been seen in the averageage of patients being hospitalised who, in addi-tion, usually have multiple comorbidities and arepolymedicated. In fact, the prevalence osteoporo-sis, as a disease linked to aging, is increased inthese individuals7. Admission to hospital repre-sents on many occasions an opportunity to diag-nose, initiate treatment and programme appropria-te monitoring in those patients with osteoporosis,with the aim of attempting to prevent the compli-cations of the disease, mainly fractures. We havecarried out this work with the aims of: a) ascertai-ning the prevalence of previous osteoporosis andof vertebral fractures in patients admitted to theinternal medicine service of a tertiary hospital; b)determining the proportion of patients dischargedwith a diagnosis of osteoporosis and the percenta-ge of those who are treated; c) quantifying the riskof fracture by applying the FRAX® tool, and; d) fin-ding out the blood levels of 25-hydroxyvitamin D(25OHD) in these patients.

Material and methodsA retrospective descriptive study was designed,based on a review of the clinical histories of allpatients admitted to the internal medicine serviceof the Marqués de Valdecilla University Hospitalduring the month of April 2012. This centre servesas a health referral centre for a population of350,000 inhabitants of Cantabria. The internalmedicine service has approximately 130 hospitalbeds.

The variable data for the study were gatheredusing a standardised protocol in a computeriseddatabase management system. These variableswere: risk factors for osteoporosis, comorbidities,drugs with an influence on bone metabolism,results of X-rays of thorax and/or dorso-lumbarspine, blood levels of 25OHD and prescribed tre-

atments, differentiating between treatments at timeof hospital admission and at discharge. Also recor-ded was the presence or not of family history ofhip fracture and personal history of fractures.

Menopause was defined as being early if it hadoccurred before the age of 45. It was consideredthat a patient had clinical osteoporosis if they hadsuffered a typical osteoporotic fracture (hip, verte-bral, humerus, forearm) earlier or had a vertebralfracture in the radiological study requested onadmission, in the absence of other causes whichmay have accounted for it (high impact trauma,bone metastasis, myeloma or other bone disea-ses).

The comorbidities analysed were: chronic obs-tructive pulmonary disease –COPD– by means ofa diagnosis based on tests of compatible respira-tory function); ischemic cardiopathy or cardiacinsufficiency, cerebrovascular accident, dementia,Parkinson’s disease; active neoplasia, or in remis-sion; venous thromboembolism; cataracts; rheu-matoid arthritis; diabetes mellitus type 1 or 2(HbA1c ≥6.5% or two tests for glycemia in fasting≥126 mg/dl or a casual glycemia of 200 mg/dl ormore than one patient with cardinal symptoms);arterial hypertension (through measurement andconfirmation of levels of TAS > 140 mmHg and/orTAD >90 mmHg persistently); dyslipemia;hyperthyroidism and hypothyroidism; hyperpa-rathyroidism; hypogonadism; intestinal malab-sorption syndrome; malnutrition, urolithiasis;chronic hepatopathy; chronic renal insufficiency(defined as a glomerular filtrate <60 ml/min/m2

according to the four variable MDRD equation,maintained for at least 3 months). The drugs eva-luated were: corticoids, beta-blockers, anticonvul-sives, statins, proton pump inhibitors (PPIs), sero-tonin re-uptake inhibitors, oral anticoagulants,opiates, hypnotics and benzodiazepines.Treatments for osteoporosis recorded in the dis-charge reports included: denosumab, selectiveestrogen receptor modulators (SERMs), strontiumranelate, teriparatide, PTH 1-84, calcitonin or bis-phosphonates.

Blood levels of 25OHD were determined usingelectrochemiluminescence (Elecsys 2010, RocheDiagnostics, GMBH, Mannheim, Germany).

A vertebral fracture was defined as a reductionin height of the vertebral body greater than orequal to 20%, assessed through a review of lateralX-rays of the thorax and/or thoracic-abdominalspine. The X-rays were reviewed independentlyby the two authors and disagreements (<5%) wereresolved by consensus. All the patients diagnosedwith osteoporosis had had at least one laboratorystudy which included biochemistry, haemogramand VG, thyroid hormone, proteinogram and 24hour calciuria.

Results300 patients were included in the study of whom157 (52.3%) were women and 143 were men. Theaverage age was 80 years. The average body massindex (BMI) was 29.3 kg/m2.


In terms of the risk factors related to osteopo-rosis or fragility fracture, the most frequent wasthe presence of cataracts (76 patients), followedby malnutrition (33), hypothyroidism (26), malab-sorption syndrome (24) and chronic hepatopathy(24). Seven patients had history of hypogonadismand hyperparathyroidism, and it was only possibleto confirm 4 women with history of early meno-pause.

In tables 1 and 2, respectively, are shown thecomorbidities of the patients included in the studyand their consumption of drugs.

A total of 34 subjects (11.3%) had had a pre-vious diagnosis of osteoporosis (5 men and 29women), although only 14 of these (4.8%) werereceiving or had received treatment for the disea-se (zoledronic acid in one patient, other bisphos-phonates in eight patients, three patients werereceiving denosumab and teriparatide and stron-tium ranelate had been subscribed in one caseeach). Of the 34 cases of osteoporosis, in 6 casesthe patients were receiving oral corticoids, in onecase for rheumatoid arthritis, and in five cases withchronic obstructive pulmonary disease (it was notpossible to quantify the exact doses of the gluco-corticoids administered). In one case there washistory of early menopause, and in another long-term hyperthyroidism. Treatment had been pres-cribed in only two of these case.

On the other hand, of the 34 patients withdiagnosis of osteoporosis, in 16 of them it wasconfirmed that they had suffered at least one ver-tebral fracture.

All the patients had an X-ray of the thoraxtaken during their admission and in 31 there wasalso an X-ray taken of the thoracic-lumbar spine.50 patients were identified as having vertebralfractures. Taking into account the total of all thepatients studied and given that only 16 had historyof radiological vertebral fractures, in 12% of allpatients the presence of vertebral fractures hadpassed unnoticed by the clinician responsible. Onthe other hand, 68% of all fractures had passedunnoticed. Of the 50 patients with vertebral frac-tures, in only one case was this attributed to mul-tiple myeloma.

Overall, and in accord with earlier data, 74cases of clinical osteoporosis (defined by verte-bral, hip humerus or wrist fracture) were identi-fied. Therefore, 40 subjects had not had a diagno-sis of osteoporosis. At the time of discharge, inonly 14 patients was the diagnosis of osteoporosisconfirmed in the clinical record, and of these, inone case no treatment was programmed.

On the other hand, the average risk of majorosteoporotic fracture (vertebral, hip, humerus orwrist) measured by application of the FRAX® scalewas 10.5% (SD: 8.7) and the risk of hip fracture5.4% (SD: 5.5). According to the FRAX® tool deve-loped for Spain, there were 27 patients with a riskhigher than 20% of major osteoporotic fracture,and 118 patients had a risk higher than 3% forfracture of the hip. The risk of fracture using theFRAX® tool could only be calculated in 174

patients, due to a lack of information in the clini-cal histories of the other subjects of the study,while those treated for osteoporosis were exclu-ded.

The average blood value of 25OHD was 16ng/ml, although this was only obtained from 45patients, this being a test which was not includedin the protocol of requests for laboratory tests onadmission. More than 80% of the patients hadvalues below 20 ng/ml, and more than 50% (26patients) had serious hypovitaminosis (≤10ng/ml). Only two patients with hypovitaminosis Ddid not receive oral supplements at discharge.

DiscussionOsteoporosis is a process which generally deve-lops asymptomatically over a long period of time,a fracture being the first sign in the majority ofcases. A predisposition to the development offractures is, from a clinical point of view, the cen-tral phenomenon of the disease. Among thesefractures, of greatest significance is fracture of thehip, which preferentially affects older people, isfrequent in women and which varies greatly in itsincidence, its mortality and in length of stay inhospital8.

Vertebral fractures are another complicationcharacteristic of osteoporosis, although more than50% of them pass unnoticed9,10. It is important toremember that the presence of an osteoporoticfracture, independently of the densitometry value,increases even more the risk of subsequent fractu-res, which means that the diagnosis is as impor-tant as the monitoring of the disease. In a studypublished by Sosa et al.11, it was concluded thatthe presence of vertebral fractures increased therisk of new fractures. Furthermore, the authorsobserved this type of fracture in 62.6% of patientshospitalised or treated for fracture of the hip.Although there is no universally accepted defini-tion of vertebral fracture, the majority of authorsare in agreement in considering that there must bea loss of height of the vertebral body of at least20%12. We know that lateral X-rays of the thoraxare a very useful tool in enabling the identifica-tion of vertebral fractures9. In our work 50 patientswere identified with fractures of this type, mostlyunnoticed by the clinician (in our service, thoracicX-rays are not analysed by a radiologist unlessspecifically requested) and as a consequence, arenot reflected in the discharge notes.

On the other hand, according to the definitionof clinical osteoporosis which we have used, 74cases were identified, which shows that almost50% were not recorded in the clinical record(there being 34 patients with a previous diagnosisof osteoporosis). Furthermore, only 13 patients inwhom their discharge notes reflected a diagnosisof osteoporosis received treatment, which indica-tes once more that, although osteoporosis is ahighly prevalent chronic disease, it continues tobe underdiagnosed and undertreated.

In respect of the results obtained using theFRAX® tool, it is known that the Spanish version of


FRAX® underestimates by 50% the number ofmajor fractures13. So, we could take into accountthe recommendations of NOF, which is to treatpatients if their risk of major fracture is ≥20% andof hip fracture ≥3%. Thus in our work, more than30% of patients (for major fracture) and more than80% (for hip fracture) had indications to receivetreatment, although it should be borne in mindthat this guide recommends starting treatmentwhen there is a diagnosis of osteoporosis, whate-ver the FRAX® indicates. This makes us think thatpatients admitted to the internal medicine serviceusually have a high risk of osteoporotic fracture.

In terms of the determination of the level of25OHD in the blood, it seems that, while this isnot normally requested routinely during hospitaladmission, the staff do usually prescribe treatmentwhen there is a deficit.

When dealing with a multifactorial disease inwhich are involved, among others, genetic andenvironmental factors, one of the keys to diagno-sis is to identify its risk factors2. Hospitalisationmay be an opportunity to diagnose osteoporosisand vertebral fracture. However, in line with thedata provided in this work the risk factors for frac-ture are not generally evaluated during admissionand osteoporosis continues to be forgotten by theclinician despite its known epidemiological, healthand social significance. Furthermore, it is a highlyprevalent disease in older patients, in whom, withincreasing frequency, are associated malnutrition,comorbidities and polymedication (a notable fin-ding in this work is the significant consumption ofhypnotics) which themselves pose additional riskof falls and, therefore, of fracture2.

However, there are highly efficacious and easymethods of treatment for osteoporosis, especiallyin polymedicated patients, which ensure thera-peutic compliance, and which may be considered,when indicated, during admission to hospital14.

Our study has various limitations. Firstly, thoseinherent in any retrospective study. In addition,not all the patients had X-rays of the lumbar spine,which means that it was not possible to determi-ne the true prevalence of vertebral fractures in thissection of the spine. Finally, the determination of25OHD in the blood was carried out in a smallsub-group of hospitalised patients, which meansthat we cannot generalise the results to all thepatients studied. However, our work group hasstudied levels of 25OHD in a broad sample ofpatients hospitalised in our internal medicine ser-vice (approximately 400 individuals) and theresults were similar (data not published).

In conclusion, osteoporosis is a disease rarelyconsidered by the clinician in patients hospitaliseddue to other causes, in spite of its having a preva-lence similar to other diseases such as diabetesmellitus, dyslipemia or dementia. As a consequen-ce, it continues to be a condition which is under-diagnosed and secondarily undertreated, in spiteof there being many therapeutic options. In thelight of the findings of our work we suggest thatthe internist, and in general, any clinician, should

Table 1. Comorbidities in the patients in the study

Disease N %

Hypertension 200 67

Dementia 112 37

Heart failure 111 37

Dyslipidemia 93 31

Diabetes mellitus 91 30

COPD 78 26

Ictus 72 24

Ischemic heart disease 68 23

Neoplasia 67 22

Chronic renal failure 60 20

Alcoholism 35 12

Thromboembolic disease 34 11

Smoking 28 9

Parkinson's disease 17 6

COPD: chronic obstructive pulmonary disease

Table 2. Consumption of drugs in hospitalizedpatients

Drug N %

PPIs 154 51

Hypnotics 126 42

Statins 90 30

Anticoagulants 56 19

Βeta-blockers 36 12

Opiates 33 11

Thyroid hormone 23 8

Corticosteroids 18 6

Anticonvulsants 9 3

PPIs: proton pump inhibitors


maintain a high level of suspicion of this diseasein hospitalised patients, in order to try to reduceassociated complications, especially fractures.

Bibliography

1. Díaz Curiel M, García JJ, Carrasco JL, Honorato J, PérezCano R, Rapado A, et al. Prevalencia de osteoporosisdeterminada por densitometría en la población feme-nina española. Med Clin (Barc) 2001;116:86-8.

2. Rosen CJ. Postmenopausal osteoporosis. N Eng J Med2005;353:595-603.

3. Ioannidis G, Papaioannou A, Hopman WM, Akhtar-Danesh N, Anastassiades T, Pickard L, et al. Relationbetween fractures and mortality: results from theCanadian Multicentre Osteoporosis Study. CMAJ2009;181:265-71.

4. Poole KE, Compston JE. Osteoporosis and its manage-ment. BMJ 2006;333:1251-6.

5. Cauley JA, Thompson DE, Ensrud KC, Scott JC, BlackD. Risk of mortality following clinical fractures.Osteoporos Int 2000;11:556-61.

6. Gullberg B, Johnell O, Kanis JA. World-wide projec-tions for hip fracture. Osteoporosis Int 1997;7:407-13.

7. Cummings SR, Melton J. Epidemiology and outcome of

osteoporotic fractures. Lancet 2002;359:1761-7.8. Serra JA, Garrido G, Vidán M, Marañón E, Brañas F,

Ortiz J. Epidemiología de la fractura de cadera enancianos en España. An Med Interna (Madrid)2002;19:389-95.

9. Hernández JL, Hidalgo I, López-Calderón M, OlmosJM, González Macías J. Diagnóstico de osteoporosismediante radiografía lateral de tórax. Med Clin (Barc)2001;117:734-6.

10. Becker C. Pathophysiology and clinical manifestationsof osteoporosis. Clin Cornerstone 2006;8:19-27.

11. Sosa M, Saavedra P. Prevalencia de fracturas vertebra-les en pacientes con fractura de cadera. Rev Clin Esp2007;207:464-8.

12. Majumdar SR, Kim N, Colman I, Chahal AM, RaymondG, Jen H, et al. Incidental vertebral fractures discove-red with chest radiography in the emergency depart-ment: prevalence, recognition, and osteoporosis mana-gement in a cohort of elderly patients. Arch Intern Med2005;165:905-9.

13. González Macías J, Marin F, Vila J, Díez Pérez A.Probability of fractures predicted by FRAX® and obser-ved incidence in the Spanish ECOSAP Study cohort.Bone 2012;50:373-7.

14. Lyles KW, Colón-Emeric CS, Magaziner JS, Adachi JD,Pieper CF, Mautalen C, et al. Zoledronic acid and cli-nical fractures and mortality after hip fracture. N EnglJ Med 2007;18:1799-809.

CLINICAL NOTE / Rev Osteoporos Metab Miner 2013 5;4:147-149147

Herrero Vicent C1, Pascual Pla FJ2, Samper Hiraldo JM3, Gavilá Gregori J1

1 Servicio de Oncología Médica2 Servicio de Urgencias Médicas3 Oncología RadioterápicaFundación Instituto Valenciano de Oncología - Valencia

Multiple skeletal-related events in apatient with breast cancer

Correspondence: Carmen Herrero Vicent - Servicio de Oncología Médica - Fundación Instituto Valenciano deOncología - Profesor Beltrán Báguena, 8 - 46009 Valencia (Spain)e-mail: [email protected]


SummaryBone metastases are common in advanced cancer, occurring up to 75% of patients with advanced breastcancer. Complications of bone metastases include bone pain, hypercalcemia and skeletal-related events(SERs), such as fracture, need for radiation or surgery to bone, or spinal cord compression. A 50 year-old patient with advanced breast cancer, who has multiple skeletal-related events and pooreroverall quality of life.

Key words: bone metastases, skeletal-related events, cancer, breast.


Introduction One of the most aggressive characteristics of cancer in general isits capacity to produce metastases, with the skeleton being one ofthe most common locations for their development. The activity ofosteoclasts produces a local destruction of the bone and, as a con-sequence, skeletal-related events (SREs) such as pathological frac-tures and medullary compressions which may require treatmentwith radiotherapy or surgery, with consequent hospitalisation1.

Presented here is the case of a patient with advanced bre-ast cancer with multiple SREs with affectation of her quality oflife, and which required an efficacious treatment based on amultidisciplinary approach.

Clinical caseA patient of 50 years of age diagnosed in December 2002 withinfiltrating ductal carcinoma of the left breast, hormonalreceptor-positive with the clinical state T3 N0 M0, treated withinduction chemotherapy with 4 cycles of fluorouracil-epirubi-cin-cyclophosphamide, modified radical mastectomy, 4 cyclesof docetaxel and adjuvant radiotherapy in the costal wall, leftsupraclavicular fossa and internal mammary nodes up to 50Gy. This was followed by hormone therapy with aromataseinhibitors and periodic checks.

In April 2005 the patient reported mechanical lumbalgiawhich did not recede with WHO first or second level analge-sia. A magnetic resonance (MR) scan was carried out in thespine, which found a single metastasis in L4, which was con-firmed histologically. An intervention was performed with afixation followed by radiotherapy on L3-L5 receiving 30 Gy,and chemotherapy initiated with vinorelbine associated withzoledronic acid 4 mg intravenous every 28 days over 2 years(24 doses in total) followed by periodic checks.

In February 2013 the patient complained of pain of a mecha-nical nature in the upper left limb which developed over a num-ber of hours. A simple radiography of the left humerus was per-formed which showed a fracture of the inferior third of thediaphysis, which was fixed with an intramedullary nail, along withradiotherapy (Figure 1a). The biopsy was compatible with metas-tasis originating from the breast. In the axial thoracic-abdominal-pelvic computerised tomography (CT) scan there was evidence ofprogression in the bone, confirmed by bone tracking (BT).

During admission the patient reported pain in the glenohu-meral joint, and after the carrying out of an X-ray a fracture ofthe right acromion was observed which required fixing.

In April 2013 the patient presented with difficulty walkingand acute mechanical pain in the proximal part of the left legthat was not eased with WHO level 3 analgesia. In the X-ray afracture of the diaphysis of the left femur was found, on whichan intramedullary fixation was carried out (Figure 1b).

During admission she presented with pain in the upperdorsal zone with a positive spinal apophysis percussion test,due to which a CT scan was requested, which showed a bulgein the posterior wall of D3 and D7, which was confirmedthrough MR (Figure 2). After evaluation for radiotherapy it wasdecided to administer 20 Gy to D2-D3 and D7, and 20 Gy tothe left femur. At the time of producing this work (June 2013)the patient was in treatment with capecitabine, despite presen-ting a score of 3 on the Performance Status scale, secondary tothe multiple skeletal-related events which resulted in depen-dency for the activities of daily living.

DiscussionPhysiologically, there is a balance between bone formationand resorption. The osteoblast line cells are involved in oste-

Figure 1. (a) X-rays of left humerus, inwhich are observed a fracture in themiddle third and the fixation with anintramedullary nail. (b) X-rays of leftfemur with lytic metastasis and intra-medullary nail

a

b

Figure 2. Magnetic resonance of thespine, which evidences medullarycompression at the second, third andseventh dorsal vertebrae


oblast function and differentiation by means of theRANK factor (Ligand Receptor of the Activator ofNuclear Kappa-B factor) present in their membra-ne. The bonding of RANK to its ligand (RANKL)stimulates differentiation, the survival of the oste-oclast precursor cells and the activity of the matu-re osteoclasts, thus increasing the expansion of theosteoclast mass and bone resorption2.

On the other hand, osteoprotegin (OPG) hasbeen identified as a protein which inhibits thedevelopment of osteoclasts. When there is suffi-cient OPG in the environment this protein bondswith the RANKL of the osteoclasts, impeding theirinteraction with the osteoclast precursors, whichslows the process of bone resorption. Changes inthe RANKL/OPG quotient are decisive in thepathogeny of bone loss, from osteoporosis tobone metastasis.

Bone is a target tissue for metastasis in breastcancer. This is due, in part, to the irrigation of thebone itself, and to growth factors IGF-1, FGF andPDGF, which exert an attraction on cancerouscells and are a suitable medium for cell growth3.

The bone balance is altered by the arrival ofthe tumorous cells. It is necessary that the neo-plastic cells are retained in the sinusoids of thebone medulla, that they migrate, that they passthrough the vascular wall and adhere to the extra-cellular matrix of the bone surface of the perios-teum in order to be able to stimulate the osteo-blasts and osteoclasts. In breast cancer there maybe an increase in bone resorption over formation,favouring the formation of osteolytic bone metas-tases. This imbalance is related to an increase inmarkers for bone resorption such as urinary N-ter-minal telopeptide of collagen type 1 (uNTX), C-terminal telopeptide of collagen type 1 (CTX), thealkaline phosphatases, (AFs), the amino-terminalpropeptide of procollagen type 1 (PINP) or tartra-te-resistant acid phosphatase (TRAP-5b)4.

Clinical trials have shown that the bisphospho-nates inhibit the resorption of bone mediated bythe osteoclasts, which means that it could be a the-rapeutic option for the prevention of bone lossinduced by oncological treatment secondary to hor-monal deprivation, especially if the patient has alow bone mineral density or has risk factors for thedevelopment of fractures from minimal traumas. Inthe initial stages of non-metastatic breast cancer,zoledronic acid administered every 6 weeks, oralibandronate monthly and weekly risedronate havebeen shown to prevent bone loss associated withthe use of aromatase inhibitors in breast cancer inpostmenopausal women. On the other hand, SREsoccur in 64% of patients with breast cancer who arenot treated with bisphosphonates, which is why theAmerican Society for Clinical Oncology (ASCO)recommends treatment with intravenous bisphos-phonates in patients with pain or destruction ofbone evidenced by radiography5-7.

Other, more recent treatments are those mono-clonal antibodies such as denosumab, which havea great affinity with RANKL, impeding theRANKL/RANK interaction on the surface of the

osteoclasts, thus diminishing bone resorption. In aphase II clinical trial carried out in patients withbreast cancer and bone metastasis in which werecompared 120 mg subcutaneous denosumab, plusan intravenous placebo, and 4 mg intravenouszoledronic acid, adjusted for renal function, plusa subcutaneous placebo every 4 weeks, it wasshown that denosumab reduced the time until thefirst bone event by up to 23% (HR 0.82; CI 95%,0.71-0.95; p=0.1 superiority) and the risk of multi-ple events by up to 18% (HR 0.77; CI 95%, 0.66-0.89; p=0.001), compared with the zoledronic acid[8]. These data translate into a greater quality oflife for the patient9,10. In addition, unlike zoledro-nic acid, denosumab is not nephrotoxic, whichmeans that there is no requirement to adjust accor-ding to creatinine clearance, it is administeredsubcutaneously and has lower toxicity.

The better understanding of bone metabolism,advances in molecular biology and a better cha-racterisation of the signaling systems of theRANK/RANKL/OPG pathways represent an advan-ce in the treatment of bone metastases, as well asin the prevention of states of osteopenia and oste-oporosis secondary to oncology treatment, giventhat these negatively influence the morbimortalityof our patients.

Bibliography

1. Coleman R. Potential use of bisphosphonates in theprevention of metastases in early stage breast cancer.Clin Breast Cancer 2007;7:S29-35.

2. Roodman GD. Mechanism of bone metastasis. N EnglJ Med 2004;350:1655-64.

3. Mundy GR, Chen D, Zhao M, Dallas S, Xu C, Harris S.Growth regulatory factors and bone. Rev EndrocrMetab Disord 2001;2:105-15.

4. Demers LM, Costa L, Lipton A. Biochemical markersand skeletal metastases. Cancer 2000;88:2010-26.

5. Hillner BE, Ingle JN, Chlebowski RT, Gralow J, YeeGC, Janjan NC, et al. American Society of ClinicalOncology 2003 update on the role of bone healthissues in women with breast cancer. J Clin Oncol2003;21:4042-57.

6. Lipton A, Theriault RL, Hortobagy GN, Simeone J,Knight RD, Mellars K, et al. Pamidronate prevents ske-letal complications and is effective palliative treatmentin women with breast carcinoma and osteolytic bonemetastases: Long term follow-up of two randomized,placebo-controlled trials. Cancer 2000;88:1082-90.

7. Gálvez-Muñoz E, Rodríguez-Lescure A. Papel de losbifosfonatos en el tratamiento adyuvante del cáncer demama. Med Clin 2010;135:70-4.

8. Stopeck A, Lipton A, Body J, Steger G, Tonkin K, deBoer R, et al. Denosumab compared with zoledronicacid for the treatment of bone metastases in patientswith advanced breast cancer: a randomized, double-blind study. J Clin Oncol 2010;28:5132-9.

9. Martín M, Bell R, Bourgeois H, Brufsky A, Diel I, EniuA, et al. Bone-related complications and quality of lifein advanced breast cancer: Results from RandomizedPhase III Trial of Denosumab versus Zoledronic Acid.Clin Cancer Res 2012;18:4841-9.

10. Von Moos R, Body JJ, Egerdie B, Stopeck A, Brown JE,Damyanov D, et al. Pain and health-related quality oflife in patients with advanced solid tumours and bonemetastases: integrated results from three randomized,double-blind studies of denosumab and zoledronicacid. Support Care Cancer 2013:21:3497-507.

SPECIAL ARTICLE / Rev Osteoporos Metab Miner 2013 5;4:151-157151

Roig Escofet D, García Borrás JWith the collaboration of: del Pino Montes J, Sosa Henríquez M, González Macías J, Moro Álvarez MJ, Herrero L, CalvoCatalá J, Giménez Úbeda E, Quesada Gómez M, Díaz Curiel M, Lluch Mesquida P, Gómez Alonso C, Olmos Martínez JM,Torrijos Eslava A, Guañabens Gay N, Nogués Solán X

The history of SEIOMM (1987-2013)

During the 1980s, the medical professionals asso-ciated with diseases of the locomotor apparatusbegan to pay attention to metabolic bone diseases,and among these, a pathology very common inadvanced age, osteoporosis, which, until this timehad passed almost unnoticed due to the absenceof precise methods of diagnosis. Bone fractures, toa great extent brought on by osteoporosis, werefrequent and provoked disorders and disabilities,above all in women after the menopause. Theappearance of highly precise methods for thediagnosis of osteoporosis and of efficacious drugsfor its treatment led to an increase in interest inthis pathology on the part of medical specialistsdirectly or indirectly associated with the locomo-tor apparatus such as internists, rheumatologists,endocrinologists, gynaecologists and nephrolo-gists. In Spain this situation resulted in the forma-tion of the Spanish Society for Bone and MineralMetabolism Research (SEIOMM) and led to a phar-maceutical company offering DXAdensitometry at 14 Spanish hospitals,facilitating the diagnosis and study ofosteoporosis.

A management board presidedover by Horacio Rico Lenza, and for-med of Luisa Traba (Vice President),Sergio Serrano (Secretary) and EmmaRosa Hernández (Treasurer) establis-hed the basis of the constitution ofthis multidisciplinary organisationand organised the first symposium inBarcelona on the 6th December1987, which saw a large number ofparticipants. This symposium wasthe opportunity to hold theAssembly at which was appointedSEIOMM’s first board of directors.

Earlier, in April 1987, the EuropeanFoundation for Osteoporosis andBone Diseases had been formed, withthe main objective of promoting theunderstanding of the physiopatho-logy, diagnosis, treatment and pre-

vention of osteoporosis and other bone diseases.To achieve this it was considered necessary tosupport biomedical, clinical, nutritional and epide-miological research into these diseases and to pro-mote their understanding, establishing educationand training for doctors and other health profes-sionals in this field. The same activities were takenon by SEIOMM and have been carried out, as faras possible, since the Society’s foundation.SEIOMM is now a benchmark at an internationallevel.

From December 1987 to December 1991During this period the board of directors wasmade up of Daniel Roig Escofet (President),Ramon Peréz Cano (Vice President), María LuisaMariñoso Barba (Secretary) and María TeresaGonzaléz Álvarez (Treasurer). In addition, boardmembers were appointed, each representing anAutonomous Community, whose role was to act

as a link between the mem-bers in their respectiveCommunities and the board ofdirectors.

One of the first activities ofthe board of directors was toappoint a committee whichwould study the previous arti-cles of association of theSociety and propose modifica-tions which they considerednecessary. In accordance withthis, new articles were draftedwhich were submitted forapproval by the ExtraordinaryAssembly held in Seville on3rd May 1989.

These articles had 35 sec-tions, from which some speci-fics may be highlighted. Themain objective of SEIOMMwas the promotion of basicand clinical research into oste-oporosis and other bone dise-

A MANAGEMENT BOARD PRE-SIDED OVER BY HORACIO RICOLENZA, AND FORMED OF LUISATRABA (VICE PRESIDENT),SERGIO SERRANO (SECRETARY)AND EMMA ROSA HERNÁNDEZ(TREASURER) ESTABLISHED THEBASIS OF THE CONSTITUTION OFTHIS MULTIDISCIPLINARY ORGA-NISATION AND ORGANISED THEFIRST SYMPOSIUM INBARCELONA ON THE 6THDECEMBER 1987, WHICH SAWA LARGE NUMBER OF PARTICI-PANTS. THIS SYMPOSIUM WASTHE OPPORTUNITY TO HOLDTHE ASSEMBLY AT WHICH WASAPPOINTED SEIOMM’S FIRSTBOARD OF DIRECTORS


ases. The national symposium of the Societywould be held biennially, and a MonographMeeting would be held on a current theme in theinter-symposium years. To each of these activitieswould be added a meeting of the GeneralAssembly. Extraordinary Assemblies could becalled for the modification of the articles of asso-ciation or to deal with matters of vital importanceand whose resolution could not wait for the nextordinary General Assembly. The board of directorswould be made up of a President, Vice President,Secretary, Treasurer and a representative of eachAutonomous Community; but a permanent execu-tive board was also established with four repre-sentatives from the Autonomous Communities.This would have decision-making powers. Theboard of directors would produce the agenda forthe General Assembly of SEIOMM, which wouldcoincide with the national symposium or themonograph meeting. The scientific programme ofthe symposia would be produced by the organi-sing committee and would need to be approvedby the board of directors of the Society. It was alsodetermined that the President would be able tohold office for a maximum of 4 years. These arti-cles were approved by an absolute majority of theAssembly.

It was considered that SEIOMM should esta-blish and maintain contacts with other institutionsor societies with similar objectives, both nationallyand abroad. The first international contact wasmade with the European Foundation forOsteoporosis and Bone Disease. This occurred ata meeting held in Davos (Switzerland) in April1988, which was attended by representatives ofdifferent national societies in Europe with similarobjectives. The President of SEIOMM Daniel RoigEscofet attended as the Society’s representative.Another Spanish representative, Aurelio RapadoErrazti, President of another Spanish association,the Spanish Association for Osteoporosis andMetabolic Bone Diseases (AHOEMO) also atten-ded. This association had been established in 1988as a federation of a number of Spanish scientificsocieties. Its objective was to inform the generalpublic on the prevalence and serious effects ofosteoporosis, as well as to give information to thepress and other communications media, and spon-sor research into all aspects of the disease throughgrants, prizes and large population studies. Its acti-vities were social in character, whereas those ofSEIOMM were exclusively scientific. Subsequentlya number of activities were carried out jointly.

The relationship with the EuropeanFoundation resulted in SEIOMM organising an“International Symposium on bone metabolicdiseases” in the Platja d’Aro (Girona) in October1990, with the aim of making the Society betterknown externally. Attendance was high and spe-cialists in these diseases attended from variousEuropean countries, as well as from SouthAmerica, since SEIOMM had also established rela-tionships with the Iberoamerican Society ofOsteology and Mineral Metabolism (SIBOMM), of

which Dr Díaz Curiel was secretary and in whichSEIOMM had a representative.

At the Assembly, Professors Horacio RicoLenza and Miguel Garrido Peralta were madehonorary members.

During this period SEIOMM organised, in addi-tion to the aforementioned international congress,a symposium in Seville (1989) and another inOviedo (1991).

From November 1991 to October 1995At the III Symposium held in Oviedo in November1991 a new board of directors was appointed,now being formed by Jorge Cannata Andía(President), Concepción de la Piedra Gordo (VicePresident), José Bernadino Díaz López (Secretary)and Javier del Pino Montes (Treasurer).

One of the details from this period is the parti-cipation of SEIOMM in the development of theSpanish Review of Bone Metabolic Diseases as theorgan of the Society, along with AHOEMO and theOsteoporosis Working Group of the SpanishSociety of Internal Medicine, and the appointmentof Jorge Cannata Andía as joint editor representingSEIOMM. SEIOMM continued to maintain its exter-nal relationships, and in May 1992 an InternationalSymposium on Osteoporosis was held inBarcelona attended by representatives of variousinternational societies. Dr Concepción de la PiedraGordo was appointed as a member of theSIBOMM board, representing SEIOMM.

The introduction of densitometry in a numberof Spanish hospitals facilitated the development ofwork relating to osteoporosis. Two types of rese-arch were requested of those receiving of the den-sitometers. One of these was a free choice. Theother was common to all the groups and consis-ted of the measurement of the bone density ofgroups of people of different ages, with the aim ofgetting an idea of the normal values for bone massin our country. This latter work, originally concei-ved of as having a transverse design, continued ina second phase with a longitudinal design inwhich the development of bone mass monitoredover three years was studied.

In 1992, as a result of this work the “Study ofbone density in the Spanish population. A multi-centric osteoporosis research project” was publis-hed, with the participation of the following hospi-tals: the La Paz Hospital (Madrid), the San CarlosUniversity Hospital (Madrid), the Jiménez DíazFoundation (Madrid), the Clinical UniversityHospital (Salamanca),the Insular Hospital (LasPalmas de Gran Canaria), the Prínceps d’EspanyaHospital (Hospitalet), the Santa Creu i Sant PauHospital (Barcelona), the Esperanca Hospital(Barcelona), the La Fe Hospital (Valencia), theVirgen Macarena University Hospital (Seville), theUniversity Hospital (Granada), the GeneralHospital of Asturias (Oviedo), the Miguel ServetHospital (Zaragoza) and the University Clinic(Pamplona). Also involved, as well as SEIOMM,were the Osteoporosis Working Group of theSpanish Society for Internal Medicine (GTO),

152


AHOEMO and the medical department of thepharmaceutical company Rhóne Poulenc Rorer.

The results of the study were published invarious reviews and in the Spanish Review ofMetabolic Bone Diseases. Notable among thesewas the article “Normal values for bone mineraldensity in the Spanish adult population”. And in1996, another work on this subject: “New frontiersin the study of bone density in the Spanish popu-lation” was published.

In 1993 SEIOMM awarded prizes to works on“Vitamin D metabolites and osteoporosis”. Thefirst prize was shared between two studies:“Osteoporotic hip fractures in old people. A socialproblem of physiopathology and prevention” byJ.M. Quesada and J. Alonso, and “Levels of25(OH)D in postmenopausalwomen and old people: itsrelationship with bone den-sity” by Maria E. Martínez,M.T. del Campo, M.J Sanchéz-Cabezudo, J.A. García, J.Coya, M.T. Sánchez-Calvín, A.Torijos and L. Munuera. Inaddition, three second prizeswere awarded.

During this presidency theAHOEMO was transformed.In 1993 it changed its nameto the Hispanic Foundation for Osteoporosis andMetabolic Bone Diseases (Fundación Hispana deOsteoporosis y Enfermedades Metabólicas Óseas[FHOEMO] ), with a scientific committee formedby four members of SEIOMM (Jorge CannataAndía, Bernadino Díaz López, Jesús GonzálezMacías and Daniel Roig Escofet) and four othermembers of different scientific societies previouslyassociated with AHOEMO. The desirability ofdeveloping a protocol or document defining therelationship between SEIOMM and FHOEMO inorder to facilitate bilateral collaboration was raised

At the executive board meeting of 19th May1993 it was decided to name Drs Aurelio RapadoErrazti and Daniel Roig Escofet as honorary mem-bers for their outstanding work and effort in acti-vities related to bone and mineral metabolism.

In this period SEIOMM symposia were held inCórdoba (1993) and Alicante (1995) and twomonograph meetings, which the articles of asso-ciation had established, were organised, one on“Glucocorticoids and bone” in Zaragoza (1994)and the other on “Paget’s Disease” in Salamanca.

From October 1995 to October 1999A new board of directors, whose appointmentcoincided with the Alicante symposium, was for-med by Jesús González Macías (President), AdolfoDíez Pérez (Vice President), Manuel SosaHenríquez (Secretary) and Manuel Díaz Curiel(Treasurer).

In this period the normal activities of theSociety continued, predominantly the dissemina-tion of new developments in bone diseasesthrough courses, conferences and written mate-

rials, relationships with other organisations, andthe promotion of research. A letter was sent to allhospital directors general to explain the need tohighlight the significance of bone densitometry.Notable among other activities was a course ondensitometry organised by the SEIOMM and ledby Luís del Río Barquero, in Barcelona inNovember 1997. Also, a monograph meeting orga-nised jointly by the IOF (InternationalOsteoporosis Foundation), FHOEMO andSEIOMM held in Madrid and presided over by DrAntonio Torrijos Eslava on “the Menopause”.

Relationships with overseas societies relatedwith osteoporosis continued. As a minimum, arepresentative of SEIOMM went to Paris every sixmonths to attend meetings with the European

Foundation for Osteoporosis and Bone Disease(EFFO), which transformed during this period intothe International Osteoporosis Foundation (IOF),although there was no Spanish member appointedto its scientific committee. At the III IberoamericanCongress on Osteology and Mineral Metabolismorganised in Mexico in 1996 by SIBOMM, DrManuel Díaz Curiel was appointed as the SEIOMMdelegate to that Society. In addition, Dr AdolfoDíez Pérez became the representative of SEIOMMto the European Calcified Tissue Society, and waschosen as a permanent member of the council ofthat society. There were also conversations withProfessor Gennari, President of the InternationalFederation of Societies on Skeletal Diseases(IFSSD) and with the World Federation onOsteoporosis (WFO) to try to integrate SEIOMMwith these societies on condition that the Societywould be able to play an active part, and be repre-sented on the boards of directors or the executivecouncils of these societies. These conversationswere not fruitful, but in the end these societieswere integrated into EFFO, subsequently calledthe IOF, and disappeared.

At the General Assembly of SEIOMM held inMadrid on 16th October 1998, a modification ofthe statues was approved which created workinggroups with the aim of channelling the scientificactivities of the members into one or other line ofresearch according to their interests. At the end ofthis period there were already four functioningworking groups, dedicated to “bone densito-metry”, “ quality of life”, “management of clinicalhistory” and “research into ultrasound and bonemetabolism”, coordinated respectively by Luis del

THE INTRODUCTION OF DENSITOMETRY IN A NUMBER OF SPANISH HOSPITALSFACILITATED THE DEVELOPMENT OF WORK RELATING TO OSTEOPOROSIS. TWOTYPES OF RESEARCH WERE REQUESTED OF THOSE RECEIVING OF THE DENSITOME-TERS. ONE OF THESE WAS A FREE CHOICE. THE OTHER WAS COMMON TO ALL THEGROUPS AND CONSISTED OF THE MEASUREMENT OF THE BONE DENSITY OFGROUPS OF PEOPLE OF DIFFERENT AGES, WITH THE AIM OF GETTING AN IDEA OFTHE NORMAL VALUES FOR BONE MASS IN OUR COUNTRY


Río Barquero, Adolfo Díez Pérez and the last twoby Manuel Sosa Henríquez. Over time othergroups were added such as one on “risk factors”coordinated by Carmen Valdés Llorca.

The board of directors carried out an internalaudit aimed at understanding the state of theSociety for the administration. A detailed study wasthen proceeded with whose results were discussedat the meeting of the permanent executive boardwhich took place in Santander on 16th February1998. The following actions were carried out:

a) Regularisation of SEIOMM through its regis-tration with the Register of Associations and com-pliance with the corresponding employment laws.

b) Regularisation of the situation of the secre-tary, proceeding with their contract and registeringwith the Social Security department.

c) Contracting with consultancies specialisingin human resources, finance and accountancy sothat monthly accounts could be managed, andannual accounts and a budget for the followingyear prepared.

In this period another honorary member wasappointed: Dr Horacio Rico Lenza. SEIOMM wasalso modernised: an e-mail was set up for the ser-vices of members to enable communication withthe board of directors and an internet web pagewas established for SEIOMM, developed by XavierNogués.

And there was a change: the biennial scientificmeetings until then called symposia became con-gresses. The congresses were held in Granada(1997) and Sitges (1999) and the two monographicmeetings in Madrid, on on “Biochemical markersfor bone remodelling” and the other on“Densitometry”. The latter was timed to coincidewith a joint meeting with FHOEMO and EFFO.

From October 1999 to November 2003A new board of directors was chosen: Adolfo DíezPérez (President), José Manuel Quesada Gómez(Vice President), Nuria Guañabens Gay(Treasurer) and Xavier Nogués Solán (Secretary).In a motion to the General Assembly of SEIOMMheld on 5th October 2002, the Society was notifiedthat the President, Adolfo Díez Pérez had resigneddue to his new position in a multinational com-pany based in the U.S, and the presidency passedto the Vice President, José Manuel QuesadaGómez.

This period saw the death of two significantpersonalities in the field of bone diseases, andhonorary members of SEIOMM: Dr Horacio RicoLenzo, President of the first management board ofSEIOMM, and Dr Aurelio Rapado Errazti, Presidentof FHOEMO. The latter position passed to DrManuel Díaz Curiel.

Relations with national and overseas bodieswere maintained, notably, an accord with theInternational Bone and Mineral Society (IBMS) bywhich for three years from January 2001 membersof SEIOMM also had full rights to the IBMS, recei-ving the review Bone free. In addition, Dr ManuelDíaz Curiel was appointed as President of

SIBOMM. SEIOMM participated in a congress ofthe Spanish Society of Internal Medicine and inanother of the Spanish Society of Family andCommunity Medicine. The relationship with FHO-EMO was maintained, with each Society reflectingits own chatacter: scientific exclusivity forSEIOMM and social exclusivity for FHOEMO.SEIOMM as generator of conferences, scientificmeetings and other events aimed at professionals,and FHOEMO spreading knowledge of the disea-ses in the population through social and culturalactivities.

The working groups already established weresustained. In 1999 a SEIOMM working group onprotocols and clinical history was created, coordi-nated by Manuel Sosa Henríquez. The result ofthis work was SEIOMM’s Clinical History, whosefirst version was presented at the congress held inMenorca in 2001 and subsequently modified at themonographic meeting in Toledo in 2002. ThisClinical History was presented on a CD, with thecollaboration of the Italfármaco laboratories.

A work entitled “Guide to Clinical Practice”produced by the working group on “Managementof Clinical History” was published in the SpanishReview of Bone Metabolic Diseases and theSpanish Clinical Review. The content of the guidewas developed over a period of two years. A draftwas presented at the congress held in Menorca in2001 and, and subsequently a debate was organi-sed in a forum open to all SEIOMM members atthe monographic meeting in Toledo. The intro-duction explained that the progressive increase inthe incidence of osteoporosis, parallel to thedemographic of aging in Spain, its morbidity andmortality, as well as its health and economicimpact had led SEIOMM to develop this guide asa first step aimed at the population group mostaffected. It offered an indicative framework, inwhich the interested professional groups coulddevelop action protocols adapted for each health-care environment.

The Guide to Clinical Practice was developedby a group of experts from different specialities(internists, rheumatologists, endocrinologists,gynaecologists, nephrologists and specialists infamily and community medicine), coordinated byan expert in evidence-based medicine. The groupwas formed by J. Calaf (gynaecologist), J. Cannata( nephrologist), B. Díaz (internist) A. Díez Pérez(internist), J. González Macías (internist), N.Guañabens (rheumatologist), F. Hawkins (endocri-nologist), A. Morales (rheumatologist), M. MuñozTorres (endocrinologist), X. Nogués (internist), J.M. Nolla (rheumatologist), P. Orozco (family doc-tor), R. Pérez Cano (internist), J. del Pino (inter-nist), J. M. Quesada (endocrinologist) and M. Sosa(internist).

The Society, alone or in collaboration with thepharmaceutical industry, stimulated active rese-arch in different areas of activity. 2003 was thethird anniversary of the SEIOMM-MSD researchcalls, while the well-established FAES andItalfármaco research prizes continued, as well as

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those from the Lilly Foundation which had beenpresented at all the congresses.

Apart from research, SEIOMM carried out anumber of educational activities, among whichwas the II Densitometry Course (BoneMeasurement Technique Accreditation) in April2001 in Barcelona under the direction of Luís delRío.

After many years in the post of editor of theSpanish Review of Bone Metabolic Diseases,Aurelio Rapado was succeeded by Manuel DiazCuriel, and later, by Federico Hawkins Carranzajointly with Esteban Jódar Gimeno as editorialsecretary. On a negative note, the lack of originalarticles put the continued publication of thereview at serious risk.

The number of members of the society increa-sed progressively from its foundation. In thisperiod there were 317 members.

At the National Assembly held in Seville in2000 it was agreed that members of the Societywho retired would become Emeritus Members. Inthis period a congress was organised in Ciutadella(Menorca) in 2001, and another in Maspalomas(Gran Canaria) in 2003. The V MonographicMeeting was held in Seville (2000) on the themeof “Osteoporosis in males”. This meeting wascoordinated with the annual meeting of FHOEMO.The VI Monographic Meeting took place in Toledo(2002).

During this period it was proposed that theoutgoing president would become part of thefollowing board of directors, since the experiencethey had gained in the previous years could beuseful.

From November 2003 until October 2007At the General Assembly held on 21stNovember2003 in Maspalomas (Gran Canaria) anew board of directors was appointed. This con-sisted of Nuria Guañabens Gay (President),Manuel Muñoz Torres (VicePresident), Carlos Gómez Alonso(Secretary) and Lius del RíoBarquero (Treasurer).

In relation to the Society´sfinances, the meeting of the boardof directors held in Cáceres in 2004discussed the difficulties in carryingout financial management within aproper legal framework. In order tofacilitate this, an agreement wasreached between the board ofdirectors and Pharma Consult forthe management of the congresses.The SEIOMM would receive thebenefits of the congress, once thecorresponding share for the localcommittee organiser and for FHOE-MO had been discounted, with theaccounting and financial manage-ment of all the expenditure andincome being the responsibility ofPharma Consult S.A.

During this period there was new revision of thearticles of association, which occurred at a meetingcalled in Madrid to coincide with the congress of2005. The following changes were approved:

- The national congress to be held annually,with the monographic meetings ceasing.

- The term of office of the board of directorswas reduced from 4 years to 3.

- The maximum period for those representingSEIOMM on other societies would be 3 years.

- The election of president a year before thechange of board of directors, to enable them tofamiliarise themselves with the essentials ofSEIOMM.

- That the procedures of the scientific program-mes be regulated.

A key activity was the updating of the guide toclinical practice for postmenopausal osteoporosisand its broadening to include corticoid and maleosteoporosis. A scientific committee was appoin-ted, coordinated by Jesús García Macías and for-med of Guañabens Gay, Muñoz Torres, del RíoBarquero and Díaz López. The updated guide waspresented at the SEIOMM congress of 2007.

The relationships with other societies conti-nued. Xavier Nogués Solán was the delegate forSEIOMM to the European Society for CalcifiedTissue and Luis del Río to SIBOMM, continuing hisprevious relationship with the IOF and FHOEMO.

At the 2007 SEIOMM congress a round tablewas organised jointly with the ASBMR and aSEIOMM round table was organised at the IBMScongress in Montreal in June 2007, as well as atperiodic meetings with the IOF.

The review Calcified Tissue International wasadded to previous subscriptions to foreign reviewssuch as Bone and Osteoporosis International, towhich members of the SEIOMM had free accessduring this period.

In 2004 SEIOMM began awarding grants tomembers for their attendance at the annual con-

gress of the ASBMR. Thesegrants have continued to thepresent day,

The SEIOMM congressestook place in Madrid (2005), inMalaga (2006) and in Valencia(2007). In this period there wasalready only one monographicmeeting (the final one) whichtook place in Cáceres on thetheme of “Bone formation fromthe basic to the clinical”. A newcourse for accreditation in den-sitometry was run by Luis delRío and Xavier Nogués. Thereissue of the course on techni-ques for the measurement ofbone mass, with the collabora-tion and accreditation of theInternational Society of ClinicalDensitometry was proposed.

The prizes from Italfarmáco,communicated orally and

THE SOCIETY, ALONE OR INCOLLABORATION WITH THEPHARMACEUTICAL INDUSTRY,STIMULATED ACTIVE RESE-ARCH IN DIFFERENT AREAS OFACTIVITY. 2003 WAS THETHIRD ANNIVERSARY OF THESEIOMM-MSD RESEARCHCALLS, WHILE THE WELL-ESTA-BLISHED FAES ANDITALFÁRMACO RESEARCH PRI-ZES CONTINUED, AS WELL ASTHOSE FROM THE LILLYFOUNDATION WHICH HADBEEN PRESENTED AT ALL THECONGRESSES


through posters, from FAES Research 2004-2005,from SEIOMM-MSD, and the SEIOMM prize for theyoungest researcher, continued.

The number of members increased up to 364during this period.

From October 2007 to October 2010The board of directors during this period was for-med by Manuel Sosa Henríquez (President), Javierdel Pino (Vice President), María Jesús Gómez deTejada Romero (Secretary) and Esteban JódarGimeno (Treasurer).

At the last Assembly of his period in office thepresident, Manuel Sosa Henríquez gave a sum-mary of the activities carried out, stating that theyhad they had met all the targets set:

- Provision of regular detailed information onthe management of the Society mainly throughbulletins and through the web.

- Creation of a new web site, which since thenhas been kept updated.

- In 2009 the Spanish Foundation for Bone andMineral Metabolism Research (FundaciónEspañola de Investigación Ósea y MetabolismoMineral [FEIOMM]) was founded, associated withSEIOMM, its first president being Manuel SosaHenriquez, with social objectives and dissemina-tion activities appropriate to foundations. In termsof the Foundation’s operational functions,FEIOMM research grants were created aimed atbasic, genetic and clinical research.

- Development of new Articles for SEIOMM,and coordinated with those of FEIOMM.

- The production of the Society’s own review(Review of Osteoporosis and Mineral Metabolism)given the difficulty in moving forward on thisbefore.

- Stimulating and facilitating research, withalmost 120,000 euros available for grants and thecreation of 10 working groups. In 2009 theSEIOMM’s committee of experts published the“Guide to clinical practice in postmenopausalosteoporosis, osteoporosis due to corticoids, andin men” (Rev Osteoporos Metab Miner 2009 1;153-60).

Up to this time, the administrative work of theSociety had been carried out by someone with thetrust of the Secretary of the board of directors intheir home city. This meant that each time theboard changed, all the documentation needed tobe moved. It was proposed, therefore, that thelocation of the secretariat be permanent. It wasthen decided that the company SANED wouldprovide this function and that they would make aperson available to SEIOMM who would performthese administrative tasks.

The relationships with the other national andinternational societies participating in the study ofosteoporosis and mineral metabolism continued.Notable among the international societies were theInternational Bone & Mineral Society, theInternational Osteoporosis Foundation, theEuropean Calcified Tissue Society, theInternational Society for Clinical Densitometry and

the Iberoamerican Society of Osteoporosis andMineral Metabolism. Adolfo Díez Pérez wasappointed as a member of the board of directors ofthe International Bone and Mineral Society, andManuel Díaz Curiel and Nuria Guañabens Gay tothe scientific committee of the InternationalOsteoporosis Foundation. Cooperative researchwork was carried out with other scientific societies,such as the “Consensus Document on Vitamin D”and the “Study of the Prevalence of Non-vertebralFractures in Patients in Treatment with Corticoids”.The board of directors aimed to keep all the asso-ciates informed and an email address was appro-ved for the dissemination of specific information.In addition, a SEIOMM Information Bulletin wascreated, which started to be produced in 2009 ona quarterly basis, through which provided conciseinformation to all the associates on all mattersregarding SEIOMM in the previous quarter, withthe intention that all associates would be fullyinformed and up to date with everything related toSEIOMM through the web site.

In a restricted zone of the web site, access wasgiven to Navibone, which allowed access tovarious reviews of interest such as OsteoporosisInternational, Calcified Tissue International andthe Journal of Bone and Mineral Metabolism. Inaddition, by being members of the IBMS, thanksto funding from the Nycomed Laboratories,SEIOMM members had access to the reviews Boneand BoneKey. Also, thanks to the work of AdolfoDíez Pérez, access was also available to the reviewProgress on Osteoporosis.

The documents most consulted through theweb site were SEIOMM´s clinical guides, followedby the information bulletins and the NOF Guides.The “Spanish Review of Bone Metabolic Diseases”had to cease publication due to financial difficul-ties and a lack of original articles. In 2009 the“Spanish Review of Osteoporosis and MineralMetabolism” was produced, replacing the afore-mentioned publication, issued three times a year,the third edition each year dedicated to recordingthe communications presented at the SEIOMMcongress. Manuel Sosa was (and continues to be)the editor of the review. In addition to the printedform, an online version was produced, with itsown web site accessible through a link from theSEIOMM web site. The Review was made openaccess for better dissemination, and is available intwo languages: Spanish and English.

During this period, the following projects wereestablished: a clinical case competition promotedby the Nycomed Laboratories; a blog on the website which could be called “Talk to an expert”; anon-line course on Medicine and Law. A collabora-tion agreement was signed between the IBMS andSEIOMM aimed at developing an IBMS-SEIOMMjoint round table, each society providing 50% ofthe costs. The first of these round tables was heldin 2009 during the Santander conference on thetheme “primary hyperparathyroidism”. Drs JanBolersven from Norway and Serge Ferrari (VicePresident) participated on behalf of the IBMS, and

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Drs Manuel Muñoz Torres, Estebán Jódar Gimenoand Manuel Sosa Hénriquez on behalf ofSEIOMM. This collaboration has continued at eachyear´s congress to the present day.

The number of members increased, reaching447 in this period.

In November 2008 the congress was held inOviedo in October 2009 the XIV congress inSantander, and in 2010 in Salamanca. In 2008 thecongress of the European Calcified Tissue Societytook place in Barcelona, A new course for accre-ditation in “Techniques for the Measurement ofBone Mass” was organised, led by Luis del Río andXavier Nogués.

At the Salamanca congress Juan José GarcíaBorrás requested that it be put on record that:“support be given to retired members who, due tothe lack of support from the pharmaceuticalindustry do not attend the congresses. Given thefavourable financial policy which has been put inplace by the board of directors and the creation ofthe Foundation, and since some funding has beenrequested for basic research, I ask that the inte-rests retired members be taken into account, andthat they are brought to the congresses, and thatat the next executive meeting considers this pleaand brings its decision to the next Assembly”.

During this period the FAES FARMA-SEIOMMprizes were awarded. Seven ASBMR grants conti-nued and a new Amgen-SEIOMM prize establis-hed. At the Santander congress of 2009, theItalfármaco Laboratories were honoured forhaving supported the prizes for the best oral andposter communications at the SEIOMM congressesfor more than 15 years without interruption.

Carlos Lozan Tonkin, Jordi Farrerons Minguellaand Juan José García Borrás were awarded hono-rary membership.

From October 2010The board of directors during this period is madeup of Javier del Pino Montes (President), JosepBlanch Rubió (Vice President), María Jesús MoroÁlvarez (Secretary) and Carmen Valero Díaz deLamadrid (Treasurer).

It was stated at the congress in Coruña in 2011that FEIOMM had responsibility for the incomeand expenditure of the congresses, which meansthat some of the benefits which are obtained fromthem should be used to accomplish the actionplans of FEIOMM, as well as funding its internalfunctions.

At this congress it was agreed to increase themembership fee which had remained unchangedfor 15 years. After a number of interventions, thechange from 27 to 35 euros in 2012, and then to40 euros in 2013, was approved. At a meetingduring this period some working committees wereestablished in accord with the objectives develo-ped using the Metaplan method, a method ofgroup moderation for problem-solving. The follo-wing groups were approved: economic manage-ment, training and congresses, research commit-tee, external relations and working groups. Based

on these objectives a strategic plan was developedwith a horizon of 2014.

In addition, a group of senior members ofSEIOMM was formed on the initiative of, and incollaboration with, Dr Juan José García Borráswho was appointed as coordinator of the group.

Relationships with other scientific societiescontinued. In collaboration with the SpanishSociety for Osteoporotic Fractures (SEFRAOS) andother related societies the “Blue Book onOsteoporotic Fractures in Spain” was produced. Acollaboration agreement was signed with AEEM tocollaborate in scientific activities related to pos-tmenopausal osteoporosis. SEIOMM maintained itsrelationships as part of SIBOMM and the IOF. Inrelation to the IBMS, due to a change in its com-munication strategy, it swapped its relationshipwith Elsevier for one with Nature, and stoppedreceiving Bone, this being substituted byBoneKey.

With regard to research, the ASBMR grantscontinued to be given and a revision was announ-ced of SEIOMM´s Guides to Clinical Practice,coordinated by Jesús González Macías.

In this period the SEIOMM congresses wereheld in La Coruña (2011), Cuenca (2012) and inTarragona (2013). In 2012 a SIBOMM congresswas held in Madrid.

During this period improvements were madeto information systems and information techno-logy in relation to Web 2.0, including restructuringand redesigning the SEIOMM-FEIOMM andReview (Review of Osteoporosis and MineralMetabolism) web site, and the creation ofSEIOMM groups on Facebook and Linkedin. Acloud computing service was initiated for the useof members. The BIDI code was incorporated inthe documents of the congresses and otherSEIOMM documents. SEIOMM and FEIOMM wereput on a sound footing, with the establishment oftheir own headquarters (Paseo de la Castellana135, Madrid) and an executive secretary (DªLorena Herrero).

A group of senior members and a young rese-archers group have been formed, with meetingsbeing organised post ASBMR.

The Cuenca congress approved the modifica-tion of the articles of association which wouldintroduce the following changes:

1. A new home for SEIOMM headquarters.2. Change to the closure of the accounts to

enable the most up to date financial report to begiven to the Assembly on June 30th each year.

3. An addition to the board of directors of twovoting members was proposed, as well as creatingthe position of President Elect (the futurePresident) to be incorporated into the board ofdirectors, so that the strategy and activities of theSociety may be planned with a horizon of twoperiods of office.

Note: this historical summary will be updatedwith the passage of time and as new databecomes available.

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