Case Report

Hypophosphatasia in an Adult:A Case Report

Masaomi NANGAKU,Noriharu SATO*, Kentaro SUGANOand Fumimaro TAKAKU

In this rare case of adult hypophosphatasia, no radiological abnormalities of the skeletal system couldbe detected even by dual energy X-ray absorptiometry. Severe dental caries was the sole clinical manifestation,indicating this case as an "odontohypophosphatasia". The levels of serum osteocalcin were low, which maybe a useful biochemical marker to diagnose hypophosphatasia. Southern blot analyses of the genomic DNArevealed no gross abnormalities. Thus, hypophosphatasia in this patient was presumed to be caused by pointmutations or small deletions. In a review of previous reports, an increased incidence amongwomenwas found.

Key words: Alkaline phosphatase, Bone mineralization, Genetic disease, Osteocalcin, Metabolic bonedisease

Hypophosphatasia is a rare inherited metabolicbone disease first described by Rathbun in 1948 (1).Defective bone mineralization coupled with a lowplasma alkaline phosphatase (Alp) and an increasedurinary phosphorylethanolamine (PEA) arecharacteristic of the disease (2). Three clinical typesof hypophosphatasia have been recognized:infantile, childhood, and adult (2). Adult

hypophosphatasia is the most rare form and hasbeen defined as ''first diagnosed in adult life". Theclinical and biochemical abnormalities appear to beless severe in adult hypophosphatasia than those inthe other types of hypophosphatasia.Herein we report a patient with the adult type ofhypophosphatasia. Bone mineral content of thepatient was measured using dual energy X-rayabsorptiometry (DEXA). Osteocalcin of her serumwas also measured. In addition, the leukocyte Alpgene structure was analyzed by meansof restrictionenzyme mapping.

METHODS

Total serum Alp activity was assayed using the

procedure of Bessey et al (3). Alp isozymes wereseparated by cellulose-acetate electrophoresis. PEAwas quantified using column chromatography (4).Serumosteocalcin concentrations were determinedby radioimmunoassay (RIA) with a Midori Juji RIAkit (CIS: Compagnie Oris Industrie SocieteAnonyme, Saclay, France).

Genomic DNAfrom leukocytes was preparedusing a standard method. The DNAswere digestedwith the following four restriction enzymes: EcoRI,BamHI, Bglll, and Hindlll. DNA fragments

obtained by restriction enzymeswere electrophoresedin 0.8% agarose gel and blotted onto nitrocellulosepaper. Southern blot analysis was performed usinga 32P-labeled probe, which contained a near full-length L/B/K Alp CDNAsequence (5) (kindlyprovided by Drs. Weiss and Harris). The nitro-cellulose paper was finally washed in O.l x SSC/0.1% SDSat 55°C, and was autoradiographed for3-5 days.

CASE REPORT

The patient, a 47-year-old Japanese female, wasFrom The Third Department of Internal Medicine, Faculty of Medicine and *Institute of Medical Science, University ofTokyo, Tokyo

Received for publication February 26, 1990; Accepted for publication August 30, 1990Reprint requests should be addressed to Masaomi Nangaku, MD,The First Department of Internal Medicine,

Faculty of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113, Japan

Jpn J Med Vol 30, No 1 (January, February 1991) 47,

Nangaku et al

the third child of non-consanguineous parents. Shehad no history of delayed growth or of rickets duringchildhood. At 20 yrs of age she suffered fromappendicitis complicated by peritonitis. She

developed dental problems at about 22 yrs of age,when fillings did not hold and her teeth becamecarious. Five yrs before admission, she noticedhematuria, which was diagnosed as idiopathic

hematuria. The decreased activity of serum Alp (5 1U/l: normal 66 to 220) was overlooked at that time.One yr before admission, intermittent abdominalpain developed. The pain was thought to be derivedfrom adhesion of the intestinal tracts due to previousperitonitis, but hypophosphatasia was thensuspected due to the low Alp levels for a greater than5-year duration at this time.There was no positive family history of any bonediseases, but her mother had complete dentures byage 20. Her youngest brother and youngest sisterdied in infancy from unknowncauses. She was notmarried and had no children.On physical examination the patient was 145 cmtall and weighed 51 kg. Examination of the lungs,heart, and abdomenwas unremarkable. Menstrualcycles were irregular but present. All but six teethhad been extracted. An electrocardiogram, plainX-ray films of the chest and of the abdomen werenormal. An ultrasonographic examination of theabdomen revealed a right renal cyst. Skeletalradiographs showed no signs of osteomalacia. Bone

Table 1. Bone mineral content

1. R igh t h ip sca n B M D (g / c m 2)

f e m o r a l n e c k 0 . 7 4 7

t r o c h a n t e r m a j o r 0 . 7 4 1

W a r d ' s 0 . 7 3 0

2. L ef t h ip s ca n

f e m o r a l n e c k 0 . 7 2 9

t r o c h a n t e r m a j o r 0 . 7 2 0

W a r d ' s 0 . 7 4 7

3. S p in e sca n

L 2 1. 0 2 8 (1 .0 2 0 ｱ 0 .13 9 )

L 3 1 . 0 9 8 ( 1 . 0 3 1 ｱ 0 . 1 4 3 )

L 4 0 . 9 6 9 ( 0 . 9 7 8 ｱ 0 . 1 1 4 )

L 2 -4 1. 0 2 8 (1 .0 12 ｱ 0 .12 4 )

4 . W h o l e b o d y s c a n B M C : 2 1 2 3 . O g B M D : 0 . 7 6 7 g / c m 2

BMC,bone mineral content: BMD,bone mineral densityReference data of 10 healthy age-matched womenare givenon the right as mean±SD

mineral content was measured by DEXA(NorlandXR-26) (Table 1). Bone mineral density was judgedas normalbased on the reference values kindly madeavailable by Dr. Arai of Hokkaido University.A routine laboratory evaluation, including serumcalcium, phosphorus, and magnesium levels, wasnormal except for serum Alp levels, which were con-sistently low, 46-50 U/l. Electrophoretic fractiona-tion of the serum Alp yielded isozymes composedof 59% liver, 35% bone, and 6% intestine. Urinaryexcretions of PEAwere also persistently high,200-250 ^mol/day (normal: ^ 100) (Table 2). Herserum osteocalcin was demonstrated to be low.To examine the spectrum of mutations that causeadult hypophosphatasia, L/B/K Alp DNAwasanalyzed in leukocytes from the patient. Southernblot analyses of the L/B/K Alp gene from thepatient and from controls revealed identical restric-tion patterns (Fig. 1).

Fig. 1. Southern blot analysis of the L/B/K Alp gene.Identical restriction patterns are shown. Digested with EcoRI,2.5, 10.5, and 14.5 kb fragments are demonstrated. Digestedwith BamHI, 6.5, 7.4, and 17.0 kb fragments are revealed.Digested with Bglll, 2.5, 3.2, 8.8, and 9.7 kb fragments areexhibited. Digested with Hindlll, 3.6, 6.2, and 13.5 kbfragments are illustrated. Migration of DNAsize markers(in kilo base pairs) is indicated on the right. Our patient, lanes1, 4, 7, 10; HL60 (leukemic cell line), lanes 2, 5, 8, ll;Control, lanes 3, 6, 9, 12.

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Adult Hypophosphatasia

Table 2. Laboratory data

Corrected calciumPhosphorusMagnesiumAlpPTH-C

PTH-M

CalcitoninO steocalcin25(OH)vitamin DNAP score9.0 mg/dl4.3 mg/dl

1.9 mg/dl46-50 U/l (66-220)*320 pg/ml (430-1,860)255 pg/ml (180-560)90.0 pg/ml ( £ 184)1.7 ng/ml**

18 ng/ml (10-30)267 (200-320)

Urinary calcium: calcium/creatinine 0. 101Urinary phosphate: TmPO4/GFR4.4mg/dlUrinary PEA 200-250 /imol/day ( ^ 100)

* Normal values in parentheses** Reference data on 7 healthy women; 5.2± 1.3

DISCUSSION

The adult form of hypophosphatasia may includea history of early loss of dentition, spontaneousfractures, often with a history of "rickets" inchildhood, and radiolucent zones in the skeleton.Somehave been recognized by chance measurementof plasma Alp. Subnormal serum Alp activity isassociated with several conditions, such as inanition,hypothyroidism, scurvy, pernicious anemia,

magnesiumdeficiency, and excess of vitamin Dingestion (6). Each of these factors was excluded inthis patient.

To date 50 cases have been reported in somedetail in the English literature. Another 2 cases havebeen reported in Japan (7, 8) (Table 3). The mostcommonsymptomswere reported to be edentia andpathological fractures. Other rare complicationsinclude chondrocalcinosis (9-12), calcification ofintervertebral disks or of spinal ligaments (1 3-16),renal calculus (15), autoimmune hemolytic anemia(17), hyperparathyroidism (1 8), and hyperthyroidism(7).

ThoughWhyteet al in a review described anequal incidence in either sex (15), they suggested thepossibility that the adult form of hypophosphatasiacould affect womenmore severely in a later report(1 1). Wendling also suggested an increased incidencein women(19). Reviewing the case reports, we foundan apparent preponderance of women, especially inthose over the age of 50. The male-to-female ratio

Number ofthe patients

10

5-13Male

H] Female

_UL-20 -30 -40 -50 -60 -70 à"

£81 Age

Fig. 2. Age distribution of adulthypophosphatasia. An apparent pre-ponderance of women, especially thoseover the age of 50, is shown. Dotted bars,males; Hatched bars, females.

is approximately 1:3. Age distribution of theproposita is shown in Fig. 2. The average age of theproposita was 49.4 yrs old. The male-to-female

ratio of the proposita was almost the same as thatof all the patients. In addition, three of four maleproposita over the age of 50 were recognized bychance measurement of Alp. The true reason for thisfemale preponderance is unclear. In post-menopausal women, the tendency to developosteoporosis may promote some latent cases.Conversely, bone diseases may be accelerated oraggravated during this period in women, thus theyare more likely to consult a physician.Weinstein and Whyte suggested heterogeneity ofadult hypophosphatasia (6). Analysis of theliterature suggested that roughly two types of adulthypophosphatasia exist, a biphasic type and amonophasic type. In the first type, childhoodhypophosphatasia exists and then becomesquiescent, only to re-emerge after adolescence as theadult of the childhood illness. Skeletal deformitiesare also apparent. It is usually inherited as anautosomal recessive trait. The second type has a lateronset and takes a mild course. There is no historyof childhood "rickets" and no skeletal deformitiesare apparent. It is usually inherited as an autosomaldominant train. The patient had no history of"rickets" nor skeletal deformities. Her mother couldhave suffered from the same disease in view of herdental problems. Thus, this case appears to fall intothe second group.A new X-ray-based instrument for the measure-ment of bone mineral, DEXA,is now available (20,

Jpn J Med Vol 30, No 1 (January, February 1991) 49

Nangaku et al

Table 3. Reported Cases of Adult HypophosphatasiaC a se S ex / A g e C h ild h o o d S k eletal P a th o lo g ica l D en ta l G en etics

N o. A g e " R ick ets" d e fo rm itie s fra c tu res d iso rd e rs su sp e cte d

1 * M a c e y : 1 9 4 0 ( P r o c S t a f f M / 3 6 + ? ?

2 * M a c e y : 1 9 4 0 M e e t i n g s M a y o C l i n ) M / ? + ? ?

3 * H e n n e m a n : 1 9 5 6 ( A m J M e d ) M / 4 4 + ? ?

4 * D e n t : 1 9 5 6 ( C I B A F o u n d a t i o n S y m p o s i u m ) F / 4 2 + + A R

5 * O w e n : 1 9 5 8 ( C l i n R e s ) F / 3 0 + + 9

6 * B e i s e l : 1 9 6 0 ( A m J M e d ) M / 3 7 + + S p

7 B e t h u n e : 1 9 6 0 ( A m J M e d ) F / 4 1 + A R

8 * S i l v e r m a n : 1 9 6 2 ( A r c h I n t e r n M e d ) M / 2 8 + + A D

9 * P i m s t o n e : 1 9 6 6 ( A n n I n t e r n M e d ) F / 5 1 + A R

1 0 * B i r t w e l l : 1 9 6 7 ( A r c h I n t e r n M e d ) M / 4 9 + ? S p

1 1* J a rd o n : 1 9 70 (J B o n e Jt S u re ) F / 54 + + A R

1 2 * O 'D u ffy : 19 7 0 (A rth ri tis R h e u m ) F / 5 1 + ? A R

1 3 * M a c p h e r s o n : 1 9 7 2 ( J C a n A s s o c R a d i o l ) F / 5 3 + + A R

1 4 * F a a s : 1 9 7 4 ( C l i n O r t h o p ) F / 7 0 + ? 9

1 5 * S o r e n s e n : 1 9 7 5 ( A c t a M e d S c a n d ) F / 5 3 + + ?

1 6 * W h y t e : 1 9 7 9 ( M e d i c i n e ) F / 5 8 + + A D

17 W h y te : 19 7 9 M / 5 7 + A D

18 W h y te : 19 7 9 M /4 8 + A D

19 W h y te : 19 7 9 F / 4 4 + A D

2 0 W h y te : 19 7 9 M /4 0 + A D

2 1 W h y te : 19 7 9 F / 3 3 A D

2 2 W h y te : 19 7 9 F / 2 4 + A D

2 3 W h y te : 19 7 9 F / 2 3 A D

2 4 W h y te : 19 7 9 F / 2 5 A D

2 5 * A n d e r t o n : 1 9 7 9 ( J B o n e J t S u r g ) F / 6 2 + ? ?

2 6 * A n d e r t o n : 1 9 7 9 F / 6 7 + ? ?

2 7 * W e i n s t e i n : 1 9 8 1 ( A r c h I n t e r n M e d ) F / 5 8 + S p

2 8 * E a d e : 1 9 8 1 ( A n n R h e u m D i s ) M / 5 7 + ? ?

2 9 * W h y t e : 1 9 8 2 ( J C l i n E n d o c r i n o l M e ta b ) F / 5 3 + + A D

3 0 W h y t e : 1 9 8 2 ( J C l i n E n d o c r i n o l M e t a b ) F / 6 2 + + A D

3 1 W h y te : 19 8 2 F / 5 6 + A D

3 2 W h y te : 19 8 2 M / 5 1 A D

3 3 W h y te : 19 8 2 F / d e a d A D

3 4 W h y te : 19 8 2 F / 4 2 +

3 5 * W h y t e : 1 9 8 2 F / 3 7 + ?

3 6 * W h y t e : 1 9 8 2 M / 4 0 + ?

3 7 * W h y t e : 1 9 8 2 F / 1 9 + ?

3 8 * C o t o : 1 9 8 3 ( S o u t h M e d J ) M / 6 0 4 - + 9

3 9 * P illa n s: 19 8 3 (G a stro e n tero lo g y ) M / 6 9 S p

4 0 * E b e r l e : 1 9 8 4 ( K l i n W o c h e n s c h r ) M / 8 0 + A D

4 1 E b e rle : 19 8 4 M / 8 0 + A D

4 2 E b e rle : 19 8 4 F / 4 3 + A D

4 3 E b e rle : 19 8 4 F / 2 5 + A D

4 4 * M a c f a r l a n e : 1 9 8 5 ( N e t h J M e d ) F / 2 4 + + A R

4 5 * S t i n s o n : 1 9 8 5 ( N E n g l J M e d ) F / 3 7 + ? ?

4 6 * B r e n t o n : 1 9 8 6 ( C l i n R h e u m D i s ) F / 4 5 + + ?

4 7 * B r e n t o n : 1 9 8 6 ( C l i n R h e u m D i s ) F / 5 9 + ?

4 8 * C o e : 1 9 8 6 (J B o n e J t S u r e) F / 4 6 + ? ?

4 9 * P a lm er : 19 8 8 (J A G S ) M / 9 0 4 - + S P

5 0 * H a r p e r : 1 9 8 9 ( F o o t A n k l e ) F / 3 9 + + S p

5 1 * A k a i : 1 9 8 1 ( O r t h o p S u r e ) F / 6 8 4 - + ?

5 2 * S a k u r a i : 1 9 8 8 ( J J p n S o c I n t e r n M e d ) F / 2 2 S p

Skeletal deformities include deformed long bones, abnormal skull, and lordosis. *proposita. AD, Autosomal Dominant;AR, Autosomal Recessive; Sp, Sporadic

50 Jpn J Med Vol 30, No 1 (January, February 1991)

Adult Hypophosphatasia

21). In the analysis of the bone mineral density(BMD) of this patient by DEXA, the BMDwaswithin normal limits, and there were no radiologicalfeatures suggesting metabolic bone disease. In somecases of hypophosphatasia described thus far, dentalcaries was the only symptom which could be relatedto hypophosphatasia (15, 22). Even bone biopsyspecimens showed normal findings (22). The termodontohypophosphatasia is suggested in thosereports where the dental findings are the pre-dominant manifestations of the disease (23). Someof the patients were totally asymptomatic, althoughit was apparent that they were biochemically affected(8, 15). The present patient fits in this type of

hypophosphatasia (odontohypophosphatasia) , andit is not surprising that no radiological abnormalitiesof the skeletal system could be detected.

The serum levels of osteocalcin were alsomeasured in this patient. Osteocalcin, also referredto as "bone Gla-protein (BGP)", represents one ofthe most abundant non-collagenous bone proteinsand is presumed to be produced by osteoblasts (24);it serves as a specific chemical index of bone turn-over (25, 26). A highly significant correlation wasdemonstrated between serum osteocalcin and bonemineralization in a calcium kinetic study (27).

Several reports indicated that serum osteocalcin islow in hypoparathyroidism, hypothyroidism, andCushing's syndrome (25, 28, 29). In the present

patient the serum concentrations of osteocalcin werelow, which may suggest a decreased bone mineraliza-tion rate in this disease. Thus, some adult hypo-phosphatasia cases, including the present case, maybe characterized as a disease in which the bonemineralization rate rather than the bonemineraliza-tion itself is disturbed without any obvious endo-crinological or nutritional abnormalities.The specific gene defects in hypophosphatasia areunknownbut are thought to occur either at theliver/bone/kidney (L/B/K) Alp locus or withinanother gene that regulates L/B/K Alp expression.Weiss et al observed a point mutation of the CDNAin a patient with an infantile form of the disease (30).They could not detect the same mutation in thegenomic DNAof unrelated individuals with variousforms of hypophosphatasia, which is thought to beconsistent with genetic heterogeneity for the disease.Weiss et al also analyzed L/B/K Alp DNA, RNA,

and enzyme activity in cultured skin fibroblasts from14 unrelated patients with infantile hypophos-

phatasia, and concluded that most cases of severehypophosphatasia are caused by point mutations or

small deletions in the L/B/K Alp gene (31). Here,southern blot analyses of the L/B/K Alp genes fromthis patient were performed, and the restriction

patterns obtained were identical to controls. Theserestriction patterns showed good agreement with therestriction map of the L/B/K gene reported by Weisset al (32). Although we can not refute the sugges-tion that there are large deletions or rearrangementsin fragments not shown in our southern blot

analyses, these results suggest that adult hypophos-phatasia is caused by point mutations or small dele-tions in the L/B/K Alp gene as proposed in perinatalor infantile forms of hypophosphatasia (31).

CONCLUSIONS

In conclusion, here a rare case of adulthypophosphatasia was reported. The diagnosis wasconfirmed through the discovery of low serum Alpactivity and the excretions of abnormally high levelsof PEAin the urine. Reviewing previous cases, anapparent predominance of womenand heterogeneityin the adult form of this disease were found.The present results suggest that measurement of

the serum osteocalcin level may be a usefulbiochemical marker for the diagnosis of hypophos-phatasia. DNAanalysis of the present patient re-vealed no major deletion or rearrangement in theL/B/K Alp gene. In order to elucidate the patho-mechanismand determine the proper treatment,more cases must be studied.

ACKNOWLEDGEMENTS:We thank Drs. ToshitakaNakamura, Tetsuo Nakamura, and Toshio Matsumoto fortheir helpful discussions.

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