IbfisttwrpgcCfrpscpc

I

S

A

0©

5

The (Pro)Renin Receptor and the Kidney

Atsuhiro Ichihara, MD, PhD, FAHA, Yuki Kaneshiro, MD, Tomoko Takemitsu, MD,Mariyo Sakoda, MD, and Hiroshi Itoh, MD, PhD

Summary: Prorenin binding to the (pro)renin receptor not only causes a nonproteolyticactivation of prorenin leading to the activation of the renin-angiotensin system (RAS), but alsostimulates the receptor’s own intracellular signaling pathways independent of the RAS.Within the kidneys, the (pro)renin receptor is present in the glomerular mesangium andpodocytes, which play an important role in the maintenance of the glomerular filtrationbarrier. Therefore, prorenin-receptor blockers, which competitively bind to the receptor asa decoy peptide, have superior benefits with regard to proteinuria and glomerulosclerosis inexperimental animal models with elevated plasma prorenin levels such as diabetes andhypertension compared with conventional RAS inhibitors, possibly by inhibiting both thenonproteolytic activation of prorenin and RAS-independent intracellular signals.Semin Nephrol 27:524-528 © 2007 Elsevier Inc. All rights reserved.Keywords: Angiotensin, mesangium, nonproteolytic activation, podocytes, prorenin

rmIHnm

R

Stek(ha(mc((c(fmednp

n the kidneys, stimulation of the (pro)reninreceptor exerts 2 major pathways: the acti-vation of the renin-angiotensin system (RAS)

y the conversion of prorenin to the activeorm of prorenin by a conformational changenstead of the proteolytic cleavage of the pro-egment of prorenin,1,2 and the stimulation ofhe RAS-independent intracellular pathways viahe (pro)renin receptor.2–5 As shown in Figure 1,hen the (pro)renin receptor binds to the handle

egion of inactive prorenin, the receptor-boundrorenin gains its enzyme activity (ability toenerate angiotensin I) without proteolyticleavage of the prosegment of prorenin inOS-7 cells,6,7 presumably as a result of a con-

ormational change. On the other hand, theeceptor triggers its own intracellular signalingathways independent of the RAS. Studies havehown that stimulation of the (pro)renin re-eptor by renin/prorenin activates tyrosinehosphorylation, leading to activation of extra-ellular-signal–related protein kinases2 and up-

nternal Medicine, Keio University School of Medicine, Shinjuku-ku, Tokyo,Japan.

upported in part by grants from the Ministry of Education, Science andCulture of Japan (16790474 and 17390249).

ddress reprint requests to Atsuhiro Ichihara, MD, PhD, FAHA, AssociateProfessor of Internal Medicine, Keio University School of Medicine, 35Shinanomachi, Shinjuku-ku, Tokyo, Japan. E-mail: atzichi@sc.itc.keio.ac.jp

270-9295/07/$ - see front matter

(2007 Elsevier Inc. All rights reserved. doi:10.1016/j.semnephrol.2007.07.005

Seminars in N24

egulates transforming growth factor-�1 andatrix proteins without involving angiotensin

I generation in human and rat mesangial cells.4

owever, the physiologic roles of the (pro)re-in receptor in the kidneys remain undeter-ined.

ENAL LOCALIZATION

tudies have shown that the (pro)renin-recep-or protein and messenger RNA (mRNA) arexpressed in the mesangium cells of humanidneys.2,4 However, we recently found thatpro)renin-receptor mRNA also is present inuman cultured podocytes (unpublished data),nd a recent preliminary study reported thatpro)renin-receptor mRNA is present in rat glo-erular epithelial cells.8 Double immunohisto-

hemical analyses using a polyclonal antiratpro)renin-receptor antibody showed that thepro)renin receptor was colocalized with podo-alyxin (a podocyte marker), but not with Thy1.1a mesangium marker), nephrin, (a podocyteoot process marker), or reca-1 (an endotheliumarker) in rat kidneys (Fig. 2A). In addition,

lectron microscopic analyses appeared to in-icate the predominant presence of rat (pro)re-in receptor in podocytes, excluding the footrocesses, and its absence in mesangial cells

Fig. 2B). Thus, rat (pro)renin receptor was

ephrology, Vol 27, No 5, September 2007, pp 524-528

paWputrtciRc

PIA

Trthtbaswbbcdrvro

sctapitcmtrsptd

Fcr

FtaMlkpattc

(Pro)renin receptor and the kidney 525

resent predominantly in the major processesnd cell bodies of podocytes in rat kidneys.ithin the glomerulus, podocytes play an im-

ortant role in the maintenance of the glomer-lar filtration barrier, and podocyte injury leadso proteinuria and initiates glomerulosclerosis,esulting in the progressive loss of renal func-ion.9 Therefore, the podocyte (pro)renin re-eptor may contribute to proteinuria and renalnjury through a RAS-dependent pathway, aAS-independent pathway, or both pathways inhronic kidney diseases.

ATHOPHYSIOLOGIC ROLESN THE KIDNEYS OF DIABETICND HYPERTENSIVE MODELS

he (pro)renin receptor binds to the handleegion of inactive prorenin, thereby activatinghe receptor-bound prorenin.6,7 Therefore, theandle region peptide competitively binds tohe receptor as a decoy peptide and inhibitsoth the nonproteolytic activation of proreninnd RAS-independent intracellular signals. Ashown in Figure 3, when the decoy peptideas administered as a prorenin-receptorlocker (PRRB) to streptozotocin-induced dia-etic rats, PRRB significantly inhibited the in-rease in renal angiotensin II levels and theevelopment of proteinuria and glomeruloscle-osis,1 suggesting that the nonproteolytic acti-ation of prorenin by binding to the (pro)renineceptor plays an important role in the devel-

igure 1. Prorenin binding to the (pro)renin receptorauses the nonproteolytic activation of prorenin andeceptor-mediated intracellular signal transduction.

pment of nephropathy. In addition, as c

hown in Figure 4, although an angiotensin-onverting enzyme inhibitor failed to inhibithe glomerulosclerosis with increased mitogen-ctivated protein kinase (MAPK) activation androteinuria that developed in streptozotocin-

nduced diabetic angiotensin II type 1a recep-or– deficient mice, despite a significant de-rease in renal angiotensin II levels, PRRBarkedly abolished these conditions, despite

he absence of any change in the increase inenal angiotensin II levels.5 Therefore, angioten-in II–independent glomerular mitogen-activatedrotein kinase activation by the (pro)renin recep-or also may contribute to the development ofiabetic nephropathy.

igure 2. (A) Double fluorescent immunohistochemis-ry images showing rat (pro)renin receptor ([P]RR, red)nd podocalyxin, nephrin, reca-1, or Thy1.1 (green).erged images were obtained for (P)RR and podoca-

yxin. (B) Electron microscopy of the podocytes in the ratidneys. The majority of (P)RR was present in the majorrocesses (MP) of podocytes, as shown by the largerrows, and a few were observed in the cell body (CB) ofhe podocyte, as shown by the small arrows, but not inhe foot processes (FP) of the podocyte, in the mesangialells (MC), or in the endothelial cells (EC). RBC, red blood

ells in capillary lumen. Scale bars: 1 �m.

mbtdcgdfrspmt

mnscste

sbpis

FaD le bar

Flmaep

Fa(w

526 A. Ichihara et al

Dilated afferent arterioles and enlarged glo-eruli are observed in the early stage of dia-

etic nephropathy, thus glomerular hyperfiltra-ion is well known to play a role in theevelopment of diabetic nephropathy. The in-reased expression of macula densa cyclooxy-enase-2 (COX-2) is observed in the kidneys ofiabetic animals10,11 and is thought to accountor the hyperfiltration state of the glomeruli as aesult of afferent arteriolar dilation. Figure 5hows that the macula densa COX-2 mRNA androtein levels were significantly higher in hu-an (pro)renin receptor gene transgenic rats

han in wild-type rats,12 suggesting the involve-

igure 3. PRRB significantly inhibited the development ongiotensin I and II levels in diabetic rats (DM). e, ControM versus control. † P � .05 for PRRB versus vehicle. Sca

igure 4. Renal expression of phosphorylated extracel-ular-signal–related protein kinases (p-ERK) and renal

orphology in control rats, untreated diabetic rats (DM),nd DM treated with PRRB, an angiotensin-convertingnzyme inhibitor (ACEi), or both. Scale bars: 25�m. Re-

rinted with permission from Ichihara et al.5 s

ent of the (pro)renin receptor in the mecha-ism of increased macula densa COX-2 expres-ion. Thus, the (pro)renin receptor also mayontribute to the glomerular hyperfiltrationtate resulting from afferent arteriolar dilationhrough an increase in macula densa COX-2xpression in diabetic animals.

The plasma prorenin levels of stroke-pronepontaneously hypertensive rats (SHRsp) haveeen reported to be high,13,14 and increasedlasma prorenin levels cause end-organ damage

n the kidneys independently of blood pres-ure.15 Studies have shown that (pro)renin re-

einuria and glomerulosclerosis and the increases in renalcontrol � PRRB; �, DM; , DM � PRRB. * P � .05 for

s: 50�m. Reprinted with permission from Ichihara et al.1

igure 5. Enhanced COX-2 expression of tubular cellsdjacent to the macula densa, shown by arrows, inpro)renin receptor ([P]RR) transgenic rats compared withild-type rats. Scale bars: 25�m. Reprinted with permis-

f protl; ,

ion from Kaneshiro et al.12

cisfisstvathtptidaPapiss

C

Ippfapba

Tpg

R

FrWp

(Pro)renin receptor and the kidney 527

eptors, which bind to prorenin and stimulatets enzyme activity, are localized in prorenin-ensitive organs such as the kidneys.2 Thesendings suggest that the (pro)renin receptorequesters prorenin and activates it on theurface of the cells of critical organs suscep-ible to end-organ damage, and that the acti-ated prorenin in turn generates angiotensin Ind II locally, thereby exerting local actionshat lead to tissue damage. In SHRsp, whichave increased renal levels of (pro)renin-recep-or mRNA, the (pro)renin receptor activatedrorenin in the glomeruli of the kidneys, whereissue angiotensin I and II concentrations werencreased markedly; consequently, end-organamage occurred (Fig. 6).16 Because proreninctivation was blocked almost completely byRRB, resulting in a decrease of angiotensin Ind II levels and attenuation of damaged mor-hology in the kidneys of the hypertensive an-

mals, the (pro)renin receptor also may play aignificant role in the development of hyperten-ive renal damage.

ONCLUSIONS

n the kidneys, the (pro)renin receptor isresent predominantly in the mesangium andodocytes, and its blockade had beneficial ef-

ects on the kidneys of experimental diabeticnd hypertensive animal models with elevatedlasma prorenin levels through inhibition ofoth the nonproteolytic activation of prorenin

igure 6. Renal morphology and (pro)renin receptor mat (SHRsp), Wister-Kyoto rats (WKY), SHRsp treated with

KY. Abbreviations: PAS, periodic acid schiff; GAPDH,ermission from Ichihara et al.16

nd RAS-independent intracellular signals.

hus, the glomerular (pro)renin receptor ap-ears to contribute to the maintenance of thelomerular filtration barrier.

EFERENCES1. Ichihara A, Hayashi M, Kaneshiro Y, et al. Inhibition of

diabetic nephropathy by a decoy peptide correspond-ing to the “handle” region for non-proteolytic activationof prorenin. J Clin Invest. 2004;114:1128-35.

2. Nguyen G, Delarue F, Burckle C, et al. Pivotal role ofthe renin/prorenin receptor in angiotensin II produc-tion and cellular responses to renin. J Clin Invest.2002;109:1417-27.

3. Schefe JH, Funke-Kaiser H, Jost A, et al. Signal trans-duction of the renin/prorenin receptor [abstract].J Hypertens. 2005;23 Suppl 2:S354.

4. Huang Y, Wongamorntham S, Kasting J, et al. Reninincreases mesangial cell transforming growth fac-tor-b1 and matrix proteins through receptor-medi-ated, angiotensin II-independent mechanisms. KidneyInt. 2006;69:105-13.

5. Ichihara A, Suzuki F, Nakagawa T, et al. Proreninreceptor blockade inhibits development of glomeru-losclerosis in diabetic angiotensin II type 1a receptordeficient mice. J Am Soc Nephrol. 2006;17:1950-61.

6. Suzuki F, Hayakawa M, Nakagawa T, et al. Human pro-renin has “gate and handle” regions for its non-proteo-lytic activation. J Biol Chem. 2003;278:22217-22.

7. Nabi AHMN, Kageshima A, Uddin M, et al. Bindingproperties of rat prorenin and renin to the recombi-nant rat renin/prorenin receptor prepared by a bacu-lovirus expression system. Int J Mol Med. 2006;18:483-8.

8. Hamming I, Contrepas A, Nguyen G, et al. Regulationof the renal renin/(pro)renin receptor (RPR) by ACEinhibition and sodium restriction [abstract]. J Am Soc

expression in stroke-prone spontaneously hypertensive, and WKY treated with PRRB. *P � .05 for SHRsp versusaldehyde-3-phosphate dehydrogenase. Reprinted with

RNAPRRB

glycer

Nephrol. 2005;16:120A.

1

1

1

1

1

1

1

528 A. Ichihara et al

9. Pavenstadt H, Kriz W, Kretzler M. Cell biology of theglomerular podocyte. Physiol Rev. 2003;83:253-307.

0. Khan KN, Stanfield KM, Harris RK, et al. Expressionof cyclooxygenase-2 in the macula densa of humankidney in hypertension, congestive heart failure, anddiabetic nephropathy. Ren Fail. 2001;23:321-30.

1. Komers R, Lindsley JN, Oyama TT, et al. Immunohis-tochemical and functional correlations of renal cyclo-oxygenase-2 in experimental diabetes. J Clin Invest.2001;107:889-98.

2. Kaneshiro Y, Ichihara A, Takemitsu T, et al. Increasedexpression of cyclooxygenase-2 in renal cortex ofhuman-prorenin-receptor-gene transgenic rats. Kid-ney Int. 2006;70:641-6.

3. Obata J, Nakamura T, Takano H, et al. Increased gene

expression of components of the renin-angiotensinsystem in glomeruli of genetically hypertensive rats.J Hypertens. 2000;18:1247-55.

4. Ichihara A, Kaneshiro Y, Takemitsu T, et al. Non-proteolytic activation of prorenin contributes to de-velopment of cardiac fibrosis in genetic hyperten-sion. Hypertension. 2006;47:894-900.

5. Veniant M, Menard J, Bruneval P, et al. Vascular dam-age without hypertension in transgenic rats express-ing prorenin exclusively in the liver. J Clin Invest.1996;98:1966-70.

6. Ichihara A, Kaneshiro Y, Takemitsu T, et al. Contri-bution of non-proteolytically activated prorenin inglomeruli to hypertensive renal damage. J Am Soc

Nephrol. 2006;17:2495-503.