T. Wang, Y. Cui, P. Liu, J. He, Q. Zhang

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1 O R I G I N A L A R T I C L E Foli Morphol. Vol. 78, No., pp. 4 3 DOI:.563/FM.8.67 Copyright 9 Vi Medic ISSN journls.vimedic.pl Distriution of epiderml growth fctor receptor, one morphogenetic protein-, nd p53 in kidney of helthy neworn, dult nd old highlnd-plteu yks T. Wng, Y. Cui, P. Liu, J. He, Q. Zhng Fculty of Veterinry Medicine, Gnsu Agriculturl University, Lnzhou, Chin [Received: 8 Mrch 8; Accepted: 3 My 8] Bckground: Kidney hs long een thought to e ody s lrgest orgn of elimintion for mintining cid-se lnce. In recent yers, the reserch on kidneys hs minly focused on the structurl chrcteristics of the kidney of single ge group nimls. In this pper we used histologicl nd immunohistochemicl methods to oserve nd compre the structure chrcteristics of yk kidney nd the expression of epiderml growth fctor receptor (EGFR), one morphogenetic protein- (BMP-) nd p53 in the kidney of yks of three different ge groups. The im of the study ws to investigte histologicl chrcteristics of ge-relted chnges in the kidney of yk nd expression nd loclistion of kidney-relted fctors. Mterils nd methods: Fifteen helthy mle nd femle yks from highlnd plteus (three groups: neworn, dult nd old yks, n = 5 per group). Histologicl methods were used to compre the relevnt chrcteristics of the kidney of yks. The immunohistochemistry method ws used to oserve the expression nd loclistion of EGFR, BMP-, nd p53 of the kidney of different ges, nd the opticl density vlue ws mesured nd nlysed y using imge nlysis softwre. Results: This is n overll oservtion of the kidney tissue section, which includes the surfce of the renl cpsule nd the internl prenchym. In the renl prenchym, there re renl corpuscles, renl tuules. The internl sustnce included cortex nd medull, which were ounded y the rched rtery. In the cortex, there were renl corpuscles, convoluted prt of renl tuules (proximl convoluted tuule nd distl convoluted tuule) nd collecting tuules. The medull included stright prts of renl tuules (proximl stright tuule nd distl stright tuule), thin segments nd collecting tuules. It ws oserved tht the orgnistionl structure of the kidney of yks did not chnge with ge, ut the degree of development of the internl structure (glomeruli, renl tuules nd collecting tuules) of the kidney chnged with ge. Immunohistochemicl results demonstrted tht EGFR nd BMP--positive rection in the neworn group ws minly distriuted in the proximl tuule epithelil cells, nd widely distriuted in the dult nd old groups. However, the p53-positive rection ws widely distriuted in the neworn, dult nd old groups. Conclusions: The results reveled tht the kidney structure tended to e completed with ge, nd the function of the kidney grdully improved. EGFR nd BMP- hd the effect of promoting kidney development. However, p53 hd een widely distriuted in the neworn kidney of the yks. It is suggested tht p53 hd een involved in cell migrtion nd metolic differentition nd self-renewl in the new stge. (Foli Morphol 9; 78, : 4 3) Key words: one morphogenetic protein-, epiderml growth fctor receptor, kidney, tumour suppressive gene p53, yk Address for correspondence: Dr. Y. Cui, College of Veterinry Medicine, Gnsu Agriculturl University, Lnzhou 737, Chin, tel: , e-mil: cuiyn369@sin.com 4

2 T. Wng et l., Distriution of EGFR, BMP-, nd p53 in yk kidneys INTRODUCTION As the lrgest excretory orgn in the ody, the kidney plys vitl role in regulting metolism nd cid-se lnce in the ody. The existing reserch on kidneys hs een minly focused on the structurl chrcteristics of the kidney of single ge group nimls such s dog, rt, mouse nd, Bctrin Cmels [, 36]. However, studies on the kidney of different ges were found only in mouse [37] nd humn [8]. So fr, since the yk (Bos grunniens) is species tht reproduced in the lpine zone of the Qinghi-Tiet Plteu nd the study focused on respirtory system [38], circultory system [3], immune system [4] nd reproductive system [5], including orgnistionl structure, ultrstructure nd some fctors. There is no detiled informtion from histologicl studies on the impct of kidney-relted fctors nd ge on yk kidneys. Epiderml growth fctor receptor (EGFR) is kind of glycoprotein, which elongs to the tyrosine kinse receptor, nd its moleculr weight is 7 kd. EGFR is locted on the surfce of the cell memrne nd is ctivted y inding to lignd, including EGF nd trnsforming growth fctor (TGF)-lph [8]. EGFR plys n importnt role in physiologicl processes such s growth, prolifertion, nd differentition of cells [3]. Studies hve shown tht EGFR is widely distriuted in the kidney [33]. The distriution of EGFR in humn kidney tissue is the sme s tht in mouse nd rt kidney tissue []. It hs recently een reported tht ctivtion of EGFR leds to phosphoryltion of Smd, the downstrem trget of one morphogenetic protein, y mitogen-ctivted protein kinse (MAPK)-dependent mechnism []. Bone morphogenetic proteins (BMPs) re group of osteogenic fctors, nd structurlly, BMPs re highly homologous to TGF-et fmily of proteins [4, 7]. BMPs exert their iologic effects vi type II nd type I serine-threonine kinse receptors [, 4, 7]. In the current study, BMPs hve een shown to ply n importnt role in the development of gut, hert, skin, teeth, lung nd kidney [6]. High-ffinity inding sites for BMP-, specific memer of this fmily of proteins, hd een identified in different cells nd tissues including the kidney [9]. BMP- my regulte oth kidney development nd dult tissue mintennce [35]. The tumour suppressive gene p53 is widely known s key regultor of poptosis y cting oth s n ctive component of the poptosis cscde nd s trnscription fctor [3]. p53 is one of the key regultory genes ctivted during kidney development which induces cell cycle rrest or poptosis to mintin genome integrity [5]. A previous study demonstrted tht p53 phosphoryltion is involved in the development of the kidney [5]. Intruterine growth restriction (IUGR) model estlished y uterine rtery ligtion oserved prllel increse in p53 protein level nd poptosis in the kidneys of IUGR rts fter irth [3]. p53 signlling is highly complex nd involved in kidney development through multiple pthwys []. In this study, we used histologicl nd immunohistochemicl methods to oserve nd compre the structure chrcteristics of yk kidney nd the expression of EGFR, BMP- nd p53 in the kidney of yks of three different ge groups. The purpose of this study ws to explore the ge-relted chnges of helthy plteu yks nd the expression of renl- -relted fctors in the kidney yks of different ges. It lid the iologicl nd pthologicl foundtion for the study of the urinry system in this plteu niml. Animls MATERIALS AND METHODS Fifteen highlnd-plteu yks were used for the study. Five of them were purchsed in Gnnn Tietn Autonomous Prefecture, nd the rest were purchsed in Qinghi Province. This study ws pproved y the Stte Forestry Administrtion, nd ll procedures were performed in complince with guidelines for the cre nd use of lortory nimls dopted y the Ministry of Science nd Technology of the People s Repulic of Chin. Fifteen cliniclly norml yks were divided into the following three ge groups: neworn ( 7 dys, n = 5), dult (3 6 yers, n = 5) nd old (7 yers, n = 5). All groups included oth mles nd femles. Ech yk ws euthnized with pentoritl sodium ( mg/kg, IV). The kidney ws hrvested t 3 min fter euthnsi. For histologic nd immunohistochemicl nlyses, smll specimens of kidney tissue were fixed in solution of 4% prformldehyde in phosphte uffer (ph, 7.3). Light microscopy Prformldehyde-fixed tissue specimens were emedded in prffin. The tissue ws then sliced into four µm-thick sections, nd the distriution of collgen nd sic structure in the kidney ws oserved y Msson s trichrome (niline lue) stining, 5

3 Foli Morphol., 9, Vol. 78, No. nd the structure of the rched rtery ws oserved y AB-PAS/Msson s trichrome (niline lue) doule stining. Hemtoxylin nd eosin (H&E) stining ws used to oserve the difference in kidney structure t different ge. The ove structures were oserved y opticl microscope (Olympus DP7, Tokyo). Immunohistochemicl nlysis For immunohistochemicl stining to investigte EGFR, BMP- nd p53 expression levels, kidney tissue sections were deprffinised in xylene nd rehydrted through grded series of lcohols. The ntigen ws repired with 3% deionised HO (5 min) nd seled with got serum (5 min). Primry rit nti-egfr, nti-bmp- nd nti-p53 polyclonl ntiodies (Bioss, Beijing, : dilution) were pplied nd sections were incuted t 4 C overnight. The sections were then incuted with the got nti-rit ntiody nd peroxidse complex. The lelled sections were then counterstined with 3-3 diminoenzidine [] nd the nucleus lightly counterstined with hemtoxylin. (Fig. C). The medull included stright prts of renl tuules (proximl stright tuule nd distl stright tuule); thin segments nd collecting tuules in the depths of the medull were generlly referred to nipple tues (Fig. D). In ddition, the corticl nd medullry tuules were rich in collgen fires. The numer of glomerulr in the kidney cortex of yk decresed with ge. The size of glomeruli incresed with ge nd the distriution of cells in the neworn yk glomeruli ws closed, especilly in the mrginl region, where the cells were closely ligned. However, the gp etween dult nd elderly glomeruli cells ecme lrger, the cells were loosely rrnged, nd the numer of cells in the mrginl zone decresed (Fig. A C). The study on the structure of renl tuules nd collecting tuules in yk kidney of ll ge groups were shown in Figure A F. The results showed tht compred with the dult nd old yk renl tuules, neworn yk renl tuulr structure hd not een fully developed, nd distl tuule wll ws not ovious, ut the proximl tuule ws ovious. Sttisticl nlysis Intensity mesurements for immunohistochemicl ssy were performed using integrted opticl density nd mesured y Imge nlysis softwre (Imge-Pro Plus version 6. Medi Cyernetics, USA). Sttisticl nlysis ws performed using the Sttisticl Pckge for Socil Science softwre (version 9., IBM Corp, Armonk). Vlues of p <.5 were considered significnt. Histologic chrcteristics RESULTS The surfce of the yk kidney ws covered y memrne tht composed of collgen fires (Fig. A) nd under the memrne ws the sustnce. Sustnce included cortex (Fig. C) nd medull (Fig. D) which were ounded y the rched rtery (Fig. B). In the cortex, there were renl corpuscles, renl tuules nd collecting tuules. The ellipsoidl renl corpuscles were respectively glomeruli nd renl cpsule from inside to outside. The glomeruli were minly composed of network of cpillries nd the wll of the renl cpsule ws rich in collgen fires (Fig. A). The end prt of renl tuules included proximl convoluted tuule nd distl convoluted tuule. In the cortex, the collecting tuules were divided into two prts, which were ow type tuules nd stright set tuules Immunohistochemicl loclistions Distriution chrcteristics of EGFR in the kidney of the yk. The EGFR positive rection ws minly distriuted in the proximl tuule epithelil cells of the neworn group (Fig. 3A, D), ut in the dult (Fig. 3B, E) nd old group (Fig. 3C, F), the renl tuulr epithelil cells were extensively distriuted nd the collecting tuule epithelil cells nd glomerulr cells demonstrted slight distriution. Distriution chrcteristics of BMP- in the kidney of the yk. The positive rection of BMP- in the neworn group minly distriuted in the proximl tuule epithelil cells (Fig. 4A, D). The positive rection of BMP- in the dult group (Fig. 4B, E) nd the old group (Fig. 4C, F) ws widely distriuted in the renl tuulr epithelil cells nd the collecting tuule epithelil cells. The mcroscopic oservtion found tht distl tuules positive rection ws more thn the proximl tuule positive rection. Distriution chrcteristics of p53 in the kidney of the yk. p53 positive rections were widely distriuted in ll ge groups of glomerulr cells, renl tuulr epithelil cells nd collecting tuule epithelil cells. The mcroscopic oservtion found tht the positive rection ws more intense in distl tuulr epithelil cells (Fig. 5A F). 6

4 T. Wng et l., Distriution of EGFR, BMP-, nd p53 in yk kidneys Figure. Representtive photomicrogrphs of kidney sections from helthy highlnd-plteu yks in two ge groups (neworn [ to 7 dys old; A nd B], old [7 to yers old; C nd D]) illustrting histologic chrcteristics (A through D). To ssess the histologic chrcteristics, sections were stined with Msson s trichrome (niline lue) nd AB-PAS/Msson s trichrome (niline lue) doule stin. Br = µm. Inserts in pnels C through D: Higher-mgnifiction views of the rched rtery, renl cpsule nd renl tuules. Br = µm. C cpsule; MC mlpighin corpuscle; G glomeruli; RC renl cpsule; PCT proximl convoluted tuule; DCT distl convoluted tuule; A rched rtery; JCM junction of cortex nd medull; CT collecting tuule; CF collgen fire; PST proximl stright tuule; DST distl stright tuule; TS thin segment. Figure. Representtive photomicrogrphs of kidney sections from helthy highlnd-plteu yks in three ge groups (neworn [ to 7 dys old; A nd D], dult [3 to 6 yers old; B nd E], or old [7 to yers old; C nd F]) illustrting histologic chnges (A through F). To ssess the histologic chnges, sections were stined with H&E stin. A through C: r = µm. D through F: r = 5 µm; G glomeruli; PCT proximl convoluted tuule; DCT distl convoluted tuule; CT collecting tuule; PST proximl stright tuule; DST distl stright tuule. 7

5 Foli Morphol., 9, Vol. 78, No. Figure 3. Representtive photomicrogrphs of kidney sections from helthy highlnd-plteu yks in three ge groups (neworn [ to 7 dys old; A nd D], dult [3 to 6 yers old; B nd E], or old [7 to yers old; C nd F]) illustrting the loclistion of epiderml growth fctor receptor (EGFR) (A through F). Immunohistochemicl stining ws used to identify EGFR, which ppers in yellow. Br = µm. Inserts in pnels A through F: Higher-mgnifiction views of the positive expression of EGFR in the kidney. Br = µm; C cortex; M medull. See Figure for the reminder of the key. Figure 4. Representtive photomicrogrphs of kidney sections from helthy highlnd-plteu yks in three ge groups (neworn [ to 7 dys old; A nd D], dult [3 to 6 yers old; B nd E], or old [7 to yers old; C nd F]) illustrting the loclistion of one morphogenetic protein- (BMP-) (A through F). Immunohistochemicl stining ws used to identify BMP-, which ppers in yellow. Br = µm. Inserts in pnels A through F: Higher-mgnifiction views of the positive expression of BMP- in the kidney. Br = µm; C cortex; M medull. See Figure for the reminder of the key. 8

6 T. Wng et l., Distriution of EGFR, BMP-, nd p53 in yk kidneys Figure 5. Representtive photomicrogrphs of kidney sections from helthy highlnd-plteu yks in three ge groups (neworn [ to 7 dys old; A nd D], dult [3 to 6 yers old; B nd E], or old [7 to yers old; C nd F]) illustrting the loclistion of p53 (A through F). Immunohistochemicl stining ws used to identify p53, which ppers in yellow. Br = µm. Inserts in pnels A through F. Higher-mgnifiction views of the positive expression of p53 in the kidney. Br = µm; C cortex; M medull. See Figures nd for the reminder of the key. Mesurements of immunohistochemistry Opticl density vlues of EGFR in the kidney of the yk Different regions. EGFR positive responses were different in different regions of the sme ge group. In the neworn group, the positive expression of EGFR in the medull ws significntly higher thn the cortex nd the junction of cortex nd medull (Fig. 6A, p <.5); In the dult group, the positive expression of EGFR in the junction of the cortex nd medull ws lower thn the cortex nd medull; In the old group, the positive expression of EGFR in the medull of the old group ws lower thn the cortex nd the junction of cortex nd medull. There ws no significnt difference etween the dult nd the old groups in three regions (Fig. 6A, p >.5). Different ge groups. With ge, the positive expression of EGFR in the cortex nd the junction of cortex nd medull of the neworn group ws significntly lower thn the dult group nd the old group (Fig. 6B, p <.5), while the EGFR positive expression in medull were the lrgest in ged, followed y the differences in the neworn, with those in the dult eing the smllest nd there were significnt differences etween the neworn nd the dult group nd the dult nd old ge groups (Fig. 6B, p <.5). Opticl density vlues of BMP- in the kidney of the yk Different regions. The positive expression of BMP- in the cortex of the neworn nd the old group ws higher thn the junction of cortex nd medull nd the medull, nd there ws no significnt difference etween the three regions (Fig. 7A, p >.5). The positive expression of BMP- in the medull of the dult group ws significntly lower thn the cortex nd the junction of cortex nd medull (Fig. 7A, p <.5). Different ge groups. The expression of BMP- positive expression in cortex, the junction of cortex nd medull nd medull were incresed with ge. The positive expression in the cortex nd medull of the old group ws significntly different from the neworn group nd the dult group (Fig. 7B, p <.5). However, there ws no significnt difference etween the different ge groups of the junction of cortex nd medull (Fig. 7B, p >.5). Opticl density vlues of p53 in the kidney of the yk Different regions. The positive expression of p53 in the cortex of the neworn group ws higher thn the junction of cortex nd medull nd the medull. The positive expression of p53 in the medull of the 9

7 Foli Morphol., 9, Vol. 78, No. A B Opticl density vlues of EGFR/ 5 µm C JCM M Neworn Adult Old C JCM M Opticl density vlues of EGFR/ 5 µm Neworn Adult Old Figure 6. The opticl density vlue distriution of epiderml growth fctor receptor (EGFR) in the different regions of the kidney of sme ge groups of yks (Fig. 4A). The opticl density vlue distriution of EGFR in the sme regions of the kidney of different ge groups of yks (Fig. 4B); C cortex; JCM junction of cortex nd medull; M medull. Dt re reported s men ± stndrd devition. Men vlues with different letters re significntly different (p <.5) nd sme letters re not significntly different (p >.5). Not significntly; Significntly different. A B Opticl density vlues of BMP-/ 5 µm C JCM M Neworn Adult Old C JCM M Opticl density vlues of BMP-/ 5 µm Neworn Adult Old Figure 7. The opticl density vlue distriution of one morphogenetic protein- (BMP-) in the different regions of the kidney of sme ge groups of yks (Fig. 6A). The opticl density vlue distriution of BMP- in the sme regions of the kidney of different ge groups of yks (Fig. 6B); C cortex; JCM junction of cortex nd medull; M medull. Dt re reported s men ± stndrd devition. Men vlues with different letters re significntly different (p<.5) nd sme letters re not significntly different (p >.5). Not significntly; Significntly different. A B Opticl density vlues of p53/ 5 µm C JCM M Neworn Adult Old C JCM M Opticl density vlues of p53/ 5 µm Neworn Adult Old Figure 8. The opticl density vlue distriution of p53 in the different regions of the kidney of sme ge groups of yks (Fig. 8A). The opticl density vlue distriution of p53 in the sme regions of the kidney of different ge groups of yks (Fig. 8B); C cortex; JCM junction of cortex nd medull; M medull. Dt re reported s men ± stndrd devition. Men vlues with different letters re significntly different (p <.5) nd sme letters re not significntly different (p >.5). Not significntly; Significntly different. dult group ws higher thn the cortex nd the junction of cortex nd medull. The positive expression of p53 in the junction of cortex nd medull of the old group ws higher thn the cortex nd medull. Different regions mong the three groups were not significnt (Fig. 8A, p >.5). Different ge groups. The positive expression of p53 in the cortex of the old group ws higher

8 T. Wng et l., Distriution of EGFR, BMP-, nd p53 in yk kidneys Opticl density vlues of ntiody/ 5 µm Neworn Adult Old Figure 9. The opticl density vlues of epiderml growth fctor receptor (EGFR), one morphogenetic protein- (BMP-) nd p53 in the kidney of different ge groups of yks. Dt re reported s men ± stndrd devition. Men vlues with different letters re significntly different (p <.5) nd sme letters re not significntly different (p >.5). Not significntly; Significntly different. thn the dult nd the neworn, nd there ws no significnt difference etween the three ge groups (Fig. 8B, p >.5). The positive expression of p53 in the junction of cortex nd medull nd medull of the neworn group ws significntly lower thn the dult nd the old (Fig. 8B, p <.5). Comprison of the opticl density vlues of EGFR, BMP- nd p53 in different ge of the yk kidneys The totl positive expression of EGFR nd BMP- incresed with ge, while the totl positive expression of p53 first incresed nd then decresed. The totl positive expression of EGFR nd p53 in the neworn ws significntly different from the dult nd the old (Fig. 9, p <.5); the totl positive expression of BMP- in the old ws significntly different from the neworn nd the dult (Fig. 9, p <.5). DISCUSSION EGFR BMP- p53 In this study, the tissue structure of yk kidney ws demonstrted y Msson s trichrome (niline lue) stin nd AB-PAS/Msson s trichrome (niline lue) doule stins. The results showed tht the kidney of yks ws like humn [8], mouse [4], rit [], Bctrin cmels [36] nd Africn white rhinoceros [6]. This result provided reference for the histologicl chnges of cliniclly norml yk kidney. Histologicl chrcteristics of postntl dy rt kidney show tht the most glomeruli, prticulrly in the outer cortex re immture ppering. Tuules re not fully mture [3]. This study found tht the kidney structure of neworn yk ws not fully developed, which ws consistent with the previous reserch. However, we lso found tht there ws no intrinsic chnge in the internl tissue structure of yk kidney, ut the development degree of the glomerulr, renl tuule nd the collection tuule hd chnged. This result is consistent with the studies of the humn kidney [8, 34]. It showed tht the kidney structure tended to e completed with ge, nd the function of the kidney grdully incresed [7]. Epiderml growth fctor receptor positive rections were widely distriuted in the kidneys, including mesngil cells, proximl tuule, nd corticl nd inner medullry collecting duct, s well s in medullry interstitil cells []. We found tht the EGFR positive rection ws minly distriuted in the proximl tuule epithelium of the neworn yk nd were widely distriuted in the dult nd old yk kidneys. Our results were not consistent with the ove reserchers. It indicted tht the function of the nephron of the neworn yk ws not fully developed nd its function ws not comprehensive. However, EGFR ws present in the kidney efore irth nd ws involved in cell prolifertion nd differentition. In the neworn, the proximl tuules re developed nd mture nd the existence of EGFR is to mintin the integrity of the mture tuule epithelium []. BMP- ws essentil for erly emryonic development [39]. In vertertes, BMP genes were multi-effect growth/differentition fctors nd expressed in mny emryonic orgns nd tissues, including epithelil nd mesenchyml cells [6]. Previous studies found tht BMP- ws highly expressed in the hert myocrdil lyer nd uterine epithelil cells of the mice [3]. Up to now, there is no relevnt literture on BMP- expression in the kidney. Our study found tht the positive rection of BMP- in the neworn group ws minly in the proximl tuule epithelium nd tht in the dult group nd the old group they were widely distriuted in the kidney. Therefore, we speculte tht BMP- my hve role in promoting kidney development. Molchdsky et l. [9] found tht p53 expression is uiquitous erly in mouse emryogenesis, fter which time expression is restricted to specific tissues during orgnogenesis. The study found tht p53 positive rection ws widely distriuted in the neworn yk, suggesting tht p53 ws involved in cell migrtion, metolism, differentition, nd self-renewl during this period [9, 3].

9 Foli Morphol., 9, Vol. 78, No. The results of the verge opticl density vlues of the kidney cortex, the junction of cortex nd medull nd medull of different ge groups indicte tht the positive expression of EGFR in the cortex nd the junction of cortex nd medull incresed with ge, while the positive expression of EGFR in the medull decresed first nd then incresed. The results indicte tht EGFR promoted kidney development [3]. EGFR hs existed from emryonic stge, nd the tissues derived from endoderm nd ectoderm cn promote mitosis nd stimulte the synthesis nd metolism, nd stimulte the prolifertion of epiderml cells, epithelil cells, interstitil cells nd vsculr endothelil cells. Therefore, there ws chnge in the degree of epithelil cell integrity with ge, nd there ws n incrementl chnge in the cortex nd the junction of the cortex nd medull, nd the EGFR content in the tuules of the medull were reduced fter irth. With ge, EGF inds to the receptor to form dimer, which ctivtes the tyrosine kinse receptor in the receptor cell. The enzyme receptor itself or sustrte phosphoryltion on tyrosine residues, result in series of downstrem signl protein nd enzyme phosphoryltion (ctivtion). By mens of MAPK, the prolifertion signl ws introduced into the nucleus, which eventully cused the cell to enter the S stge, leding to cell prolifertion nd incresing the expression of EGFR [4]. In ddition, some results suggested tht BMP- could promote emryonic development. In this study, the positive expression of BMP- in the cortex, medull nd the junction of cortex nd medull incresed with ge. Therefore, it is speculted tht BMP- hs een involved in the prolifertion nd differentition of cells in the kidney t the eginning of the emryonic period nd promoted the development of the kidneys [7]. At the sme time, the study showed tht positive expression of p53 in cortex incresed with ge, while positive expression of p53 in the medull nd the junction of cortex nd medull first incresed nd then decresed with ge. The results indicted tht the development of the cortex, the junction of cortex nd medull nd medull in the kidney ws different, nd the p53 showed different positive expression chnges with the development degree of ech structure [9]. Finlly, the results lso demonstrted tht the totl positive expression of EGFR nd BMP- incresed with ge nd the totl positive expression of p53 decresed with ge. Previous literture confirms tht EGFR nd BMP- re expressed in orgns such s kidneys [6] nd uterus [3] nd incresed expression with ge. Therefore, we hypothesize tht EGFR nd BMP- hve the effect of promoting kidney development. He et l. [] found tht p53 protein expression levels decresed in the kidneys of rts with ge, which ws consistent with our results, indicting tht p53 ws key regultory fctor in the development of kidney. CONCLUSIONS The results reveled tht the development of the kidney of the yk internl structure chnged with ge, indicting tht the yk kidney structure tended to complete with ge nd the function of kidneys grdully improved. EGFR nd BMP- positive rection in the neworn group ws minly distriuted in the proximl tuule epithelil cells, nd widely distriuted in the dult nd old groups, indicting tht EGFR nd BMP- hd the effect of promoting kidney development. However, p53 hd een widely distriuted in neworn kidney of the yks. It is suggested tht p53 hd een involved in cell migrtion nd metolic differentition nd self-renewl in the new stge. Acknowledgements This reserch ws supported y The Ntionl Nturl Science Foundtion of Chin (grnt No ). REFERENCES. Attisno L, Wrn JL. Mds nd Smds in TGF et signlling. Curr Opin Cell Biol. 998; (): 88 94, indexed in Pumed: Buchet AL, Msson R, Guffroy M, et l. Immunohistochemicl identifiction of kidney nephron segments in the dog, rt, mouse, nd cynomolgus monkey. Toxicol Pthol. ; 39(7): 5 8, doi:.77/ , indexed in Pumed: Bserg M, Hle MA, Ke X, et l. Uteroplcentl insufficiency increses p53 phosphoryltion without triggering the p53- MDM functionl circuit response in the IUGR rt kidney. Am J Physiol Regul Integr Comp Physiol. 6; 9(): R4 R48, doi:.5/jpregu.88.5, indexed in Pumed: Cirdiello F, Tortor G. EGFR ntgonists in cncer tretment. N Engl J Med. 8; 358(): 6 74, doi:.56/ NEJMr7774, indexed in Pumed: El-Dhr S, Hillird S, Aoudehen K, et l. The MDM-p53 pthwy: multiple roles in kidney development. Peditr Nephrol. 4; 9(4): 6 67, doi:.7/s y, indexed in Pumed: Gttone VH, Shermn DA, Hinton DA, et l. Epiderml growth fctor in the neontl mouse slivry glnd nd kidney. Biol Neonte. 99; 6(): 54 67, doi:.59/4353, indexed in Pumed: Gekle M. Kidney nd ging: nrrtive review. Exp Gerontol. 7; 87(Pt B): 53 55, doi:.6/j.exger.6.3.3, indexed in Pumed:73877.

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