大鼠生后發(fā)育期間胰島D細(xì)胞的胰島淀粉樣多肽和生長抑素的表達(dá)

1、    大鼠生后發(fā)育期間胰島D細(xì)胞的胰島淀粉樣 多肽和生長抑素的表達(dá)        【摘要】目的觀察大鼠生后發(fā)育期間胰島淀粉樣多肽(islet amyloid polypeptide,IAPP)與生長抑素(somatostatin,SS)在胰島D細(xì)胞的表達(dá)。方法免疫組織化學(xué)PAP鄰片單染法。結(jié)果從生后1d至成年大鼠胰島內(nèi)均有部分IAPP-IR細(xì)胞和SS-IR細(xì)胞形態(tài)相似、定位一致；外分泌部腺泡間散在分布的部分IAPP-IR細(xì)胞也呈SS免疫反應(yīng)性。結(jié)論大鼠生后發(fā)育過程

2、中胰島部分D細(xì)胞同時表達(dá)IAPP和SS，即兩者共存于胰島D細(xì)胞內(nèi)。【關(guān)鍵詞】胰島淀粉樣多肽生長抑素表達(dá)生后發(fā)育大鼠 IMMUNOHISTOCHEMICAL STUDY ON THE EXPRESSION OF IAPP AND SS IN D CELL OF PANCREAS ISLET OF POSTNATAL DEVELOPING RATGuo Shuangping,Shi Airong*,Wang Xianrong,Zhai Yuqiang,Huang Yan*(Department of Histology and Embryology,Lanzou Medical College,L

3、anzhou;*Department of Histology and Embryology,Beijing Medical University,Beijing)【Abstract】Objective To observe the expression of islet amyloid polypeptide (IAPP) and somatostatin(SS) in islet D cells of postnatal developing rats.MethodsImmunohistochemical PAP method was used to show IAAPIR(immunor

4、eactivity，IR) cells and SSIR cells in islets on adjacent sections of postnatal rats.ResultsSome islet IAPPIR cells were identically localized to SSIR cells on adjacent sections of postanatal developing rats from the first day after birth to adult.Some IAPPIR cells distributed in exocrine pancreas we

5、re also showed SS immunoreactivity.ConclusionSome islet D cells of postnatal developing rats expressed both IAPP and SS simultaneously,the two kinds hormones coexisted in D cells.【Key words】 IAPP;SS;Expression;Postanatal development;Rat胰島淀粉樣多肽(IAPP)是胰島B細(xì)胞合成和分泌的一種調(diào)節(jié)糖代謝的激素13，IAPP與胰島素(insulin,Ins)共存于胰島

6、B細(xì)胞的分泌顆粒內(nèi)，兩者常相伴釋放4。IAPP作為一種自分泌或旁分泌激素可調(diào)節(jié)Ins的分泌5，與型糖尿病的發(fā)生密切相關(guān)。IAPP與Ins共存在B細(xì)胞中的研究已較深入，但I(xiàn)APP與生長抑素(SS)共存的報(bào)道較少，尤其是大鼠生后發(fā)育過程中IAPP在胰島D細(xì)胞的表達(dá)尚未見系統(tǒng)報(bào)道。本實(shí)驗(yàn)用免疫組化法研究了大鼠生后發(fā)育期間IAPP與SS在胰島D細(xì)胞的表達(dá)，旨在為探討IAPP的生理功能提供更多的形態(tài)學(xué)依據(jù)。材料和方法1.動物分組、取材和標(biāo)本制備正常Wistar大鼠共30只，由北京醫(yī)科大學(xué)動物實(shí)驗(yàn)中心提供，分為生后1d,7d,18d,28d,45d和10月齡成年大鼠6組，每組5只。體重分別為(6.5

7、77;0.5)g、(15±2)g、(36.4±2.1)g、(58.5±3.8)g、(131.1±9.5)g、(177±10)g。取材前動物禁食24h，常規(guī)石蠟包埋，制成4m厚連續(xù)切片，各組的每只動物取1張裱于同一張載玻片上，按順序編號，取相鄰切片分別染IAPP和SS。2.免疫組織化學(xué)法采用Sternberger PAP法，主要染色步驟為：(1)切片常規(guī)脫蠟入水，甲醇-H2O2封閉，相鄰切片分別滴加SS抗體(11 500)和IAPP抗體(14 000)，4孵育過夜；(2)滴加羊抗兔IgG(150)，37孵育40min;(3)滴加PAP復(fù)合物(11

8、00)，37孵育40min。(以上抗體均由北京醫(yī)科大學(xué)石愛榮教授提供)。(4)滴加DAB-H2O2液顯色、蘇木精復(fù)染、脫水、透明、封固。對照組以PBS代替1抗。結(jié)果和討論光鏡下觀察，免疫染色物質(zhì)均位于細(xì)胞質(zhì)中，胞核內(nèi)無免疫染色物，對照組為陰性。生后1d大鼠胰島內(nèi)就可見IAPP-IR細(xì)胞和SS-IR細(xì)胞。IAPP-IR細(xì)胞多為圓形或橢圓形，呈索狀排列于胰島中央及周邊。SS-IR細(xì)胞較少，多為不規(guī)則形，散在分布于胰島周邊部。在外分泌部腺泡之間及導(dǎo)管上皮之間，也有散在的IAPP-IR細(xì)胞和SS-IR細(xì)胞。同時觀察到生后1d和7d大鼠胰島細(xì)胞分裂像多見，隨著大鼠生后發(fā)育細(xì)胞分裂像漸減少。觀察IAPP和

9、SS免疫染色的鄰片發(fā)現(xiàn)，生后發(fā)育不同時期大鼠胰腺組織內(nèi)，均可見形態(tài)和分布一致的IAPP-IR細(xì)胞和SS-IR細(xì)胞，即同一細(xì)胞同時表達(dá)IAPP和SS。在生后1d大鼠胰腺組織中，可觀察到部分SS-IR細(xì)胞與鄰片同一胰島的IAPP-IR細(xì)胞形態(tài)相似、定位一致(1)。在生后7d大鼠胰腺組織切片中，胰島部分D細(xì)胞同時表達(dá)IAPP和SS(2)。在生后18d大鼠胰腺組織切片中，觀察到部分定位一致的D細(xì)胞同時表達(dá)IAPP和SS(3)。在生后28d大鼠胰腺組織切片中，也有部分D細(xì)胞同時表達(dá)IAPP和SS(4)。在生后45d大鼠胰腺組織切片中，部分D細(xì)胞同時表達(dá)IAPP和SS(5)。在成年大鼠胰腺組織切片中，也可

10、觀察到胰島部分D細(xì)胞同時表達(dá)IAPP和SS?？傊?，在大鼠生后發(fā)育期間，部分D細(xì)胞同時表達(dá)IAPP和SS，兩者共存于D細(xì)胞。IAPP分離提純以來，關(guān)于其生化特性、組織定位、基因表達(dá)、受體分布和生理作用都進(jìn)行了深入研究13，認(rèn)為它主要由胰島B細(xì)胞合成和分泌，與胰島的其他激素協(xié)同作用，調(diào)節(jié)機(jī)體的代謝和生長發(fā)育4。近年的研究發(fā)現(xiàn)，IAPP不僅與Ins共存于胰島B細(xì)胞，而且與SS共存于胰島D細(xì)胞。De Vroede6等發(fā)現(xiàn)培養(yǎng)的大鼠胰島D細(xì)胞呈IAPP免疫反應(yīng)性。國內(nèi)也有報(bào)道IAPP與SS共存于正常大鼠胰島D細(xì)胞7。本實(shí)驗(yàn)觀察了大鼠生后發(fā)育過程中IAPP與SS在胰島D細(xì)胞的共存關(guān)系，從大鼠生后1d起直至

11、成年都有胰島部分D細(xì)胞同時表達(dá)IAPP和SS，即兩者共存于胰島D細(xì)胞。關(guān)于IAPP與SS在胰島D共存的意義尚不清楚。在生理情況下，胰島D細(xì)胞分泌的IAPP僅占胰島分泌IAPP總量的很少一部分，但在鏈佐霉素誘導(dǎo)的糖尿病大鼠，胰島B細(xì)胞被選擇性破壞。此時，IAPP的分泌主要依賴于胰島D細(xì)胞，IAPP與SS在胰島D細(xì)胞中的共存與糖尿病相關(guān)6。本實(shí)驗(yàn)觀察了在大鼠生后發(fā)育過程中胰島D細(xì)胞同時表達(dá)IAPP和SS，而SS可抑制IAPP的釋放8，SS對胰腺及胃腸組織的生長起抑制作用，是促進(jìn)組織分化的信號9。因此，我們認(rèn)為在大鼠生后發(fā)育期間胰島D細(xì)胞同時表達(dá)IAPP和SS可能與胰島的生后發(fā)育有關(guān)。1生后1d大鼠

12、胰腺相鄰連續(xù)切片顯示D細(xì)胞同時表達(dá)IAPP(1A箭頭)與SS(1B箭頭)×400Fig.1Photomicrograph demonstrated the expression of IAPP(arrow,1A)and SS(arrow,1B)in pancreas islet D cell of rat on the first day after birth. ×4002生后7d大鼠胰腺相鄰連續(xù)切片顯示D細(xì)胞同時表達(dá)IAPP(2A箭頭)與SS(2B箭頭)×400Fig.2 Photomicrograph demonstrated the expression

13、of IAPP(arrow,2A)and SS(arrow,2B)in pancreas islet D cell of rat on the 7th day after birth. ×4003生后18d大鼠胰腺相鄰連續(xù)切片顯示D細(xì)胞同時表達(dá)IAPP(3A箭頭)與SS(3B箭頭)×400Fig.3 Photomicrograph demonstrated the expression of IAPP(arrow,3A)and SS(arrow,3B)in pancreas islet D cell of rat on the 18th day after birth. &

14、#215;4004生后28d大鼠胰腺相鄰連續(xù)切片顯示D細(xì)胞同時表達(dá)IAPP(4A箭頭)與SS(4B箭頭)×400Fig.4 Photomicrograph demonstrated the expression of IAPP(arrow,4A)and SS(arrow,4B)in pancreas islet D cell of rat on the 28th day after birth. ×4005生后45d大鼠胰腺相鄰連續(xù)切片顯示D細(xì)胞同時表達(dá)IAPP(5A箭頭)與SS(5B箭頭)×400Fig.5 Photomicrograph demonstrate

15、d the expression of IAPP(arrow,5A)and SS(arrow,5B)in pancreas islet D cell of rat on the 45th day after birth. ×400作者單位：郭雙平石愛榮?王先榮翟宇強(qiáng)黃巖蘭州醫(yī)學(xué)院組織學(xué)與胚胎學(xué)教研室，蘭州730000；*北京醫(yī)科大學(xué)組織學(xué)與胚胎學(xué)系參考文獻(xiàn)1Westermark P,Wernstedt C,Wllanddet E,et al.A novel peptide in the calcitonin related peptide family as an amyloid f

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17、cat are derived from a neuropeptidelike protein also present in normal islet cells.Proc Natl Acad Sci USA,1987,84(11):38814Hartter E,Svoboda T,Ludvik B,et al.Basal and stimulated plasma levels of pancreatic amylin indicate it cosecretion with insulin in human.Diabetologia,1991,34(1):525Bennet WM,Vbe

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