馮　濤，吳曉敏，許曉玲，白佳樺，宋玉清，肖霖力，韓向敏，劉　彥*（.北京市農(nóng)林科學(xué)院畜牧獸醫(yī)研究所，北京　00097；.甘肅農(nóng)業(yè)大學(xué)動物科學(xué)技術(shù)學(xué)院，蘭州　730070）

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GT1-7細(xì)胞及其在生殖相關(guān)研究中的應(yīng)用

馮濤1，吳曉敏2，許曉玲1，白佳樺1，宋玉清1，肖霖力1，韓向敏2，劉彥1*
（1.北京市農(nóng)林科學(xué)院畜牧獸醫(yī)研究所，北京100097；2.甘肅農(nóng)業(yè)大學(xué)動物科學(xué)技術(shù)學(xué)院，蘭州730070）

摘要：GT1-7細(xì)胞是GT1細(xì)胞株的亞株，通過轉(zhuǎn)基因技術(shù)從小鼠下丘腦分離獲得的GnRH神經(jīng)元細(xì)胞系，具有高度分化的神經(jīng)內(nèi)分泌細(xì)胞典型特征。下丘腦GnRH合成和釋放對生殖功能具有重要作用，而GnRH神經(jīng)元在腦內(nèi)數(shù)量少且呈彌散分布，體內(nèi)研究較困難。目前，GT1-7細(xì)胞是研究GnRH神經(jīng)元的理想離體細(xì)胞模型，在生殖相關(guān)研究中廣泛使用。文章詳細(xì)闡述GT1-7細(xì)胞獲得過程、功能特性及其在生殖系統(tǒng)研究中應(yīng)用，以及影響GT1-7細(xì)胞活性的信號通路，以期對動物生殖調(diào)控研究提供參考。

關(guān)鍵詞：GT1-7細(xì)胞；促性腺激素釋放激素；生殖調(diào)控；信號通路

馮濤,吳曉敏,許曉玲,等. GT1-7細(xì)胞及其在生殖相關(guān)研究中的應(yīng)用[J].東北農(nóng)業(yè)大學(xué)學(xué)報, 2016, 47(1): 102-108.

Feng Tao, Wu Xiaomin, Xu Xiaoling, et al. GT1-7 cells and its application in reproduction-related researches[J]. Journal of Northeast Agricultural University, 2016, 47(1): 102-108. (in Chinese with English abstract)

雌性動物生殖活動受下丘腦-垂體-性腺軸（Hypothalamus-pituitary-gonad，HPG）精密調(diào)控，其中下丘腦促性腺激素釋放激素（Gonadotropin-re?leasing hormone，GnRH）神經(jīng)元是動物發(fā)情周期啟動關(guān)鍵部位，GnRH分泌調(diào)控是深入揭示動物情期啟動關(guān)鍵因素。近年來，分子遺傳學(xué)得到快速發(fā)展，特別是GnRH神經(jīng)元離體細(xì)胞模型—GT1-7細(xì)胞的發(fā)現(xiàn)，加速生殖內(nèi)分泌調(diào)控領(lǐng)域研究進程。永生GT1-7細(xì)胞是從轉(zhuǎn)基因小鼠下丘腦視前區(qū)分離到的GnRH神經(jīng)元細(xì)胞，具有高度分化神經(jīng)內(nèi)分泌細(xì)胞特性，能模擬GnRH神經(jīng)元功能。GT1-7細(xì)胞不但具有原始GnRH神經(jīng)元多種功能，如節(jié)律性脈沖分泌GnRH，多種信號通路調(diào)節(jié)細(xì)胞活性，能表達GnRH、Kiss-1、G蛋白偶聯(lián)受體54（GPR54）、雌激素受體α（ERα）、雌激素受體β（ERβ）及卵泡抑制素樣蛋白等多種生殖相關(guān)基因、受體及其蛋白，在動物生殖調(diào)控研究中具有重要作用。然而，GnRH神經(jīng)元作為哺乳動物中樞生殖調(diào)控體系最終共同通路，GnRH合成和分泌受諸多因子及神經(jīng)元相互作用調(diào)控，因此利用GT1-7細(xì)胞完全模擬GnRH體內(nèi)調(diào)控存在局限性。本文從GT1-7細(xì)胞來源和特性及其在生殖相關(guān)研究中應(yīng)用等方面綜述，總結(jié)影響GT1-7細(xì)胞活性信號通路，以期對動物生殖調(diào)控研究提供參考。

1 GT1-7細(xì)胞來源

GT1-7細(xì)胞是GT1細(xì)胞株1個亞株。GT1細(xì)胞株由美國Mellon等于1990年利用遺傳學(xué)靶向致腫瘤技術(shù)[1]，將致癌猿猴空泡病毒40（Simian vacuolat?ing virus，SV40）抗原（Tag）基因及其在GnRH細(xì)胞中表達調(diào)控基因構(gòu)建雜合基因后，整合到小鼠受精卵原核中，并將從轉(zhuǎn)基因小鼠子代下丘腦視前區(qū)分離得到腫瘤細(xì)胞經(jīng)進一步篩選和克隆，獲得同時表達GnRH和Tag基因穩(wěn)定細(xì)胞株。GT1細(xì)胞株有3個亞株，分別是GT1-1、GT1-3和GT1-7，均具有高度分化GnRH神經(jīng)元特征。GT1-7細(xì)胞在表型形態(tài)、超微結(jié)構(gòu)、基因表達、激素儲存以及分泌方式上均與GnRH神經(jīng)元一致，且能在體外分裂傳代，是體外研究GnRH神經(jīng)元的理想細(xì)胞模型[2]。

2 GT1-7細(xì)胞特性

GnRH神經(jīng)元是哺乳動物中樞生殖調(diào)控體系最終共同通路，僅分布于前腦嗅區(qū)、隔區(qū)及下丘腦等區(qū)域。下丘腦是由許多功能各異神經(jīng)核團組成結(jié)構(gòu)復(fù)雜器官，體內(nèi)精確研究GnRH神經(jīng)元及相關(guān)功能難度較大，通過原代培養(yǎng)方法直接研究下丘腦GnRH神經(jīng)元功能也存在很大困難。因此，目前研究多利用下丘腦永生GnRH神經(jīng)元細(xì)胞系GT1-7細(xì)胞開展相關(guān)研究。

2.1 GT1-7細(xì)胞分泌模式

GT1-7細(xì)胞表現(xiàn)GnRH神經(jīng)元形態(tài)，且保留許多原始GnRH神經(jīng)元功能[3-4]。Martinez等研究發(fā)現(xiàn)，GT1-7細(xì)胞在體外表現(xiàn)出與GnRH神經(jīng)元類似脈沖分泌模式，下丘腦GnRH神經(jīng)元分泌GnRH平均脈沖間隔25.8 min，平均持續(xù)時間18.8 min[5]。GT1-7細(xì)胞分泌GnRH平均脈沖間隔24.8~45.4 min，平均脈沖幅度14.30~191.5 pg·min·mL-1，平均脈沖持續(xù)時間為21.30 min[4-8]。可見GT1-7細(xì)胞與GnRH神經(jīng)元細(xì)胞脈沖情況基本一致，細(xì)微差異可能與培養(yǎng)液種類、細(xì)胞系傳代次數(shù)以及培養(yǎng)時間等條件有關(guān)。

2.2時鐘基因?qū)T1-7細(xì)胞分泌調(diào)節(jié)

GnRH神經(jīng)元細(xì)胞含有分子時鐘，晝夜信號可有效分配到多個組織[9]。研究表明，GT1-7細(xì)胞能表達多種時鐘基因，包括Bmal1、Clock、mCry1、mCry2、mPer1、mPer2、mPer3和CKIε，這些基因也在下丘腦視交叉上核（SCN）和外周細(xì)胞中表達[10]。在GT1-7細(xì)胞中用顯性失活Clock-Δ19基因瞬時表達能擾亂生物鐘功能進而破壞GnRH正常晝夜分泌模式，顯著降低平均脈沖頻率；過表達負(fù)調(diào)控Limb基因mCry1可顯著增加GnRH脈沖幅度，而對脈沖頻率無顯著變化[11]。Eileen等將GT1-7細(xì)胞暴露于0.1 U·mL-1凝血酶（誘導(dǎo)Ca2+內(nèi)流）也可顯著增加脈沖振幅，但對持續(xù)時間影響不顯著，推測可能與GT1-7細(xì)胞特定膜結(jié)合受體及細(xì)胞膜上時鐘基因有關(guān)[7]。Blackman等研究表明，GT1-7細(xì)胞內(nèi)cAMP水平通過環(huán)核苷酸門控通道（CNGs）參與內(nèi)源性生物鐘調(diào)節(jié)，影響GnRH分泌[12]。可見，時鐘基因表達能夠調(diào)節(jié)GT1-7細(xì)胞分泌活性，但并不改變脈沖式釋放GnRH固有特性。

2.3離子通道對GT1-7細(xì)胞分泌調(diào)節(jié)

GT1-7細(xì)胞作為神經(jīng)內(nèi)分泌細(xì)胞，其分泌激素與細(xì)胞膜電位、胞內(nèi)鈣離子濃度和興奮性等有關(guān)[13]。GT1-7細(xì)胞能表達多種質(zhì)膜通道，有自發(fā)動作電位能力[14]：鈉通道阻滯劑河豚毒素（TTX）敏感Na+通道[1]，三種不同類型向外K+通道和一個K+向內(nèi)整流器[15-16]以及電壓依賴性Ca2+通道[15,17]。GT1-7細(xì)胞自發(fā)動作電位頻率24.8 min[18]與GnRH神經(jīng)元平均脈沖頻率25.8 min[5]基本一致，因此動作電位是GnRH釋放機制一部分。GnRH分泌和鈣振蕩是“GnRH脈沖發(fā)生器”基礎(chǔ)[19-20]，搏動機制是Ca2+周期性交替[21]。鈣振蕩與Na+通道動作電位和Ca2+通過電壓門控鈣通道有關(guān)[18]。K+通道阻滯劑能引起幅度或鈣振蕩頻率明顯變化[21]。GT1-7細(xì)胞能夠表達R、L、N和T型鈣通道[14]，其中R型亞基起關(guān)鍵作用，作為主要電流調(diào)節(jié)通道與L、N和T型鈣通道共同調(diào)節(jié)GnRH釋放[22]。近年來超極化激活環(huán)核苷酸調(diào)節(jié)通道（HCN）成為研究熱點，HCN-在超極化時被激活并產(chǎn)生內(nèi)向電流，目前發(fā)現(xiàn)有四種HCN通道亞型，即HCNl、HCN2、HCN3和HCN4，且四種亞型在GTl-7細(xì)胞上均有表達[23]。

Wetsel等研究發(fā)現(xiàn)，GT1-7細(xì)胞能表達縫隙連接蛋白，連接素26樣蛋白[24]，及突觸樣連接蛋白[25]。目前研究發(fā)現(xiàn)GT1-7細(xì)胞分泌GnRH主要是分子生物學(xué)和電生理機制，但精確脈沖機制尚未確定，可能還包括經(jīng)間隙連接旁分泌因子，例如一氧化氮、電生理耦合或GnRH及從GnRH前體分子加工而得到肽類物質(zhì)，如GnRH相關(guān)肽（GAP）等。

3 GT1-7細(xì)胞在生殖相關(guān)研究中的應(yīng)用

下丘腦接受中樞神經(jīng)系統(tǒng)發(fā)布信息，通過GnRH神經(jīng)元分泌GnRH，GnRH調(diào)節(jié)垂體中促性腺激素細(xì)胞分泌促卵泡素（FSH）和黃體生成素（LH），促性腺激素（FSH，LH）作用于性腺（雄性睪丸和雌性卵巢），調(diào)節(jié)性激素分泌并影響生殖活動。垂體分泌促性腺激素可反饋調(diào)節(jié)下丘腦GnRH分泌；性腺激素也可反饋調(diào)節(jié)下丘腦和垂體相應(yīng)激素釋放。因此，在下丘腦、垂體和性腺之間形成密切相連軸線系統(tǒng)，即HPG軸。體外GT1-7細(xì)胞不但能表現(xiàn)出與GnRH神經(jīng)元類似脈沖分泌模式，還有GnRH、Kiss-1、GPR54、BMP受體、ERα、ERβ、抑制性Smads以及卵泡抑制素樣蛋白等表達[26-27]。目前認(rèn)為，GT1-7細(xì)胞是體外研究HPG軸調(diào)控機制的理想細(xì)胞模型。

3.1 GT1-7細(xì)胞與Kisspeptin/GPR54

Kisspeptin是Kiss-1基因編碼神經(jīng)內(nèi)分泌肽類激素，為GPR54內(nèi)源性配體[28]，可直接作用于下丘腦GnRH神經(jīng)元，促進GnRH分泌，激活HPG軸。Kiss-1/GPR54基因突變可導(dǎo)致特發(fā)性促性腺激素分泌不足型腺機能減退癥（Idiopathic hypogo?nadotropic hypogonadism, IHH）[29]，因此Kisspeptin/ GPR54被認(rèn)為是青春期發(fā)育啟動“分子閥門”。Kis?speptin處理GT1-7細(xì)胞，以濃度依賴性方式增加GnRH mRNA表達及GnRH分泌[27]。Ozcan等研究表明，Kisspeptin可使GT1-7細(xì)胞過表達GPR54，激活cAMP/PKA信號通路[30]，誘導(dǎo)GnRH受體表達。同時，Kisspeptin在Kisspeptin/GPR54系統(tǒng)保護機制下升高細(xì)胞內(nèi)鈣水平，通過Ca2+/PKC信號調(diào)控GnRH分泌[30]。Terasaka等通過RT-PCR證實Kiss?peptins/GPR54系統(tǒng)在GT1-7神經(jīng)元細(xì)胞表達，但體內(nèi)GnRH神經(jīng)元并未表達，有GPR54表達[27]。因此，Kisspeptins刺激促性腺激素釋放可能通過GPR54表達實現(xiàn)。Heather等對哺乳動物研究也證實Kisspeptins通過活化GPR54，使GnRH神經(jīng)元表達GPR54，刺激下丘腦GnRH釋放[31]。總之，Kiss?peptin/GPR54是青春期啟動催化劑，進一步研究可以利用GT1-7細(xì)胞為模型探究下丘腦GnRH細(xì)胞如何調(diào)控HPG軸以及青春期啟動等現(xiàn)象。

3.2 GT1-7細(xì)胞與E2/ER

動物體內(nèi)研究表明，溫和地持續(xù)注射雌激素（E2）（50 μg·kg-1，5 d）[32]及大劑量一次性注射雌激素（10 mg·kg-1）[33]，均能誘發(fā)大鼠真性性早熟。給卵巢摘除小鼠連續(xù)外源注射雌激素，可誘導(dǎo)GnRH和LH峰產(chǎn)生[34]。在體外用雌激素處理GT1-7細(xì)胞，pmol級濃度雌激素可對cAMP產(chǎn)生急速、持續(xù)、劑量依賴性抑制，而nmol級濃度雌激素可增加cAMP產(chǎn)生[35]。雖然GT1-7細(xì)胞表達ERα和ERβ，但小鼠GnRH神經(jīng)元表達ERβ，卻不表達ERα[36-37]。有研究報道GT1-7細(xì)胞中雌激素可能通過激活ERβ活性引起GPR54表達[38]。Tonsfeldt等研究還發(fā)現(xiàn)kiss?peptin引起GnRH分泌可能部分依賴雌激素活性，當(dāng)雌激素水平升高時，增加kisspeptin可引起ERα 和ERβ mRNA表達，雌激素也能增加GnRH神經(jīng)元對kisspeptin敏感性，加強GPR54表達[38]。E2能正負(fù)調(diào)控下丘腦GnRH神經(jīng)元，啟動初情期或正常生殖周期，GT1-7細(xì)胞已廣泛用于E2生殖調(diào)控研究。

3.3 GT1-7細(xì)胞與Leptin

瘦素（Leptin）作為脂肪分泌激素，作用于下丘腦，調(diào)節(jié)食物攝入和能量代謝，也是生殖系統(tǒng)重要代謝信號。只有當(dāng)體內(nèi)營養(yǎng)狀態(tài)達到一定臨界水平，足以滿足生殖需要時，才能觸發(fā)動物機體青春期啟動。Leptin對生殖活動調(diào)節(jié)主要通過調(diào)節(jié)下丘腦GnRH分泌實現(xiàn)。因肥胖基因突變而致原發(fā)性瘦素缺失ob小鼠出現(xiàn)肥胖并伴有生殖功能低下，通過單純控制飲食可使體重下降，但不能恢復(fù)生育能力，而外原性瘦素可使其恢復(fù)[39-40]。楊穎等研究表明，GT1-7細(xì)胞表達Leptin受體基因，Leptin能促使GT1-7細(xì)胞迅速釋放GnRH，呈明顯量效關(guān)系，Leptin可能通過直接作用于下丘腦GnRH神經(jīng)元，對生殖功能起調(diào)節(jié)作用[41]。雖然現(xiàn)有研究表明Leptin可直接調(diào)節(jié)GnRH分泌，但是在GnRH神經(jīng)元上并未發(fā)現(xiàn)Leptin受體表達，表明Leptin對GnRH存在中間調(diào)節(jié)機制[42]。在下丘腦視前區(qū)，nNOS神經(jīng)元上存在Leptin受體，NOS酶活性受Leptin調(diào)節(jié)。敲除Leptin基因小鼠（Lepob/ob）不育，在nNOS神經(jīng)元選擇性缺失Leptin可推遲雌鼠初情期啟動，由于阻斷Leptin信號傳導(dǎo)經(jīng)過nNOS細(xì)胞傳至GnRH，nNOS-/-Lepob/ob小鼠在Leptin處理后缺乏LH分泌[43-44]。離體GT1-7細(xì)胞何時啟動能量代謝調(diào)節(jié)，何時啟動生殖機能調(diào)節(jié)，能否系統(tǒng)模擬動物機體生長發(fā)育過程尚不清楚。假設(shè)GT1-7細(xì)胞leptin受體基因缺失，通過提供外原性leptin，觀察GT1-7細(xì)胞GnRH分泌情況，可研究能量對GnRH合成分泌以及生殖機能等影響。

3.4 GT1-7細(xì)胞與褪黑激素

褪黑激素（Melatonin，MLT）是普遍存在于動物機體的吲哚類激素，主要由松果體（Pineal gland，PG）合成和分泌，受光照調(diào)節(jié)。MLT可降低GnRH受體數(shù)量，抑制下丘腦GnRH分泌，主要通過HPG軸調(diào)控動物生殖活動。研究發(fā)現(xiàn)GT1-7細(xì)胞表達G蛋白偶聯(lián)受體褪黑激素受體1（Mt1）和2（Mt2）[45]，抑制cAMP活性，激活PKC和AMPK活性而非PKA活性，抑制GT1-7細(xì)胞GnRH分泌[46]。MLT能提高立早基因c-fos和junB表達水平[46]。Kelestimur等研究結(jié)果表明MLT通過激活PKC增加細(xì)胞內(nèi)Ca2+水平，抑制GT1-7細(xì)胞GnRH釋放[47]。

在青春期與生殖功能調(diào)控研究中，胡雅婷發(fā)現(xiàn)脂聯(lián)素通過AMPK和SP1調(diào)節(jié)GT1-7細(xì)胞Kiss-1基因表達[48]。GT1-7細(xì)胞表達脂聯(lián)素受體1和2[49]、降鈣素基因相關(guān)肽（CGRP）受體[50]、垂體腺苷酸環(huán)化酶激活多肽（PACAP）Ⅰ型受體[51]、神經(jīng)激肽3受體（NKB3R）[52]等與生殖相關(guān)基因。

以上研究表明，GT1-7細(xì)胞與體內(nèi)GnRH神經(jīng)元相似，有多種生殖相關(guān)基因及其受體基因表達，可作為HPG軸調(diào)控機理理想細(xì)胞模型研究機體生殖活動。陳名道等研究中藥補腎方二仙湯及其拆方對GT1-7細(xì)胞分泌GnRH影響[53]，揭示二仙湯及其拆方如何對下丘腦GnRH細(xì)胞發(fā)揮作用，調(diào)控HPG軸。

4影響GT1-7細(xì)胞活性信號通路

4.1 AMPK相關(guān)信號通路

腺苷酸活化蛋白激酶（AMPK），即AMP依賴蛋白激酶，由α、β和γ亞基組成異源三聚體復(fù)合體，是生物機體重要信號分子、細(xì)胞能量關(guān)鍵傳感器和調(diào)制器。AMPK活力主要通過調(diào)節(jié)其與催化相關(guān)結(jié)構(gòu)元件實現(xiàn)。

AMPK通過接受上游信號調(diào)控活力，影響GT1-7細(xì)胞GnRH分泌。研究發(fā)現(xiàn)，GT1-7細(xì)胞中，胰高血糖素樣肽-1受體（GLP-1R）活化后以時間依賴方式減少Akt磷酸化，可能通過PI3K-Akt信號通路調(diào)控AMPK活性，影響食物攝取[54]。在GT1-7細(xì)胞中，α-黑色素細(xì)胞刺激素（α-MSH）通過PKA-AMPK途徑以調(diào)節(jié)能量消耗和食物攝取[55]；葡萄糖轉(zhuǎn)運蛋白2（GLUT2）[56]通過改變細(xì)胞能量狀態(tài)或AMP/ATP比率，調(diào)控食欲相關(guān)肽（AgRP）mRNA表達并使AMPK磷酸化葡萄糖；CGRP通過其受體下調(diào)GnRH mRNA表達[50]，激活cAMP-PKA通路而非cAMP-PKC通路抑制AMPK活性[12]，降低GnRH mRNA表達。

通過激活A(yù)MPK激動其下游信號調(diào)控GT1-7細(xì)胞GnRH分泌。球狀域脂聯(lián)素[57]通過AMPK-ERK，增加轉(zhuǎn)錄因子SPl表達或磷酸化，抑制GTl-7細(xì)胞Kiss-1基因啟動子活性及其表達，影響GnRH分泌。GT1-7細(xì)胞中，球狀脂連蛋白能激活A(yù)MPK磷酸化使乙酰輔酶A羧化酶（ACC）失活，使丙二酰CoA降低，增加AgRP mRNA表達，證明在下丘腦球狀脂連蛋白調(diào)節(jié)能量平衡通過AMPK-ACC信號通路而非JAK-STAT3通路[58]，在泌乳奶牛飼喂苜蓿青貯粗飼料研究中發(fā)現(xiàn)，血清中胰島素和脂聯(lián)素濃度提高，激活奶牛乳腺胰島素信號通路及JAK2-STAT信號通路[59]。Beall等研究發(fā)現(xiàn)，GT1-7神經(jīng)元能表現(xiàn)出GE型葡萄糖傳感行為并通過AMPKα2-UCP2通路實現(xiàn)[60]。

4.2 MAPK相關(guān)信號通路

絲裂原活化蛋白激酶（MAPKs）是一組可被不同細(xì)胞外刺激，如細(xì)胞因子、神經(jīng)遞質(zhì)、激素、細(xì)胞應(yīng)激及細(xì)胞黏附等激活絲氨酸-蘇氨酸蛋白激酶。MAPK信號轉(zhuǎn)導(dǎo)通路存在于大多數(shù)細(xì)胞內(nèi)，能將細(xì)胞外刺激信號轉(zhuǎn)導(dǎo)至細(xì)胞及其核內(nèi)，并引起細(xì)胞生物學(xué)反應(yīng)。研究發(fā)現(xiàn)，在GT1-7細(xì)胞中，E2能活化ERK1/2和SAPK/JNK信號通路，但不激活p38MAPK信號通路[26]。Hayes等研究發(fā)現(xiàn)[61]，利用胰高血糖素樣肽1（GLP-1）類似物Exendin-4，能通過激活PKA抑制AMPK并激活MAPK信號通路誘導(dǎo)胰高血糖素樣肽1受體（GLP-1R）表達，抑制食物攝入量。研制藥物抑制GLP-1R表達，可有效治療肥胖癥。Terasaka等研究表明，在GT1-7神經(jīng)元細(xì)胞中，Kisspeptin能夠刺激MAPK-AKT信號，與ERK信號在功能上參與GnRH mRNA表達[27]。

在GT1-7細(xì)胞中，是否存在除AMPK及MAPK相關(guān)信號通路以外信號轉(zhuǎn)導(dǎo)，使一些神經(jīng)遞質(zhì)或激素能通過多條信號通路傳導(dǎo)共同作用影響GT1-7細(xì)胞分泌活性，尚需要深入研究。

5展　望

在脊椎動物腦組織中，一些激素和神經(jīng)遞質(zhì)通過作用于GnRH神經(jīng)元調(diào)控性腺軸。因此，GnRH神經(jīng)元是這些物質(zhì)調(diào)控性腺軸共同通路。劉金玲等研究發(fā)現(xiàn)，蛋氨酸腦啡肽（MENK）能促進小鼠骨髓來源樹突狀細(xì)胞TLR-4和TNF-α表達[62]，GT1-7細(xì)胞能否表達MENK受體，如何調(diào)控免疫系統(tǒng)有待進一步探討。在體內(nèi)研究中，神經(jīng)元之間聯(lián)系復(fù)雜，難以判斷其直接作用抑或通過中間神經(jīng)元間接作用于該GnRH神經(jīng)元，利用下丘腦永生GnRH神經(jīng)元細(xì)胞系GT1-7細(xì)胞可解決該問題。GT1-7細(xì)胞作為GnRH神經(jīng)元理想細(xì)胞模型，在研究過程中取得突破，體外培養(yǎng)GT1-7細(xì)胞也更易于試驗。但隨著研究深入，GT1-7細(xì)胞模型局限性逐漸凸顯。該細(xì)胞保持單一細(xì)胞克隆特性，只能代表一種GnRH神經(jīng)元行為特點，激酶或細(xì)胞因子參與細(xì)胞活性與細(xì)胞類型、細(xì)胞成熟程度有關(guān)，某些基因或受體表達與體內(nèi)GnRH神經(jīng)元不一致，能否系統(tǒng)而精確模擬動物機體生長發(fā)育過程中各種神經(jīng)遞質(zhì)及調(diào)控因子相互作用有待探索。

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GT1-7 cells and its application in reproduction-related researches

FENG Tao1, WU Xiaomin1,2, XU Xiaoling1, BAI Jiahua1, SONG Yuqing1, XIAO Linli1, HAN Xiangmin2, LIU Yan1(1. Institute of Animal Husbandry and Veterinary Medicine, Beijing Academy of Agriculture and Forestry Sciences, Beijing 100097, China; 2. School of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China)

Abstract:GT1-7 cells were subset strains of GT1 cell lines, which were GnRH neuron cell line isloated from the hypothalamus of transgenic mice and had the typical characteristics of highly differentiated neuroendocrine cells. GnRH synthesis and release in hypothalamus played an important role in reproductive function. However, GnRH neurons were only thousands and discrete distribution in the brain, which leads more difficulties in studying in vivo. Currently, GT1-7 cell was widely using in reproduction-related research works as an ideal GnRH cell model in vitro. In this review, the source, characteristics, applications as well as signaling pathways influencing ability of GT1-7 cells were summarized to provide references for GnRH related researches.

Key words:GT1-7 cells; gonadotropin-releasing hormone; reproductive regulation；signaling pathway

*通訊作者:劉彥，研究員，研究方向為動物繁殖學(xué)。E-mail: liuyanxms@163.com

作者簡介：馮濤（1980-），男，副研究員，博士，研究方向為動物生殖營養(yǎng)調(diào)控。E-mail: fengtao_gs@163.com

基金項目：國家自然科學(xué)基金項目（31501950）；北京市科技新星計劃項目（Z141105001814046）；奶牛產(chǎn)業(yè)技術(shù)體系北京市創(chuàng)新團隊

收稿日期：2015-06-29

中圖分類號：R58

文獻標(biāo)志碼：A

文章編號：1005-9369（2016）01-0102-07