【摘要】：低氧作為一種非特異性應(yīng)激原可誘導(dǎo)機(jī)體產(chǎn)生應(yīng)激反應(yīng)。哺乳動(dòng)物通過(guò)下丘腦-垂體-腎上腺皮質(zhì)軸(Hypothalamus-pituitary-adrenal axis, HPA)對(duì)機(jī)體面臨的各種應(yīng)激做出應(yīng)答反應(yīng)。應(yīng)激促進(jìn)下丘腦室旁核(Paraventricular nucleus, PVN)神經(jīng)元釋放下丘腦促腎上腺皮質(zhì)激素釋放激素(Corticotropin releasing factor, CRF)。 CRF主要通過(guò)和促腎上腺皮質(zhì)激素釋放激素I型受體(Corticotropin releasing factor receptor-1, CRFR1)和促腎上腺皮質(zhì)激素釋放激素II型受體(Corticotropin releasing factor receptor-2, CRFR2)結(jié)合發(fā)揮生理作用,參與神經(jīng)和體液調(diào)節(jié)。 環(huán)境低氧嚴(yán)重時(shí)可導(dǎo)致機(jī)體發(fā)生腦水腫,腦水腫的發(fā)生、發(fā)展與水通道蛋白(Aquaporins, AQPs)密切相關(guān)。AQPs是細(xì)胞膜上一類(lèi)特異性水通道蛋白,參與多種類(lèi)形細(xì)胞的水分子跨膜運(yùn)輸。其中水通道蛋白4(Aquaporin-4, AQP4)在哺乳動(dòng)物腦組織中分布最為豐富,在正常生理狀態(tài)下參與大腦水平衡調(diào)節(jié),在病理?yè)p傷時(shí)則參與腦水腫形成和消退。AQP4在星形膠質(zhì)細(xì)胞中大量表達(dá),因此星形膠質(zhì)細(xì)胞的腫脹與腦水腫有重要關(guān)系。低氧誘導(dǎo)因子-1(Hypoxia induced factor-1, HIF-1)是低氧敏感因子,參與調(diào)節(jié)機(jī)體氧平衡。低氧上調(diào)HIF-la蛋白表達(dá),使HIF-1穩(wěn)定性增加并與低氧反應(yīng)元件(Hypoxia response element, HRE)結(jié)合作用下游靶基因,調(diào)控生理或病理作用。HIF-1調(diào)控的靶基因有血管內(nèi)皮生長(zhǎng)因子(Vascular endothelial growth factor, VEGF)及其受體(Vascular endothelial growth factor receptor, VEGFR)、誘導(dǎo)型一氧化氮合酶(Inducible nitric oxide synthase, iNOS)等。iNOS是一氧化氮合酶(Nitric oxide synthase, NOS)的一種,主要參與機(jī)體免疫反應(yīng),能夠催化L-精氨酸(L-Arginine)產(chǎn)生一氧化氮(Nitric oxide, NO)。 NO是著名的氣體信號(hào)分子,可以通過(guò)NO-cGMP-PKG通路調(diào)節(jié)細(xì)胞和機(jī)體功能。CRFR1在中樞神經(jīng)系統(tǒng)和外周系統(tǒng)廣泛分布,在星形膠質(zhì)細(xì)胞中也有表達(dá)。我們實(shí)驗(yàn)室前期結(jié)果表明,低氧時(shí)下丘腦CRF分泌增加,前額葉皮層CRF基因轉(zhuǎn)錄升高。因此,我們?cè)O(shè)想機(jī)體在嚴(yán)重低氧時(shí)一方面通過(guò)CRF釋放增加,激活星形膠質(zhì)細(xì)胞CRFR1,引發(fā)胞內(nèi)信號(hào)通路；另一方面細(xì)胞在嚴(yán)重低氧時(shí)胞內(nèi)低氧誘導(dǎo)因子-1α(Hypoxia induced factor-1α, HIF-1α)激活,驅(qū)動(dòng)下游信號(hào)通路。這兩條通路共同作用于AQP4,增加AQP4水通道功能。 本研究通過(guò)低氧培養(yǎng)箱模擬星形膠質(zhì)細(xì)胞低氧環(huán)境,通過(guò)western blot、熒光標(biāo)記法和細(xì)胞免疫熒光等方法,對(duì)CRFR1如何參與低氧誘導(dǎo)的星形膠質(zhì)細(xì)胞水腫進(jìn)行了研究。研究結(jié)果發(fā)現(xiàn)AQP4與CRFR1在大鼠原代皮層星形膠質(zhì)細(xì)胞共表達(dá)；10nM和100nM CRF均能誘導(dǎo)大鼠原代星形膠質(zhì)細(xì)胞胞內(nèi)鈣離子濃度升高,該作用可以被預(yù)孵CRFR1拮抗劑CP154,526阻斷。同時(shí)發(fā)現(xiàn)大鼠原代星形膠質(zhì)細(xì)胞在低氧(1%02)下HIF-1α蛋白表達(dá)增加,但并不表達(dá)iNOS蛋白；低氧上調(diào)大鼠原代小膠質(zhì)細(xì)胞iNOS表達(dá)。這些結(jié)果提示低氧誘導(dǎo)皮層神經(jīng)元分泌CRF,通過(guò)在星形膠質(zhì)細(xì)胞G-蛋白偶聯(lián)受體CRFR1激活胞內(nèi)第二信使Ca2+,增加AQP4水通透性；低氧使胞內(nèi)HIF-1α蛋白集聚增多,誘導(dǎo)下游低氧相關(guān)靶基因表達(dá),促進(jìn)AQP4水通透性增加。 創(chuàng)新點(diǎn)： 1.發(fā)現(xiàn)低氧不激活離體培養(yǎng)大鼠原代星形膠質(zhì)細(xì)胞的iNOS-NO-PKG通路。
[Abstract]:Hypoxia, as a non-specific stress, can induce stress reaction in that body. The mammal responds to the various stresses facing the body through the hypothalamic-pituitary-adrenal axis (HPA). Stress promotes the release of the corticotropin-releasing hormone (CRF) from the paraventricular nucleus (PVN) of the hypothalamus. The CRF mainly plays a physiological role in combination with the corticotropin-releasing hormone type I receptor (CRFR1) and the corticotropin-releasing hormone type II receptor (CRFR2), and is involved in the regulation of nerve and body fluid. The development of water channel protein (AQPs) is closely related to the development of water channel protein (AQPs). AAQPs is a class of specific water-channel proteins on the cell membrane, which is involved in the cross-membrane transport of water molecules in multi-species cells. The water channel protein 4 (Aquaporin-4, AQP4) is most abundant in the brain of the mammal, and is involved in the brain water balance regulation in the normal physiological state, and is involved in the formation and elimination of the brain edema in the case of pathological injury. The expression of AQP4 in astrocyte is a significant effect on the swelling and brain edema of astrocytes. The hypoxia-inducible factor-1 (HIF-1) is a hypoxia-sensitive factor and is involved in the regulation of the oxygen level in the body. The expression of HIF-la protein is up-regulated by hypoxia, and the stability of HIF-1 is increased and the downstream target gene is combined with the hypoxia response element (HRE) to control the physiological or pathological changes. The target gene regulated by. HIF-1 has vascular endothelial growth factor (VEGF) and its receptor (VEGFR) and inducible nitric oxide synthase (iNOS). The iNOS is a kind of nitric oxide synthase (NOS), which is mainly involved in the immune response of the organism and can catalyze the production of nitric oxide (NO) in L-arginine (L-Arginine). ). NO is a well-known gas signal molecule that can be used to regulate cell and body work through the NO-cGMP-PKG pathway can. CRFR1 is widely distributed in the central nervous system and the peripheral system, and there are also tables in astrocyte Da. The early results of our laboratory indicated that the secretion of CRF in the hypothalamus of the prefrontal cortex increased, and the transcription of CRF gene in the prefrontal cortex was increased. High. Therefore, we imagine that, on the one hand, in the case of severe hypoxemia, on the one hand, the increase of CRFR1 of the astrocytes is activated and the intracellular signaling pathway is induced; on the other hand, the cells are activated by the hypoxemia induction factor-1 (HIF-1) in severe hypoxia, and the downstream signal is driven. Road. These two paths act together on AQP4 and increase AQP4 water channel work In this study, the hypoxia environment of astrocyte was simulated by a low-oxygen incubator, and the method of western blot, fluorescent labeling and cell immunofluorescence was used to study how CRFR1 was involved in hypoxia-induced astrocyte edema. The results of the study showed that AQP4 and CRFR1 were co-expressed in primary rat cortical astrocytes; both 10 nM and 100 nM CRF were able to induce an increase in intracellular calcium ion concentration in primary astrocytes of the rat, which could be pre-treated with CRFR1 antagonists CP154,52, 6. At the same time, the expression of HIF-1 was increased under hypoxia (1%02), but the iNOS protein was not expressed. S expression. These results suggest that the hypoxia-induced cortical neurons secrete CRF and increase the water permeability of AQP4 by activating the second messenger Ca2 + in the intracellular second messenger Ca2 + in the G-protein-coupled receptor CRFR1 of the astrocyte. To promote AQP4 water permeability due to expression The sex is increased. Innovation point:1. It is found that hypoxia does not activate the iNOS-N in the primary astrocytes of the isolated rat
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R363

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 吳鋼,郝亞榮,李庚山,黃從新,王晶,謝強(qiáng),黃崢嶸;低氧誘導(dǎo)心肌細(xì)胞表達(dá)血管內(nèi)皮生長(zhǎng)因子發(fā)生機(jī)制的實(shí)驗(yàn)研究[J];微循環(huán)學(xué)雜志;2004年01期

2 韓沾元，陳銘，呂及忍，，聞萍，宋雷蕾，吳奇英;低氧誘導(dǎo)兔血MDA和棘狀紅細(xì)胞增加及其被MPEG－SOD抑制[J];生理學(xué)報(bào);1995年06期

3 梅長(zhǎng)林,陳蕾;低氧誘導(dǎo)的大鼠系膜細(xì)胞增殖及基質(zhì)mRNA表達(dá)的研究[J];中華腎臟病雜志;1996年06期

4 陳偉,陳家佩;低氧誘導(dǎo)促紅細(xì)胞生成素基因表達(dá)的機(jī)制[J];國(guó)外醫(yī)學(xué).生理.病理科學(xué)與臨床分冊(cè);2000年06期

5 宗建春,吳誠(chéng)義;低氧誘導(dǎo)人乳腺癌細(xì)胞VEGF的表達(dá)及機(jī)制[J];重慶醫(yī)科大學(xué)學(xué)報(bào);2005年02期

6 范蓓,王艷霞,姚泰,朱依純;p38絲裂原素激活的蛋白激酶在調(diào)節(jié)低氧誘導(dǎo)人內(nèi)皮細(xì)胞分泌血管內(nèi)皮生長(zhǎng)因子過(guò)程中的作用(英文)[J];生理學(xué)報(bào);2005年01期

7 聶小維;孫立軍;郝躍文;楊廣笑;王全穎;;構(gòu)建人工合成的微小低氧反應(yīng)元件可調(diào)控的AAV載體[J];細(xì)胞與分子免疫學(xué)雜志;2011年03期

8 呂順艷,朱妙章,郭海濤,于軍,魏?jiǎn)⒚?血管鈉肽對(duì)中度低氧誘導(dǎo)的心肌細(xì)胞蛋白合成有抑制作用[J];生理學(xué)報(bào);2002年01期

9 呂國(guó)蔚;低氧適應(yīng)的進(jìn)化[J];首都醫(yī)科大學(xué)學(xué)報(bào);2002年02期

10 林紅,蔡英年,鄧希賢,李世強(qiáng),梁兵,周曉梅,蔡起薔,楊燕芬;低氧誘導(dǎo)肺血管平滑肌細(xì)胞誘生型一氧化氮合酶基因表達(dá)[J];基礎(chǔ)醫(yī)學(xué)與臨床;1997年03期

相關(guān)會(huì)議論文 前1條

1 劉戟環(huán);糜漫天;張乾勇;許紅霞;;牛黃酸對(duì)低氧誘導(dǎo)大鼠視網(wǎng)膜病變的防護(hù)作用及機(jī)制研究[A];中國(guó)營(yíng)養(yǎng)學(xué)會(huì)第九次全國(guó)營(yíng)養(yǎng)學(xué)術(shù)會(huì)議論文摘要匯編[C];2004年

相關(guān)重要報(bào)紙文章 前4條

1 汪敏 章米力;低氧誘導(dǎo)白血病細(xì)胞分化[N];健康報(bào);2005年

2 汪敏 章米力;治療白血病新理論[N];保健時(shí)報(bào);2005年

3 汪敏 仇逸;低氧可誘導(dǎo)白血病細(xì)胞分化[N];醫(yī)藥經(jīng)濟(jì)報(bào);2003年

4 ;鈉尿肽防治低氧性心臟病[N];中國(guó)醫(yī)藥報(bào);2004年

相關(guān)博士學(xué)位論文 前1條

1 呂順艷;血管鈉肽對(duì)低氧誘導(dǎo)的心肌細(xì)胞、心成纖維細(xì)胞以及血管平滑肌細(xì)胞生長(zhǎng)的抑制調(diào)節(jié)[D];第四軍醫(yī)大學(xué);2002年

相關(guān)碩士學(xué)位論文 前5條

1 郝躍文;分泌表達(dá)PR39的重組腺相關(guān)病毒的構(gòu)建和促進(jìn)低氧環(huán)境雞胚血管新生作用的研究[D];第四軍醫(yī)大學(xué);2008年

2 聶小維;HRE啟動(dòng)子調(diào)控分泌表達(dá)PR39重組AAV的構(gòu)建及在CHD基因治療中的應(yīng)用研究[D];第四軍醫(yī)大學(xué);2011年

3 王彥榮;氯化鈷模擬低氧誘導(dǎo)白血病細(xì)胞分化及化療增敏作用的實(shí)驗(yàn)研究[D];吉林大學(xué);2004年

4 單俊杰;金雀異黃素對(duì)缺氧誘導(dǎo)的人ARPE-19細(xì)胞內(nèi)堿性成纖維細(xì)胞生長(zhǎng)因子蛋白表達(dá)的影響[D];南京醫(yī)科大學(xué);2004年

5 夏小俊;低氧對(duì)人肝癌細(xì)胞內(nèi)ARE介導(dǎo)的NQO1酶基因表達(dá)及其對(duì)生物還原劑藥物毒性的影響[D];浙江大學(xué);2002年

本文編號(hào)：2479668