發(fā)布時間：2018-12-23 12:28

【摘要】：目的：通過體外實驗，觀察垂體腺苷酸環(huán)化酶激活多肽(pituitary adenylate cyclaseactivating polypeptide，PACAP)衍生的小環(huán)肽CHSDGIC對紫外光B照射誘導(dǎo)凋亡的大鼠視網(wǎng)膜神經(jīng)節(jié)細胞株RGC-5的保護作用，為CHSDGIC預(yù)防視網(wǎng)膜神經(jīng)節(jié)細胞病變提供實驗依據(jù)。 方法：用高糖DMEM常規(guī)培養(yǎng)RGC-5，然后將RGC-5置于紫外光B（Ultraviolet,UVB;中波312nm）照射下進行細胞凋亡誘導(dǎo)處理。(1)采用Western-blot法檢測正常條件下和UVB照射1min、3min、5min后RGC-5體內(nèi)PAC1受體的表達情況。(2)常規(guī)培養(yǎng)RGC-5，然后分為4個實驗組和1個對照組，各實驗組加入不同質(zhì)量濃度的CHSDGIC(1μmol/L、10μmol/L、100μmol/L、1000μmol/L)，對照組加入0.9%NaCl，培養(yǎng)1h，各組細胞同時接受UVB照射3min，24h后采用CCK-8試劑盒酶標儀檢測各組RGC-5存活情況；流式細胞儀Annexin-V/PI雙染法檢測RGC-5凋亡情況，主要分析10μmol/L CHSDGIC對細胞凋亡率的影響。 結(jié)果：(1) Western-Blot檢測發(fā)現(xiàn)，正常條件及UVB照射下RGC-5體內(nèi)均有PAC1受體的表達；(2) CCK-8法檢測顯示1μmol/L、10μmol/L、100μmol/L、1000μmol/LCHSDGIC的細胞存活率分別為(34.00±1.07)%、(55.20±2.23)%、(36.69±1.96)%、(42.93±1.91)%，，對照組的存活率為(24.62±1.72)%，差異有統(tǒng)計學(xué)意義(P值均0.05)，其中10μmol/L組的細胞存活率最高；(3)流式Annexin-V/PI雙染法凋亡檢測發(fā)現(xiàn)，UVB作用于RGC-5的對照組細胞凋亡率(37.40±2.40)%較正常生長的RGC-5細胞凋亡率(5.43±1.02)%高，差異有統(tǒng)計學(xué)意義(P值0.05)；10μmol/LCHSDGIC實驗組細胞凋亡率為(15.30±1.10)%，較對照組相比差異有統(tǒng)計學(xué)意義(P值0.05)。 結(jié)論：RGC-5存在PACAP的特異性受體PAC1，CHSDGIC對UVB誘導(dǎo)凋亡條件下的RGC-5有明顯的保護作用。
[Abstract]:Aim: to observe the protective effect of pituitary adenylate cyclase activated polypeptide (pituitary adenylate cyclaseactivating polypeptide,PACAP) -derived small cyclic peptide CHSDGIC on rat retinal ganglion cell line RGC-5 induced by ultraviolet B irradiation in vitro. To provide experimental evidence for the prevention of retinal ganglion cytopathia by CHSDGIC. Methods: RGC-5, was cultured with high glucose DMEM and then RGC-5 was placed in ultraviolet B (Ultraviolet,UVB;). (1) the expression of PAC1 receptor in RGC-5 was detected by Western-blot method in normal condition and 3 mins 5 min after UVB irradiation. (2) the expression of PAC1 receptor in RGC-5 was detected by routine culture of RGC-5,. Then they were divided into four experimental groups and one control group. Each group was treated with different concentration of CHSDGIC (1 渭 mol/L,10 渭 mol/L,100 渭 mol/L,1000 渭 mol/L), the control group was treated with 0.9 NaCl1 for 1 hour, and the cells in each group were irradiated with UVB for 3 min at the same time. After 24 hours, the survival of RGC-5 in each group was detected by CCK-8 kit enzyme labeling instrument. The apoptosis of RGC-5 was detected by flow cytometry (Annexin-V/PI) double staining, and the effect of 10 渭 mol/L CHSDGIC on the apoptosis rate was analyzed. Results: (1) the expression of PAC1 receptor in RGC-5 was found in normal condition and UVB irradiation by Western-Blot. (2) the cell survival rate of 1 渭 mol/L,10 渭 mol/L,100 渭 mol/L,1000 渭 mol/LCHSDGIC was (34.00 鹵1.07)%, (55.20 鹵2.23)%, (36.69 鹵1.96)%, (42.93 鹵1.91)%, respectively. The survival rate of the control group was (24.62 鹵1.72)%, and the difference was statistically significant (P < 0. 05). The cell survival rate of the 10 渭 mol/L group was the highest. (3) the apoptosis rate of the control group treated with UVB was (37.40 鹵2.40)% higher than that of the normal growth RGC-5 cell (5.43 鹵1.02)%, and the apoptosis rate of the control group was (37.40 鹵2.40)% higher than that of the normal growth group (5.43 鹵1.02)%). The difference was statistically significant (P 0.05). The apoptotic rate of the 10 渭 mol/LCHSDGIC experimental group was (15.30 鹵1.10)%, which was significantly higher than that of the control group (P < 0. 05). Conclusion: PAC1,CHSDGIC, a specific receptor for PACAP in RGC-5, has an obvious protective effect on RGC-5 induced by UVB.
【學(xué)位授予單位】：暨南大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R774.1

【參考文獻】

相關(guān)期刊論文 前3條

1 朱益華,蔣幼芹,劉忠浩;視神經(jīng)再生[J];國外醫(yī)學(xué).眼科學(xué)分冊;2000年04期

2 丁勇;湯翠翠;余榕捷;黃霖;陳建蘇;趙秀麗;徐錦堂;;小環(huán)肽CHSDGIC對兔角膜堿燒傷愈合的作用[J];眼科新進展;2011年10期

3 趙秀麗;陳建蘇;余榕捷;丁勇;鄭佩娥;李曉霞;田振彩;戴靜;;PACAP38對兔角膜瓣術(shù)后三叉神經(jīng)節(jié)細胞及角膜神經(jīng)生長的影響[J];眼科研究;2010年08期

本文編號：2389985