發(fā)布時(shí)間：2018-09-14 20:30

【摘要】：目的： 通過(guò)分析用氯胺酮刺激人膀胱SV-HUC-1細(xì)胞后,誘導(dǎo)其發(fā)生細(xì)胞凋亡,為初步探討氯胺酮相關(guān)性泌尿系統(tǒng)損傷的發(fā)病機(jī)制提供理論基礎(chǔ)。 方法： 培養(yǎng)人膀胱SV-HUC-1永生化上皮細(xì)胞系,用不同濃度(Ommol/L、0.5mmol/L、1mmol/L、2mmol/L)的氯胺酮刺激SV-HUC-1細(xì)胞24小時(shí),用Annexin V-FITC/PI雙染法流式細(xì)胞術(shù)檢測(cè)細(xì)胞凋亡率,Western免疫印跡法檢測(cè)Bax、Bcl-2、pro-caspase-3、Cleaved Caspase-3蛋白表達(dá)的相對(duì)含量,并選取上述實(shí)驗(yàn)中結(jié)果最佳的氯胺酮濃度作為下一步實(shí)驗(yàn)的工作濃度,繼續(xù)作用于SV-HUC-1細(xì)胞不同的時(shí)間后行上述實(shí)驗(yàn)。實(shí)驗(yàn)數(shù)據(jù)使用SPSS19.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)分析。 結(jié)果： 1.氯胺酮對(duì)SV-HUC-1細(xì)胞凋亡率的影響 0mmol/L、0.5mmol/L、1mmol/L、2mmol/L的氯胺酮作用SV-HUC-1細(xì)胞24小時(shí)后,流式細(xì)胞術(shù)檢測(cè)結(jié)果顯示各組細(xì)胞凋亡比率分別為(3.33±0.24)%、(5.16±0.57)%、(9.39±0.52)%、(11.33±0.40)%,各組間比較P值均0.05。相關(guān)分析結(jié)果顯示,SV-HUC-1細(xì)胞凋亡率與氯胺酮質(zhì)量濃度呈正相關(guān)(r=0.829,P0.05)。 用2mmol/L的氯胺酮分別作用SV-HUC-1細(xì)胞0h、12h、24h、48h后,流式細(xì)胞術(shù)檢測(cè)顯示各組細(xì)胞凋亡比率分別為(3.72±0.22)%、(6.45±0.32)%、(11.18±0.45)%、(12.24±0.40)%,各組間比較P值均0.05。相關(guān)分析結(jié)果顯示,SV-HUC-1細(xì)胞凋亡率與氯胺酮作用時(shí)間呈正相關(guān)(r=0.762,P0.05)。 2.氯胺酮對(duì)SV-HUC-1細(xì)胞Bax、Bcl-2、pro-caspase-3、Cleaved Caspase-3蛋白的表達(dá)的影響 O mmol/L、0.5mmol/L.1mmol/L.2mmol/L的氯胺酮作用于SV-HUC-1細(xì)胞24小時(shí)后,Bax和Cleaved Caspase-3蛋白(17KD.19KD)的表達(dá)較對(duì)照組明顯增多,Bax/Bcl-2比值增大(P0.05),且隨氯胺酮濃度的升高而表達(dá)增高(rBax=0.986,r19KD=0.888,r17KD=0.769,r Bax/Bcl-2=0.982,P值均0.05)；Bcl-2和pro-caspase-3蛋白表達(dá)較對(duì)照組逐漸下降(P0.05),且隨氯胺酮濃度的升高而表達(dá)下降(r Bcl-2=-0.963,r pro-caspase-3=-0.894,P值均0.05)。 氯胺酮(2mmol/L)作用于SV-HUC-1細(xì)胞Oh、12h、24h、48h后,Bax和Cleaved Caspase-3蛋白(17KD.19KD)的表達(dá)較對(duì)照組增多,Bax/Bcl-2比值增大(P0.05),隨作用時(shí)間的延長(zhǎng)而表達(dá)增多(r Bax=0.951,r19KD=0.787,r17KD=0.942,r Bax/Bcl-2＝0.979,P值均0.05)；Bcl-2和pro-caspase-3蛋白表達(dá)較對(duì)照組逐漸下降(P0.05),且隨作用時(shí)間的延長(zhǎng)而表達(dá)下降(r Bcl-2:=-0.949,r pro-caspase-3=-0.914,P值均0.05)。 結(jié)論： 氯胺酮能夠誘導(dǎo)人膀胱SV-HUC-1細(xì)胞系發(fā)生細(xì)胞凋亡,并呈濃度依賴性和時(shí)間依賴性。說(shuō)明了氯胺酮可能誘導(dǎo)尿路上皮細(xì)胞發(fā)生凋亡,導(dǎo)致尿路上皮層變薄或上皮缺損,進(jìn)一步對(duì)上皮下肌層組織造成損害,這可能是氯胺酮相關(guān)性泌尿系統(tǒng)損傷,在疾病發(fā)病早期的一個(gè)重要機(jī)制。
[Abstract]:Aim: to investigate the mechanism of Ketamine associated urinary system injury by inducing apoptosis of human bladder SV-HUC-1 cells stimulated by ketamine. Methods: human bladder SV-HUC-1 immortalized epithelial cell lines were cultured. SV-HUC-1 cells were stimulated with different concentrations of ketamine (Ommol/L,0.5mmol/L,1mmol/L,2mmol/L) for 24 hours. Annexin V-FITC/PI double staining flow cytometry was used to detect apoptosis rate and Western blot was used to detect the relative content of Bax,Bcl-2,pro-caspase-3,Cleaved Caspase-3 protein. The optimal concentration of ketamine was selected as the working concentration of the next step. The experiment was carried out after the SV-HUC-1 cells continued to be treated for different time. The experimental data were analyzed by SPSS19.0 statistical software. Results: 1. Effect of ketamine on apoptosis of SV-HUC-1 cells. 24 hours after Ketamine treated SV-HUC-1 cells with 1 mmol / L Ketamine, the apoptotic rates were (3.33 鹵0.24), (5.16 鹵0.57), (9.39 鹵0.52), (11.33 鹵0.40), respectively (P < 0.05). The results of correlation analysis showed that the apoptosis rate of SV-HUC-1 cells was positively correlated with the mass concentration of ketamine (P 0.05). Flow cytometry showed that the apoptotic rates of SV-HUC-1 cells in each group were (3.72 鹵0.22), (6.45 鹵0.32), (11.18 鹵0.45), (12.24 鹵0.40), respectively. The results of correlation analysis showed that the apoptosis rate of SV-HUC-1 cells was positively correlated with the time of ketamine treatment (P 0.05). Effect of ketamine on the expression of Bax,Bcl-2,pro-caspase-3,Cleaved Caspase-3 protein in SV-HUC-1 cells the expression of Cleaved Caspase-3 and Bax protein (17KD.19KD) in SV-HUC-1 cells increased significantly after treatment with ketamine O mmol/L,0.5mmol/L.1mmol/L.2mmol/L for 24 hours (P0.05), and increased with the concentration of ketamine (P0.05). The expression of rBax=0.986,r19KD=0.888,r17KD=0.769,r Bax/Bcl-2=0.982,P was higher than that of the control group (rBax=0.986,r19KD=0.888,r17KD=0.769,r Bax/Bcl-2=0.982,P = 0. 05). Compared with the control group, the expression of Bcl-2 and pro-caspase-3 protein decreased gradually (P0.05), and decreased with the increase of ketamine concentration (r Bcl-2=-0.963,r pro-caspase-3=-0.894,P). The expression of Bax and Cleaved Caspase-3 protein (17KD.19KD) in SV-HUC-1 cells treated with ketamine (2mmol/L) was higher than that in control group (P0.05), and increased with the prolongation of time (r Bax=0.951,r19KD=0.787,r17KD=0.942,r Bax/Bcl-2=0.979,P). Compared with the control group, the expression of Bcl-2 and pro-caspase-3 protein decreased gradually (P0.05), and decreased with the prolongation of the action time (r Bcl-2:=-0.949,r pro-caspase-3=-0.914,P). Conclusion: ketamine can induce apoptosis of human bladder SV-HUC-1 cell line in a concentration and time dependent manner. It is suggested that ketamine may induce apoptosis of urinary tract epithelial cells, resulting in thinning or epithelial defect of urinary tract epithelium, and further damage to the myometrium of epiglottis, which may be the injury of urinary system associated with ketamine. An important mechanism in the early stages of disease.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R694.3

【參考文獻(xiàn)】

相關(guān)期刊論文 前4條

1 文博;黃小佳;李曉銘;丘捷文;邱建忠;余貴亮;許詳;李春;;RTX膀胱灌注治療氯胺酮相關(guān)性下尿路癥狀的臨床研究[J];臨床泌尿外科雜志;2013年02期

2 黃亮;李東杰;唐正嚴(yán);王桂林;丁見;羅寬;黃珂;周杰;黎勇林;;氯胺酮(K粉)相關(guān)性膀胱炎3例診治分析并文獻(xiàn)復(fù)習(xí)[J];中南大學(xué)學(xué)報(bào)(醫(yī)學(xué)版);2013年09期

3 羅玉琴;魏輝;黃小佳;袁志敏;方梅;;氯胺酮相關(guān)性膀胱炎的尿動(dòng)力檢查及分析[J];臨床泌尿外科雜志;2013年11期

4 王和燕;劉忠芳;;497例氯胺酮(K粉)濫用對(duì)泌尿及消化系統(tǒng)損害的調(diào)查報(bào)告[J];中國(guó)藥物濫用防治雜志;2011年01期

，

本文編號(hào)：2243792