硫化氢上调SIRT1拮抗甲醛诱导PC12细胞内质网应激

发布时间：2018-03-08 21:03

本文选题：硫化氢　切入点：甲醛　出处：《南华大学》2013年硕士论文　论文类型：学位论文

【摘要】：目的：本课题以甲醛损伤PC12细胞为甲醛神经毒性的细胞模型，探讨硫化氢是否能通过调控SIRT1的表达而拮抗甲醛诱导的PC12细胞的内质网应激。方法： CCK8法检测细胞的活性；PI染色流式细胞仪和Hoechst33258染色分析细胞凋亡情况；Western Blotting检测GRP78, CHOP, Cleaved-caspase-12和SIRT1蛋白的表达。结果： 1、H_2S抑制甲醛诱导的细胞毒性 FA (120or240μmol/L,24h)可浓度依赖性诱导PC12细胞活性下降及PC12细胞的凋亡，而NaHS（100,200or400μmol/L）可浓度依赖性抑制FA (120or240μmol/L,24h)诱导的PC12细胞的活性下降及PC12细胞凋亡，表明H_2S能够对抗FA对PC12细胞的毒性作用。 FA (120μmol/L)处理细胞12h、24h、36h后，FA导致的PC12细胞的活性的下降及凋亡呈时间依赖性加重，NaHS (200μmol/L)逆转FA导致的PC12细胞的活性的下降及凋亡的作用呈时间依赖性增强。 2、甲醛对PC12细胞产生内质网应激(ERS) FA (60,120or240μmol/L)处理PC12细胞24小时后GRP78, CHOP andCleaved-caspase-12蛋白表达呈浓度依赖性上调，表明FA诱导PC12细胞产生ERS。 3、硫化氢抑制甲醛诱导的PC12细胞的内质网应激 H_2S（100or200μmol/L）可浓度依赖性抑制FA (120μmol/L,24h)诱导的PC12细胞中GRP78, CHOP and Cleaved-caspase-12蛋白表达的上调。结果表明H_2S拮抗FA诱导的PC12细胞ERS。 4、H_2S通过上调SIRT1抑制甲醛诱导的PC12细胞内质网应激。 4.1H_2S促进PC12细胞高表达SIRT1，，减轻甲醛对SIRT1表达的抑制作用 H_2S (100,200or400μmol/L,24h)处理PC12细胞后浓度依赖性地上调SIRT1蛋白表达。H_2S(200μmol/L)上调SIRT1蛋白表达并可逆转FA(120μmol/L,24h)作用PC12细胞后对SIRT1蛋白表达的抑制作用。结果表明H_2S促进PC12细胞高表达SIRT1，减轻了FA对SIRT1蛋白表达的抑制作用。 4.2SIRT1抑制剂可阻断H_2S对甲醛诱导的PC12细胞ERS的抑制作用 NaHS（200μmol/L）可对抗FA (120μmol/L)导致的PC12细胞活性下降及凋亡，而加入sirtino（l15μmol/L）预处理2h后NaHS对抗FA导致的PC12细胞的活性下降及凋亡的作用被部分阻断。 NaHS（200μmol/L）可抑制FA (120μmol/L,24h)诱导的PC12细胞中GRP78,CHOP and Cleaved-caspase-12蛋白表达的上调。而加入sirtinol（15μmol/L）预处理PC12细胞2h后NaHS抑制FA诱导的PC12细胞中GRP78, CHOP andCleaved-caspase-12蛋白表达的上调的作用被阻断，此结果表明H_2S可以通过上调SIRT1抑制FA诱导的PC12细胞的ERS。结论： 1、甲醛诱导PC12细胞产生内质网应激,硫化氢拮抗甲醛诱导的PC12细胞的内质网应激。 2、硫化氢通过上调SIRT1拮抗甲醛诱导PC12细胞内质网应激。
[Abstract]:Objective:. To investigate whether hydrogen sulfide can antagonize the endoplasmic reticulum stress induced by formaldehyde in PC12 cells by regulating the expression of SIRT1. Methods:. The expression of GRP78, CHOP, Cleaved-caspase-12 and SIRT1 proteins was detected by CCK8 and Hoechst33258 staining. Results:. Inhibitory effect of H2S on the cytotoxicity induced by formaldehyde. FA (120 or 240 渭 mol / L ~ (24 h)) could induce the decrease of PC12 cell activity and the apoptosis of PC12 cells in a concentration-dependent manner, while NaHSN _ (100) (200 渭 mol / L) inhibited the decrease of PC12 cell activity and PC12 cell apoptosis induced by FA (120 or 240 渭 mol / L ~ (24) h) in a concentration-dependent manner, which indicated that H _ (2) S could antagonize the toxicity of FA to PC12 cells. FA (120 渭 mol 路L ~ (-1)) treated with FA for 12 h or 24 h or 36 h, decreased the activity of PC12 cells in a time-dependent manner and increased the apoptosis of PC12 cells induced by FA in a time-dependent manner. The effect of FA on the activity of PC12 cells was reversed in a time-dependent manner, and the effect of apoptosis was enhanced in a time-dependent manner. 2, formaldehyde produces endoplasmic reticulum stress (ERS) in PC12 cells. PC12 cells were treated with FA 60 or 120 or 240 渭 mol / L for 24 hours. The expression of CHOP andCleaved-caspase-12 protein was up-regulated in a concentration-dependent manner, indicating that FA induced PC12 cells to produce ERSs. Inhibitory effect of hydrogen sulfide on endoplasmic reticulum stress induced by formaldehyde in PC12 cells. The expression of GRP78 and CHOP and Cleaved-caspase-12 in PC12 cells induced by FA (120 渭 mol / L for 24 h) was inhibited in a concentration-dependent manner. The results showed that H2S antagonized the expression of PC12 cells induced by FA. 4. H2S upregulated SIRT1 to inhibit endoplasmic reticulum stress induced by formaldehyde in PC12 cells. 4.1 H2S promotes the overexpression of SIRT1 in PC12 cells and alleviates the inhibitory effect of formaldehyde on SIRT1 expression. After treated with H2S200 渭 mol / L for 24 h, H2S increased the expression of SIRT1 protein in a concentration-dependent manner. H2SN / L (200 渭 mol / L) upregulated the expression of SIRT1 protein and reversed the inhibitory effect of FA(120 渭 mol / L on the expression of SIRT1 protein in PC12 cells. The results showed that HSn2S promoted the overexpression of SIRT1 in PC12 cells and alleviated the expression of FA in PC12 cells. Inhibition of SIRT1 protein expression. 4.2SIRT1 inhibitor can block the inhibitory effect of HSPs on formaldehyde-induced ERS in PC12 cells. NaHS(200 渭 mol / L could antagonize the decrease of PC12 cell activity and apoptosis induced by FA (120 渭 mol / L). However, after pretreatment with sirtino(l15 渭 mol / L for 2 h, the effect of NaHS on the decrease of PC12 cell activity and apoptosis induced by FA was partially blocked. NaHS(200 渭 mol / L inhibited the up-regulation of GRP78 chop andCleaved-caspase-12 protein expression in PC12 cells induced by FA 120 渭 mol / L for 24 h, but NaHS inhibited the up-regulation of NaHS on FA-induced NaHS andCleaved-caspase-12 expression in PC12 cells after pretreatment with sirtinol(15 渭 mol / L for 2 h. These results suggest that H2S can up-regulate the inhibition of PC12 cells induced by FA by SIRT1. Conclusion:. 1. Formaldehyde induced endoplasmic reticulum stress in PC12 cells, and hydrogen sulfide antagonized endoplasmic reticulum stress in PC12 cells induced by formaldehyde. 2. Hydrogen sulfide antagonizes endoplasmic reticulum stress induced by formaldehyde by up-regulating SIRT1.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R114

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