内毒素经MyD88信号通路介导肝星状细胞增殖与凋亡的研究

发布时间：2018-02-09 19:52

  本文关键词： 内毒素 肝星状细胞 髓样分化因子88 toll样受体4　出处：《暨南大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的 观察不同浓度内毒素对TLR4和MyD88，及其依赖MyD88信号通路上多种细胞因子表达的影响，以及对大鼠肝星状细胞（HSCs）增殖与凋亡的影响，探讨MyD88在内毒素诱导HSCs凋亡信号转导中的作用，，探索肝纤维化发病机制。 方法 体外培养大鼠HSC株HSC-T6，取细胞浓度与吸光值呈直线关系且斜率最大处的细胞浓度作为接种浓度，将内毒素按浓度0.1，1.0，10.0EIU/mL（分别为内毒素I组、II组和III组）各作用于HSCs，以未加干预的HSCs为正常对照组(第IV组)，采用RT-PCR及WesternBlot术分别检测TLR4和MyD88的mRNA与蛋白表达的变化，以及依赖MyD88信号通路的相关因子TRAF6、TNF-α、NF-кΒ的mRNA与蛋白表达的变化，并同时以流式细胞术观察HSC凋亡的变化。 结果 1.内毒素I组、II组和III组中，MyD88、TLR4、NF-κB的mRNA及蛋白的表达量较对照组均有所增高（P0.05），其中，内毒素II组升高更明显（P0.01）。 2.内毒素I组、II组中，TRAF-6、TNF-α的mRNA及蛋白的表达量较对照组有所增高（P0.05），而在内毒素III组中，其mRNA及蛋白的表达量无明显影响（P＞0.05）。 3.内毒素I组、II组中HSC的平均吸光度显著高于对照组(P0.05)。而毒素III组中HSC的平均吸光度无明显增高(P＞0.05)，提示一定浓度的内毒素能够促进肝星状细胞的增殖。 4.内毒素I组、II组中HSC凋亡减少（P0.05），其中内毒素II组中HSC凋亡减少更明显，（P0.01），而内毒素III组中HSC凋亡无明显影响（P0.05）。 结论 一定浓度的内毒素可以促进TLR4、MyD88及其信号通路相关因子TRAF6、TNF-α、NF-кΒ的表达，促进肝星状细胞的增殖，抑制其凋亡，内毒素可能由此机制促进肝纤维化发生发展。
[Abstract]:Purpose. To investigate the effects of different concentrations of endotoxin on the expression of cytokines in TLR4, MyD88and its dependent MyD88 signaling pathway, and on the proliferation and apoptosis of rat hepatic stellate cells (HSCs), and to explore the role of MyD88 in the signal transduction of HSCs apoptosis induced by endotoxin. To explore the pathogenesis of liver fibrosis. Method. Rat HSC strain HSC-T6 was cultured in vitro. The cell concentration which had a linear relationship with the absorptivity and the highest slope was taken as the inoculation concentration. Endotoxin was treated with 0.1 ~ 1.0 ~ 1.0 ~ 10.0 EIU-mL (endotoxin I group II and III group) respectively, and HSCs without intervention was used as normal control group (group IV). The changes of mRNA and protein expression of TLR4 and MyD88 were detected by RT-PCR and WesternBlot, respectively. The changes of mRNA and protein expression of TRAF6TNF- 伪 TNF- 伪 尾 were also observed by flow cytometry. The apoptosis of HSC was also observed by flow cytometry. Results. 1. The expression of mRNA and protein of MyD88TLR4NF- 魏 B in group I and group III were higher than those in control group (P 0.05), especially in group II (P 0.01). 2. The expression of mRNA and protein of TRAF-6 TNF- 伪 in lipopolysaccharide I group was higher than that in control group (P 0.05), but the expression of mRNA and protein in endotoxin III group had no significant effect (P > 0.05). 3. The average absorbance of HSC in group I was significantly higher than that in control group (P 0.05), but the average absorbance of HSC in III group was not significantly increased (P > 0.05), suggesting that endotoxin at a certain concentration could promote the proliferation of hepatic stellate cells. 4. The apoptosis of HSC in group I was decreased by P0.05, and the apoptosis of HSC was decreased more significantly in group II, but the apoptosis of HSC in group III was not significantly affected by P0.05. Conclusion. A certain concentration of endotoxin can promote the expression of TLR4- MyD88 and its signal transduction related factor TRAF6- TNF- 伪 -NF- 尾, promote the proliferation of hepatic stellate cells and inhibit its apoptosis. Endotoxin may promote the occurrence and development of hepatic fibrosis.
【学位授予单位】：暨南大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R575.2

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