自噬对镉致大鼠肾小管上皮细胞凋亡的影响及其调控机制
发布时间：2018-07-31 14:45
【摘要】：镉是环境中常见的高毒性重金属污染物,急性或慢性镉暴露能引起多种器官和组织的损伤。近年来,由于工业生产中镉生产量和使用量的激增及相关工业废物带来的污染加重,使得环境中镉含量呈快速上升趋势;镉在环境中的半衰期长达10-30年,进入环境后不能像有机污染物一样被微生物降解,并且可以通过食物链效应最终蓄积在人体中。因此,镉暴露给公众健康带来的危害引起了广泛关注。肾脏是镉毒性作用的靶器官,国内外学者从环境污染、职业性暴露和动物试验等多等方面对镉所致肾毒性机理进行了广泛、深入地研究;但这些研究多集中于凋亡机理的探究,而对镉所致肾脏毒性过程中自噬的机理、作用及其与凋亡关系的探究较少。1.镉诱导大鼠肾脏皮质和肾小管上皮细胞发生自噬分为体外和体内两部分。体外试验采用机械筛网结合酶消化法建立原代大鼠肾小管上皮细胞(rPT cells)培养模型,在传一代细胞增殖活性最强时间进行镉染毒处理,同时选择NRK-52E细胞株进行同样处理。CCK-8法测定不同浓度的镉在不同染毒时间对rPT细胞和NRK-52E细胞活性的影响;MDC荧光探针法检测不同浓度的镉在不同染毒时间对rPT细胞和NRK-52E细胞自噬水平的影响;2.5 μ和5 μM镉处理rPT细胞、5 ⒚M和10 μM镉处理NRK-52E细胞6 h,免疫印迹法检测自噬相关蛋白LC3Ⅱ、p62和Beclin-1表达水平变化。结果表明,2.5 μM和5 μM镉处理rPT细胞、5 μM和10μ 镉处理NRK-52E细胞6h,细胞活性最强,自噬水平最高,极显著高于对照组(P0.01);2.5μM和5μM镉处理rPT细胞、5μM和10μM镉处理NRK-52E细胞12h,细胞自噬水平最低,极显著低于对照组(P0.01)。体内试验选择SD大鼠32只,随机分为4组:对照组(DDW)和镉组(CdAc2,50mg/kg·bw),饲养1周;对照组(DDW)和镉组(CdAc2,50mg/kg·bw),饲养8周。取肾脏皮质,RT-PCR法检测自噬相关基因LC3、p62和Beclin-1表达量变化;免疫印迹法检测自噬相关蛋白LC3Ⅱ、p62和Beclin-1表达水平变化。结果表明,镉染毒处理1周后,大鼠肾脏皮质自噬水平极显著高于对照组(P0.01);与对照组相比Beclin-1表达水平极显著升高(P0.01)。镉染毒处理8周,大鼠肾脏皮质自噬水平极显著低于对照组(P0.01)。体外和体内研究表明,低剂量镉短时间处理可以诱导肾脏皮质和肾小管上皮细胞发生自噬,而长时间处理则抑制自噬。2.镉诱导大鼠肾脏皮质和肾小管上皮细胞发生凋亡分为体外和体内两部分。体外试验采用机械筛网结合酶消化法建立原代大鼠肾小管上皮细胞(rPT cells)培养模型,在传一代细胞增殖活性最强时间进行镉染毒处理,同时选择NRK-52E细胞株进行同样处理。DAPI染色法检测不同浓度的镉在不同染毒时间对rPT细胞和NRK-52E细胞凋亡的影响;流式细胞术检测2.5 μM和5μ 镉处理rPT细胞、5(μM和10 μM镉处理NRK-52E细胞12 h,细胞凋亡率变化;免疫印迹法检测2.5 μM和5 μM镉处理rPT细胞、5 μM和10 μM镉处理NRK-52E细胞12 h,线粒体凋亡途径相关蛋白Cyt C、Cleaved caspase-9和Cleaved caspase-3表达水平变化,内质网凋亡途径相关蛋白ATF-6、GRP78、IRE1α和Cleavedcaspase-12表达水平变化,Fas/FasL凋亡途径相关蛋白Fas、FasL、FADD、Cleavedcaspase-8表达水平变化。结果表明,2.5μM和5μM镉处理rPT细胞、5μM和10μM镉处理NRK-52E细胞6h,细胞凋亡率最低(P0.01),6h之后凋亡率开始升高,12h细胞凋亡率最高(P0.01);2.5μM和5μM镉处理rPT细胞、5μM和10 μM镉处理NRK-52E细胞12 h,与对照组相比,线粒体凋亡途径、内质网凋亡途径、Fas/FasL凋亡途径相关蛋白表达水平都显著或极显著升高(P0.05或P0.01)。体内试验选择SD大鼠32只,随机分为4组:对照组(DDW)和镉组(CdAc2,50 mg/kg·bw),饲养1周;对照组(DDW)和镉组(CdAc2,50mg/kg·bw),饲养8周。饲养1周后取肾脏皮质,RT-PCR法检测凋亡基因Caspase-3表达量变化;免疫印迹法检测凋亡蛋白Cleaved caspase-3表达水平变化。饲养8周后取肾脏皮质,RT-PCR法检测线粒体凋亡途径相关基因Cyt C、Caspase-9和Caspase-3表达量变化,内质网凋亡途径相关基因ATF-6、GRP78、IRE1α和 Caspase-12 表达量变化,Fas/FasL 凋亡途径相关基因 Fas、FasL、FADD、Caspase-8表达量变化;线粒体凋亡途径相关蛋白Cyt C、Cleaved caspase-9和Cleaved caspase-3表达水平变化,内质网凋亡途径相关蛋白ATF-6、GRP78、IRE1α和Cleaved caspase-12 表达水平变化,Fas/FasL 凋亡途径相关蛋白 Fas、FasL、FADD、Cleaved caspase-8表达水平变化。结果表明,镉染毒处理1周,与对照组相比,大鼠肾脏皮质细胞凋亡无显著变化(P0.05);镉染毒处理8周,与对照组相比,线粒体凋亡途径、内质网凋亡途径、Fas/FasL凋亡途径相关基因和蛋白表达水平都显著或极显著升高(P0.05或P0.01)。体外和体内研究表明,低剂量镉短时间处理激活细胞的保护机制,减少细胞凋亡,而长时间处理则同时激活线粒体凋亡途径、内质网凋亡途径和Fas/FasL凋亡途径,诱导细胞发生凋亡。3.镉诱导细胞自噬与细胞凋亡的关系采用机械筛网结合酶消化法建立原代大鼠肾小管上皮细胞(rPT cells)培养模型,在传一代细胞增殖活性最强时间进行处理。流式细胞术检测自噬抑制剂3-MA对镉处理rPT细胞6h凋亡率的影响、自噬激活剂RAPA对镉处理rPT细胞12h凋亡率的影响;免疫共沉淀法检测2.5 μM和5 μM镉处理rPT细胞6 h,Beclin-1与线粒体凋亡途径相关蛋白Cyt C、Caspase-3、Cleaved caspase-3、Caspase-9、Cleaved caspase-9、Bcl-2、Bax,与内质网凋亡途径相关蛋白 ATF-6、GRP78、IRE1α、Caspase-12、Cleavedcaspase-12 及 Fas/FasL 凋亡途径相关蛋白Fas、FasL、FADD、Caspase-8、Cleavedcaspase-8的相互作用;免疫印迹法检测Beclin-1敲减对镉诱导的自噬和凋亡的影响;DCFH-DA探针法检测镉诱导ROS生成量的变化;透射电子显微镜检测包裹线粒体的溶酶体数量变化。结果表明,自噬抑制剂3-MA极显著促进镉诱导rPT细胞凋亡(P0.01),同时自噬激活剂RAPA极显著抑制镉诱导rPT细胞凋亡(P0.01);2.5 μM和5 μM镉处理rPT细胞6 h,Beclin-1与线粒体凋亡途径相关蛋白CytCcyto、Caspase-9、Caspase-3、Baxmito,与内质网凋亡途径相关蛋白Caspase-12,与Fas/FasL凋亡途径相关蛋白Cleaved caspase-8存在明显相互作用,与Bcl-2Totai和Bcl-2Mito也存在明显相互作用,但呈负相关剂量效应关系,与其他蛋白不存在或仅存在极微弱相互作用;Beclin-1敲减导致LC3Ⅱ蛋白表达极显著下降(P0.01),p62蛋白表达极显著升高(P0.01),Cleavedcaspase-12、Cleavedcaspase-8、Cleavedcaspase-3表达极显著升高(P0.01);2.5 μM Cd和5 μM Cd处理rPT细胞6 h,5 μM镉组与2.5 μM镉组相比,ROS 水平显著降低(P0.05),包裹线粒体的溶酶体数量显著增多(PP0.05)。说明,自噬在镉诱导rPT细胞凋亡过程中通过Beclin-1发挥保护作用,同时通过清除受损线粒体、降低ROS水平发挥保护作用。
[Abstract]:Cadmium is a common high toxic heavy metal pollutant in the environment. Acute or chronic cadmium exposure can cause a variety of organs and tissue damage. In recent years, the cadmium content in the environment is rising rapidly because of the increase of cadmium production and use and the pollution caused by industrial wastes in industrial production. The half-life of cadmium in the environment is long. For 10-30 years, after entering the environment, it can not be degraded by microorganisms like organic pollutants and can eventually be stored in the human body through the food chain effect. Therefore, the harm caused by cadmium exposure to public health has attracted wide attention. The kidney is the target organ of the toxic effect of cadmium, and the domestic and foreign scholars from the environmental pollution, occupational exposure and animal test. The mechanism of renal toxicity induced by cadmium is extensively studied in many aspects, but these studies focus on the mechanism of apoptosis, and the mechanism of autophagy, the role of autophagy and the relationship with apoptosis in renal toxicity induced by cadmium are less.1. induced autophagy in the renal cortex and tubular epithelial cells of rats induced by cadmium in vitro And in the two part of the body. In vitro, the primary rat renal tubular epithelial cell (rPT cells) culture model was established by mechanical screening and enzyme digestion. In the strongest time of the first generation, the cell proliferation activity was treated with cadmium. At the same time, the NRK-52E cell line was selected for the same treatment.CCK-8 method to determine the different concentration of cadmium in different time to rP The effect of T cell and NRK-52E cell activity, MDC fluorescence probe was used to detect the effect of cadmium on the autophagy level of rPT cells and NRK-52E cells at different time of exposure. 2.5 Mu and 5 mu M cadmium treated rPT cells, 5 M and 10 micron cadmium treatment NRK-52E cells 6 h. The results showed that 2.5 M and 5 M cadmium treated rPT cells, 5 mu M and 10 mu cadmium treated NRK-52E cell 6h, the cell activity was the strongest, the level of autophagy was the highest, which was significantly higher than that of the control group (P0.01); 2.5 mu M and 5 micron cadmium treated rPT cells, 5 mu M and 10 micron cadmium treated cells, the level of autophagy was the lowest, significantly lower than the control group. 32 SD rats were randomly divided into 4 groups: the control group (DDW) and the cadmium group (CdAc2,50mg/kg. BW) for 1 weeks; the control group (DDW) and the cadmium group (CdAc2,50mg/kg. BW) were reared for 8 weeks. The renal cortex was taken to detect the expression of autophagy related genes LC3, p62 and Beclin-1, and the immunoblotting was used to detect the autophagy related protein LC3 II. The results showed that the autophagy level of renal cortex in rats was significantly higher than that of the control group (P0.01) after 1 weeks of cadmium treatment, and the expression level of Beclin-1 was significantly higher than that of the control group (P0.01). The autophagy level of renal cortex in rats was significantly lower than that of the control group (P0.01). In vitro and in vivo studies showed that low dose of cadmium in vitro and in vivo Time treatment can induce autophagy in renal cortex and renal tubular epithelial cells, while long time treatment inhibits the apoptosis of renal cortical and tubular epithelial cells of.2. induced by autophagy into two parts in vitro and in vivo. In vitro, the primary rat renal tubular epithelial cells (rPT cell) were established by mechanical sieve binding enzyme digestion method (rPT). S) culture model, cadmium treatment at the strongest time of cell proliferation in the first generation, and the effect of NRK-52E cell line on the same treatment.DAPI staining method to detect the apoptosis of rPT cells and NRK-52E cells at different concentration of cadmium at different time of exposure; the flow cytometry was used to test 2.5 mu M and 5 micron cadmium to treat rPT cells, 5 (mu M and 10 um M cadmium). The apoptosis rate of NRK-52E cells was 12 h and the rate of apoptosis was changed; rPT cells treated with 2.5 and 5 M were detected by Western blot and 12 h in NRK-52E cells treated with cadmium and 10 micron M. The apoptotic pathway related protein Cyt C, the apoptosis pathway related protein of the endoplasmic reticulum, and the apoptosis pathway related protein of the endoplasmic reticulum The expression level of aspase-12 expression, Fas/FasL apoptosis pathway related protein Fas, FasL, FADD, Cleavedcaspase-8 expression level changes. The results showed that 2.5 mu M and 5 mu M treated rPT cells, 5 mu M and 10 mu M cadmium treatment NRK-52E cells, the apoptosis rate began to rise, the apoptosis rate was the highest, 2.5 Mu and 5 mu cadmium cadmium. Treatment of rPT cells, 5 mu M and 10 u M cadmium treated NRK-52E cells 12 h. Compared with the control group, the mitochondrial apoptosis pathway, the apoptosis pathway of endoplasmic reticulum, and the expression level of Fas/FasL apoptosis pathway related proteins were significantly or significantly increased (P0.05 or P0.01). In vivo, 32 rats were randomly divided into 4 groups: control group (DDW) and cadmium group (CdAc2,50 mg/kg. For 1 weeks, the control group (DDW) and the cadmium group (CdAc2,50mg/kg BW) were reared for 8 weeks. After 1 weeks of feeding, the renal cortex was taken and the Caspase-3 expression of apoptosis gene was detected by RT-PCR method. The expression of Cleaved caspase-3 expression of apoptotic protein was detected by Western blot. After 8 weeks, the renal cortex was taken and RT-PCR method was used to detect the Cyt C of the mitochondrial apoptotic pathway related genes and C. C Changes in the expression of aspase-9 and Caspase-3, the changes in the expression of ATF-6, GRP78, IRE1, and Caspase-12 of the apoptosis pathway related genes of the endoplasmic reticulum, Fas/FasL apoptosis pathway related genes Fas, FasL, FADD, Caspase-8 expression. The expression levels of ATF-6, GRP78, IRE1, and Cleaved caspase-12, the expression level of Fas/FasL apoptosis pathway related proteins Fas, FasL, FADD, Cleaved caspase-8 were changed. The results showed that there was no significant change in renal cortical cell apoptosis (P0.05) with the control group for 1 weeks, compared with the control group (P0.05); cadmium treatment for 8 weeks, and control. Compared with the group, the apoptosis pathway of mitochondria, the apoptosis pathway of endoplasmic reticulum, and the level of Fas/FasL apoptosis pathway related genes and protein expression were significantly or significantly increased (P0.05 or P0.01). In vitro and in vivo, the study in vitro and in vivo showed that low dose cadmium was used for short time treatment to activate cell protection mechanism and reduce cell apoptosis, while long time treatment simultaneously activated mitochondria withering. The pathway of death, the apoptosis pathway of endoplasmic reticulum and the apoptosis pathway of Fas/FasL, inducing cell apoptosis and.3. cadmium induced autophagy to induce cell autophagy and cell apoptosis, the primary rat renal tubular epithelial cell (rPT cells) culture model was established by mechanical sieve binding enzyme digestion method, and the best time for cell proliferation in the first generation was processed. Flow cytometry The effect of autophagy inhibitor 3-MA on the apoptosis rate of 6h in rPT cells treated with cadmium, the effect of autophagy activator RAPA on the apoptosis rate of 12h in rPT cells treated with cadmium; the immunocoprecipitation method was used to detect the rPT cell 6 h of 2.5 mu M and 5 micron M, and Beclin-1 and mitochondrial apoptosis pathway related proteins Cyt. Bax, the interaction of ATF-6, GRP78, IRE1 alpha, Caspase-12, Cleavedcaspase-12 and Fas/FasL apoptosis pathway related protein Fas, FasL, FADD, Caspase-8, Cleavedcaspase-8, and immunoblotting to detect the effect of cadmium induced autophagy and apoptosis The changes in the amount of the lysosomes were detected by transmission electron microscopy. The results showed that the autophagy inhibitor 3-MA significantly promoted the apoptosis of rPT cells (P0.01) induced by cadmium, while the autophagy activator RAPA significantly inhibited the apoptosis of rPT cells induced by cadmium (P0.01); 2.5 mu M and 5 u M treated rPT cell 6 h, Beclin-1 and mitochondrial apoptosis Diameter related proteins CytCcyto, Caspase-9, Caspase-3, Baxmito, and the apoptosis pathway related protein Caspase-12 of endoplasmic reticulum, which interact with Fas/FasL apoptotic pathway related protein Cleaved caspase-8, and have obvious interaction with Bcl-2Totai and Bcl-2Mito, but there is a negative correlation of dose effect, which does not exist or exists only with other proteins. The expression of LC3 II protein decreased significantly (P0.01), the expression of p62 protein increased significantly (P0.01), the expression of Cleavedcaspase-12, Cleavedcaspase-8 and Cleavedcaspase-3 increased significantly (P0.01), and the 2.5 mu M Cd and 5 micron M cells treated 6 cells, and 5 mu cadmium group was significantly lower than the 2.5 micron cadmium group. 0.05) the number of lysosomes in the parcel mitochondria increased significantly (PP0.05). It shows that autophagy plays a protective role through Beclin-1 during the apoptosis of rPT cells induced by cadmium, and plays a protective role by removing the damaged mitochondria and reducing the level of ROS.
【学位授予单位】：扬州大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R114
[Abstract]:Cadmium is a common high toxic heavy metal pollutant in the environment. Acute or chronic cadmium exposure can cause a variety of organs and tissue damage. In recent years, the cadmium content in the environment is rising rapidly because of the increase of cadmium production and use and the pollution caused by industrial wastes in industrial production. The half-life of cadmium in the environment is long. For 10-30 years, after entering the environment, it can not be degraded by microorganisms like organic pollutants and can eventually be stored in the human body through the food chain effect. Therefore, the harm caused by cadmium exposure to public health has attracted wide attention. The kidney is the target organ of the toxic effect of cadmium, and the domestic and foreign scholars from the environmental pollution, occupational exposure and animal test. The mechanism of renal toxicity induced by cadmium is extensively studied in many aspects, but these studies focus on the mechanism of apoptosis, and the mechanism of autophagy, the role of autophagy and the relationship with apoptosis in renal toxicity induced by cadmium are less.1. induced autophagy in the renal cortex and tubular epithelial cells of rats induced by cadmium in vitro And in the two part of the body. In vitro, the primary rat renal tubular epithelial cell (rPT cells) culture model was established by mechanical screening and enzyme digestion. In the strongest time of the first generation, the cell proliferation activity was treated with cadmium. At the same time, the NRK-52E cell line was selected for the same treatment.CCK-8 method to determine the different concentration of cadmium in different time to rP The effect of T cell and NRK-52E cell activity, MDC fluorescence probe was used to detect the effect of cadmium on the autophagy level of rPT cells and NRK-52E cells at different time of exposure. 2.5 Mu and 5 mu M cadmium treated rPT cells, 5 M and 10 micron cadmium treatment NRK-52E cells 6 h. The results showed that 2.5 M and 5 M cadmium treated rPT cells, 5 mu M and 10 mu cadmium treated NRK-52E cell 6h, the cell activity was the strongest, the level of autophagy was the highest, which was significantly higher than that of the control group (P0.01); 2.5 mu M and 5 micron cadmium treated rPT cells, 5 mu M and 10 micron cadmium treated cells, the level of autophagy was the lowest, significantly lower than the control group. 32 SD rats were randomly divided into 4 groups: the control group (DDW) and the cadmium group (CdAc2,50mg/kg. BW) for 1 weeks; the control group (DDW) and the cadmium group (CdAc2,50mg/kg. BW) were reared for 8 weeks. The renal cortex was taken to detect the expression of autophagy related genes LC3, p62 and Beclin-1, and the immunoblotting was used to detect the autophagy related protein LC3 II. The results showed that the autophagy level of renal cortex in rats was significantly higher than that of the control group (P0.01) after 1 weeks of cadmium treatment, and the expression level of Beclin-1 was significantly higher than that of the control group (P0.01). The autophagy level of renal cortex in rats was significantly lower than that of the control group (P0.01). In vitro and in vivo studies showed that low dose of cadmium in vitro and in vivo Time treatment can induce autophagy in renal cortex and renal tubular epithelial cells, while long time treatment inhibits the apoptosis of renal cortical and tubular epithelial cells of.2. induced by autophagy into two parts in vitro and in vivo. In vitro, the primary rat renal tubular epithelial cells (rPT cell) were established by mechanical sieve binding enzyme digestion method (rPT). S) culture model, cadmium treatment at the strongest time of cell proliferation in the first generation, and the effect of NRK-52E cell line on the same treatment.DAPI staining method to detect the apoptosis of rPT cells and NRK-52E cells at different concentration of cadmium at different time of exposure; the flow cytometry was used to test 2.5 mu M and 5 micron cadmium to treat rPT cells, 5 (mu M and 10 um M cadmium). The apoptosis rate of NRK-52E cells was 12 h and the rate of apoptosis was changed; rPT cells treated with 2.5 and 5 M were detected by Western blot and 12 h in NRK-52E cells treated with cadmium and 10 micron M. The apoptotic pathway related protein Cyt C, the apoptosis pathway related protein of the endoplasmic reticulum, and the apoptosis pathway related protein of the endoplasmic reticulum The expression level of aspase-12 expression, Fas/FasL apoptosis pathway related protein Fas, FasL, FADD, Cleavedcaspase-8 expression level changes. The results showed that 2.5 mu M and 5 mu M treated rPT cells, 5 mu M and 10 mu M cadmium treatment NRK-52E cells, the apoptosis rate began to rise, the apoptosis rate was the highest, 2.5 Mu and 5 mu cadmium cadmium. Treatment of rPT cells, 5 mu M and 10 u M cadmium treated NRK-52E cells 12 h. Compared with the control group, the mitochondrial apoptosis pathway, the apoptosis pathway of endoplasmic reticulum, and the expression level of Fas/FasL apoptosis pathway related proteins were significantly or significantly increased (P0.05 or P0.01). In vivo, 32 rats were randomly divided into 4 groups: control group (DDW) and cadmium group (CdAc2,50 mg/kg. For 1 weeks, the control group (DDW) and the cadmium group (CdAc2,50mg/kg BW) were reared for 8 weeks. After 1 weeks of feeding, the renal cortex was taken and the Caspase-3 expression of apoptosis gene was detected by RT-PCR method. The expression of Cleaved caspase-3 expression of apoptotic protein was detected by Western blot. After 8 weeks, the renal cortex was taken and RT-PCR method was used to detect the Cyt C of the mitochondrial apoptotic pathway related genes and C. C Changes in the expression of aspase-9 and Caspase-3, the changes in the expression of ATF-6, GRP78, IRE1, and Caspase-12 of the apoptosis pathway related genes of the endoplasmic reticulum, Fas/FasL apoptosis pathway related genes Fas, FasL, FADD, Caspase-8 expression. The expression levels of ATF-6, GRP78, IRE1, and Cleaved caspase-12, the expression level of Fas/FasL apoptosis pathway related proteins Fas, FasL, FADD, Cleaved caspase-8 were changed. The results showed that there was no significant change in renal cortical cell apoptosis (P0.05) with the control group for 1 weeks, compared with the control group (P0.05); cadmium treatment for 8 weeks, and control. Compared with the group, the apoptosis pathway of mitochondria, the apoptosis pathway of endoplasmic reticulum, and the level of Fas/FasL apoptosis pathway related genes and protein expression were significantly or significantly increased (P0.05 or P0.01). In vitro and in vivo, the study in vitro and in vivo showed that low dose cadmium was used for short time treatment to activate cell protection mechanism and reduce cell apoptosis, while long time treatment simultaneously activated mitochondria withering. The pathway of death, the apoptosis pathway of endoplasmic reticulum and the apoptosis pathway of Fas/FasL, inducing cell apoptosis and.3. cadmium induced autophagy to induce cell autophagy and cell apoptosis, the primary rat renal tubular epithelial cell (rPT cells) culture model was established by mechanical sieve binding enzyme digestion method, and the best time for cell proliferation in the first generation was processed. Flow cytometry The effect of autophagy inhibitor 3-MA on the apoptosis rate of 6h in rPT cells treated with cadmium, the effect of autophagy activator RAPA on the apoptosis rate of 12h in rPT cells treated with cadmium; the immunocoprecipitation method was used to detect the rPT cell 6 h of 2.5 mu M and 5 micron M, and Beclin-1 and mitochondrial apoptosis pathway related proteins Cyt. Bax, the interaction of ATF-6, GRP78, IRE1 alpha, Caspase-12, Cleavedcaspase-12 and Fas/FasL apoptosis pathway related protein Fas, FasL, FADD, Caspase-8, Cleavedcaspase-8, and immunoblotting to detect the effect of cadmium induced autophagy and apoptosis The changes in the amount of the lysosomes were detected by transmission electron microscopy. The results showed that the autophagy inhibitor 3-MA significantly promoted the apoptosis of rPT cells (P0.01) induced by cadmium, while the autophagy activator RAPA significantly inhibited the apoptosis of rPT cells induced by cadmium (P0.01); 2.5 mu M and 5 u M treated rPT cell 6 h, Beclin-1 and mitochondrial apoptosis Diameter related proteins CytCcyto, Caspase-9, Caspase-3, Baxmito, and the apoptosis pathway related protein Caspase-12 of endoplasmic reticulum, which interact with Fas/FasL apoptotic pathway related protein Cleaved caspase-8, and have obvious interaction with Bcl-2Totai and Bcl-2Mito, but there is a negative correlation of dose effect, which does not exist or exists only with other proteins. The expression of LC3 II protein decreased significantly (P0.01), the expression of p62 protein increased significantly (P0.01), the expression of Cleavedcaspase-12, Cleavedcaspase-8 and Cleavedcaspase-3 increased significantly (P0.01), and the 2.5 mu M Cd and 5 micron M cells treated 6 cells, and 5 mu cadmium group was significantly lower than the 2.5 micron cadmium group. 0.05) the number of lysosomes in the parcel mitochondria increased significantly (PP0.05). It shows that autophagy plays a protective role through Beclin-1 during the apoptosis of rPT cells induced by cadmium, and plays a protective role by removing the damaged mitochondria and reducing the level of ROS.
【学位授予单位】：扬州大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R114
【相似文献】
相关期刊论文 前10条
1 王骏;黄锋先;余学清;;骨髓间充质干细胞共培养减轻顺铂所致的肾小管上皮细胞损伤[J];中国病理生理杂志;2008年07期
2 吴雄飞,王东,刘宏,范连慧,金锡御,宋关斌,吴泽志;肾小管上皮细胞模拟缺血缺氧时黏附力的改变[J];中华肾脏病杂志;2001年06期
3 黄全勇,黄巨恩;庆大霉素对培养的肾小管上皮细胞毒性作用的研究进展[J];解剖学研究;2003年01期
4 陈玲,贾汝汉,邱昌建,丁国华;脱氢抗坏血酸对高糖诱导肾小管上皮细胞产生氧自由基的影响[J];中华肾脏病杂志;2005年10期
5 李开龙,王建民,赵玲,张建国,何娅妮,杨聚荣,陈林;缺血再灌注损伤后肾小管上皮细胞的衰老演变及其意义[J];中华肾脏病杂志;2005年10期
6 罗昌霞;张永;张建鄂;李涛;;白蛋白诱导肾小管上皮细胞凋亡的实验研究[J];中国医师杂志;2006年05期
7 宋新文;邓存良;盛云建;陈枫;王明勇;史小玲;;外周血单个核细胞对肾小管上皮细胞凋亡影响研究[J];中国煤炭工业医学杂志;2006年03期
8 田少江;丁国华;陈铖;贾俊亚;梁伟;;表面活性蛋白A调节肾小管上皮细胞巨噬细胞炎症蛋白2的表达及其机制的探讨[J];中华肾脏病杂志;2006年05期
9 刘丽敏;刘华钢;黄巨恩;李校X;肖健;;酸性成纤维细胞生长因子对体外培养肾小管上皮细胞存活与增殖的影响[J];中国药理学通报;2007年06期
10 刘华钢;黄巨恩;刘丽敏;王慧杰;胡文娟;李校X;肖健;;酸性成纤维细胞生长因子对庆大霉素和顺铂所致肾小管上皮细胞损伤的保护作用[J];现代生物医学进展;2007年12期
相关会议论文 前10条
1 于澄钒;黄晓波;许清泉;陈亮;胡卫国;王晓峰;;纳米细菌对肾小管上皮细胞的损伤作用及晶体滞留的影响研究[A];第十七届全国泌尿外科学术会议论文汇编[C];2010年
2 刘燕;蒲蕾;杨立川;刘先蓉;樊均明;;洛伐他汀对转化生长因子-β诱导的肾小管上皮细胞-肌成纤维细胞转分化的影响[A];“中华医学会肾脏病学分会2004年年会”暨“第二届全国中青年肾脏病学术会议”论文汇编[C];2004年
3 朱加明;丁小强;张晓丽;刘春凤;;己糖胺通路活化介导肾小管上皮细胞表型和功能的改变[A];中华医学会肾脏病学分会2006年学术年会论文集[C];2006年
4 李健;李晓玫;王海燕;;低密度脂蛋白诱导的肾小管上皮细胞活化介导肾间质成纤维细胞生物学表型变化[A];中国细胞生物学学会第七次会议论文摘要汇编[C];1999年
5 朱吉莉;丁国华;王学玉;;血管紧张素Ⅱ通过抑制蛋白激酶B诱导肾小管上皮细胞凋亡[A];“中华医学会肾脏病学分会2004年年会”暨“第二届全国中青年肾脏病学术会议”论文汇编[C];2004年
6 杨晓;张春;邓安国;朱忠华;;结缔组织生长因子在肾小管上皮细胞细胞外基质积聚中的作用[A];“中华医学会肾脏病学分会2004年年会”暨“第二届全国中青年肾脏病学术会议”论文汇编[C];2004年
7 张瑜;周建华;;肝素对肾小管上皮细胞膜结合补体调节蛋白表达的影响及意义[A];中华医学会第十四次全国儿科学术会议论文汇编[C];2006年
8 杨雅丽;程晓霞;;肾小管上皮细胞凋亡在肾间质纤维化中的作用及其防治的研究进展[A];2009全国时间生物医学学术会议论文集[C];2009年
9 谭玉莉;黄妍;杨亦彬;;鱼胆汁毒素致肾小管上皮细胞死亡方式及实施干预的试验研究[A];贵州省医学会肾脏病学分会2008年学术年会论文汇编[C];2008年
10 乔f^;陈香美;吴镝;丁瑞;师锁柱;谢院生;洪权;吕杨;王兆霞;尹忠;;衰老大鼠肾脏缺血/再灌注损伤中肾小管上皮细胞不同凋亡途径的研究[A];“中华医学会肾脏病学分会2004年年会”暨“第二届全国中青年肾脏病学术会议”论文汇编[C];2004年
相关重要报纸文章 前1条
1 华红梅;药损肾先“知”[N];医药养生保健报;2006年
相关博士学位论文 前10条
1 倪s，
本文编号：2155918
本文链接：https://www.wllwen.com/shoufeilunwen/yxlbs/2155918.html
教材专著
