【作者】 周洪文；

【导师】 甘华；

【作者基本信息】 重庆医科大学 ， 内科学， 2010， 硕士

【摘要】 目的:探讨连续性静脉静脉血液滤过(continuous veno-venous hemofiltration,CVVH)对多脏器功能衰竭(multiple organ dysfun- ction syndrome,MODS)患者血清体外诱导的脐静脉内皮细胞凋亡及内质网应激的影响及其机制。方法:实验分为健康对照组、CVVH治疗前组、CVVH治疗6h组、CVVH治疗20h组,分别用10名健康志愿者的血清及10名MODS患者CVVH治疗前、治疗6h、治疗20h的血清体外诱导脐静脉内皮细胞(HUVEC)24h,TUNEL法检测HUVEC细胞凋亡率,免疫荧光观察细胞GRP78蛋白表达,RT-PCR检测细胞GRP78、CHOP、TNF-α、IL-6 mRNA表达量,比色法检测健康志愿者及MODS患者CVVH治疗不同时间组血清中SOD活性、MDA含量。ELISA法检测检测健康志愿者及MODS患者CVVH治疗不同时间组血清中TNF-α、IL-6含量。结果:与健康对照组相比,CVVH治疗前组脐静脉内皮细胞凋亡率明显升高(p<0.01),细胞GRP78、CHOP、IL-6、TNF-αmRNA表达量也显著增高(p<0.01),免疫荧光显示脐静脉内皮细胞GRP78蛋白表达明显增多。与CVVH治疗前组比较,CVVH治疗6h组、20h组脐静脉内皮细胞凋亡率明显下降(p<0.01),细胞GRP78、CHOP、IL-6、TNF-αmRNA表达量明显减少(p<0.01),治疗20h组较治疗6h组进一步减少(p<0.01)。免疫荧光显示脐静脉内皮细胞GRP78蛋白表达治疗6h组较治疗前组明显减少,治疗20h组较治疗6h组进一步减少。与健康对照组比较,CVVH治疗前组患者血清SOD活性明显下降(p<0.01),MDA含量明显增高(p<0.01),血清中TNF-α、IL-6含量也明显增高(p<0.01)。经过CVVH治疗,MODS患者血清SOD活性明显升高(p<0.01), MDA含量明显下降(p<0.01),TNF-α、IL-6含量也明显下降(p<0.01)。结论:MODS患者血清体外诱导的HUVEC细胞凋亡与内质网应激有关,CVVH治疗能减轻氧化应激,清除炎症介质,减轻内质网应激程度,从而减少内皮细胞凋亡。更多还原

【Abstract】 AIM:To investigate the impact and mechanism of continuous veno-venous hemofiltration(CVVH) on endoplasmic reticulum stress(ERS) and apoptosis of HUVEC cells which induced by serum of patients with multiple organ dysfunction syndrome(MODS).METHODS:10 patients with MODS who received CVVH and 10 healthy volunteers were involved in this study. This experiment was divided into four groups: healthy control,group of pre-CVVH,group of 6-hour-CVVH and group of 20-hour-CVVH. Serum of healthy volunteers and serum of patients with MODS at pre- CVVH,6-hour-CVVH and 20-hour-CVVH were used to induced HUVEC cells respectively in vitro for 24h. The cell apoptosis was examined by TUNEL assay,the intracellular GRP78 protein was observed by immunocytochemistry,the mRNA expression of GRP78, CHOP,TNF-α,IL-6 of cells were measured by RT-PCR, The level of Serum SOD,MDA was detected by colorimetric method, the level of TNF-α,IL-6 of serum was detected by ELISA. RESULTS:Compared with healty control, the group of pre-CVVH showed higher cellular apoptosis rate(p<0.01),the mRNA expression of GRP78,CHOP,TNF-α,IL-6 of cells increased significantly(p<0.01), protein expression of GRP78 increased obviously.Compared with group of pre-CVVH, cellular apoptosis rate decreased significantly(p<0.01)and the mRNA expression of GRP78,CHOP,TNF-α,IL-6 of cells decreased significantly(p<0.01)either in the group of 6-hour-CVVH or group of 20-hour-CVVH. The mRNA expression of GRP78,CHOP,TNF-α,IL-6 of cells in group of 20-hour-CVVH were significantly lower than those in group of 6-hour-CVVH(p<0.01). Immunocytochemistry showed protein expression of GRP78 in group of 6-hour-CVVH was significantly lower than that in group of pre-CVVH, protein expression of GRP78 in group of 20-hour-CVVH was significantly lower than that in group of 6-hour-CVVH. Compared with healthy control, the level of SOD of serum decreased significantly and the level of MDA of Serum increased significantly in the group of pre-CVVH(p<0.01), the level of TNF-α,IL-6 of serum also increased significantly in the group of pre-CVVH(p<0.01),after CVVH, the level of SOD of serum increased significantly and the level of MDA of serum decreased significantly(p<0.01), the level of TNF-α,IL-6 of serum also decreased significantly (p<0.01).CONCLUSION:Apoptosis of HUVEC cells induced by serum of patients with MODS was associated with ERS,CVVH can relieve redox state, remove inflammatory cytokine, relieve ERS, this may be one of mechanisms of CVVH relieve damage of HUVEC cells induced by serum of patients with MODS.更多还原