异阿克替苷对LPS诱导脓毒症性急性肾损伤的保护作用及机制研究

发布时间：2018-10-05 13:05

【摘要】：目的:探究异阿克替苷对内毒素导致的脓毒症性急性肾损伤(AKI)的治疗作用及作用机制,为脓毒症性AKI的治疗提供实验依据。方法:本实验采用单次腹腔注射LPS(10mL/kg)的方法制备小鼠脓毒症性AKI模型。将60只雄性小鼠随机分为6组:空白组,模型组(LPS组),异阿克替苷(Isoacteoside,IAC)高、中、低剂量组(200、100、50mL/kg,IAC 组),阳性药(地塞米松,dexamethasone,DEX,5mL/kg)组(DEX组),每组10只。IAC组在造模前的3h,造模后的3h、6h腹腔皮下注射给药,阳性药组于造模前的1h给药,其他各组同时给予等体积生理盐水。造模后观察小鼠的一般状况,造模12h后处死小鼠,取材,测量各组小鼠的肾指数,测定各组小鼠血清肌酐(Crea)、谷草转氨酶(ALT)、谷丙转氨酶(AST)、血尿素氮(BUN)的水平,HE染色观察肾脏病理组织形态学异常;应用酶联免疫法测定小鼠血清中的肿瘤坏死因子(TNF-a)及白细胞介素-1β(IL-1β)的水平;免疫印迹法检测p-IκB-a和NF-κB/p65蛋白的表达水平。结果:1.HE染色结果显示.与空白组相比LPS组小鼠肾组织出现肾小管上皮细胞肿胀,肾间质水肿,肾小球萎缩,有少量炎性细胞浸润;与LPS组相比,IAC组和DEX组均能有效改善小鼠肾组织的肾间质水肿和炎性细胞浸润情况。2.测量各组小鼠肾指数的结果为:与空白组相比,LPS组肾指数明显升高;与LPS组相比,各给药组小鼠肾指数明显下降,均具有统计学意义,且IAC各剂量组呈一定剂量依赖性。3.测量各组小鼠BUN和Crea水平的结果:LPS组小鼠BUN和Crea含量与空白组相比显著上升;与LPS组相较,IAC组和DEX组血清Crea和BUN水平均显著下降,具有统计学意义。4.测量各组小鼠ALT和AST活性的结果:与空白组相比,LPS组血清ALT和AST活性均显著上升;与LPS组相比,IAC组和DEX组ALT和AST活性均显著下降,均具有统计学意义。5.测量各组小鼠血清TNF-α及IL-1β含量的结果:LPS组与空白组相比TNF-α和IL-1β水平均显著上调;与LPS组相比,IAC组和DEX组均能显著下调血清中TNF-α和IL-1β的水平,均具有统计学意义。6.测量各组小鼠肾组织NF-κB信号通路蛋白表达水平的结果:LPS组的细胞总p-IκB-a和核NF-κB/P65蛋白表达水平相比空白组显著上调;与LPS组比较,IAC组和DEX组的细胞总p-IκB-a和核NF-κB/P65蛋白表达水平有显著下调。可见,异阿克替苷和地塞米松抑制了 NF-κB信号通路的活化。结论:1.异阿克替苷能改善肾功能,减轻肾损伤,明显抑制TNF-α和IL-1β释放,具有保护脓毒症性急性肾损伤的作用。2.异阿克替苷能通过抑制NF-κB信号通路的活化,抑制脓毒症小鼠炎症反应保护脓毒症性急性肾损伤。
[Abstract]:Aim: to investigate the therapeutic effect and mechanism of isoapatidine on (AKI) induced by endotoxin in septic acute renal injury, and to provide experimental evidence for the treatment of septic AKI. Methods: the septic AKI model of mice was established by single intraperitoneal injection of LPS (10mL/kg). Sixty male mice were randomly divided into 6 groups: blank group, model group (LPS group), iso-azotoside (Isoacteoside,IAC) high, middle, high, middle and middle groups. In the low dose group (200,100ml / kg), the positive drug (dexamethasoneone dexone 5mL / kg) group (DEX group), 10 rats in each group were given intraperitoneally subcutaneously at 3 hours before modeling, 3 hours after modeling, and 1 hour before modeling in the positive drug group, and at the same time, the other groups were given the same volume of normal saline at the same time. The general condition of the mice was observed after modeling. The mice were killed 12 hours later and the kidney index of each group was measured. Serum creatinine (Crea),) glutamic oxaloacetic transaminase (ALT),) (ALT), alanine aminotransferase (AST),) blood urea nitrogen (BUN) were measured by HE staining. The levels of tumor necrosis factor (TNF-a) and interleukin-1 尾 (IL-1 尾) in serum of mice were determined by enzyme linked immunosorbent assay (Elisa), and the expression of p-I 魏 B-a and NF- 魏 B/p65 were detected by Western blot. Results 1. The results of HE staining showed that. Compared with the blank group, LPS group showed tubuloepithelial cell swelling, renal interstitial edema, glomerular atrophy and a small amount of inflammatory cell infiltration. Compared with LPS group and DEX group, the renal interstitial edema and inflammatory cell infiltration in mice were significantly improved. The results of measuring the renal index of each group were as follows: compared with the blank group, the renal index in the lipopolysaccharide group was higher than that in the control group, and the renal index in each administration group was significantly lower than that in the LPS group, and the renal index of each dose group of IAC was in a dose-dependent manner. Compared with the control group, the BUN and Crea levels in the BUN and Crea groups were significantly increased, and the serum Crea and BUN levels in the LPS and DEX groups were significantly lower than those in the LPS group, with statistical significance. 4. The results of measuring ALT and AST activities in each group: compared with the blank group, the serum ALT and AST activities in the lipopolysaccharide group increased significantly, and the ALT and AST activities in the LPS group and DEX group decreased significantly (P < 0.05). Results the levels of TNF- 伪 and IL-1 尾 in serum of mice in the control group were significantly higher than those in the control group, and the levels of TNF- 伪 and IL-1 尾 in serum were significantly decreased in LPS group and DEX group (P < 0.05), and were significantly lower than those in the control group (P < 0.05), and the levels of TNF- 伪 and IL-1 尾 in the control group were significantly lower than those in the control group (P < 0.05). The expression level of NF- 魏 B signaling pathway protein in the kidney tissue of mice in each group was measured. The expression of total p-I 魏 B-a and nuclear NF- 魏 B/P65 protein in the cells of the control group was significantly higher than that in the control group. Compared with LPS group, the expression of total p-I 魏 B-a and nuclear NF- 魏 B/P65 protein in LPS group and DEX group were significantly down-regulated. The activation of NF- 魏 B signaling pathway was inhibited by isoazoside and dexamethasone. Conclusion 1. Isoazoside can improve renal function, alleviate renal injury, inhibit the release of TNF- 伪 and IL-1 尾, and protect acute renal injury induced by sepsis. Isoazoside can protect septic acute renal injury by inhibiting the activation of NF- 魏 B signaling pathway and inhibiting inflammatory response in septic mice.
【学位授予单位】：黑龙江中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R459.7

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