基于TGR5介导的NLRP3炎症小体探讨芪参汤抑制巨噬细胞M1型极化改善非酒精性脂肪性肝炎的机制

目的:观察芪参汤对TGR5介导的NLRP3炎症小体的影响,从而明确其抑制巨噬细胞M1型极化改善非酒精性脂肪性肝炎的分子机制。方法:小鼠巨噬细胞RAW264.7随机分为空白组、模型组、芪参汤组、TGR5激动剂组和芪参汤+TGR5激动剂组。除空白组外,余下各组通过棕榈酸诱导构建巨噬细胞NLRP3活化模型,并给予相应的药物进行干预。分别采用酶联免疫吸附试验检测巨噬细胞上清中TNF-α、IL-6、IL-1β和CXCL2的含量,流式细胞术检测巨噬细胞极化标记分子CD86和iNOS的表达水平,Western blot检测TGR5/STAT1/STAT6信号通路和NLRP3炎症小体蛋白的表达丰度。结果:与空白组相比,模型组巨噬细胞TNF-α、IL-6、IL-1β、CXCL2的含量和CD86、iNOS阳性表达的巨噬细胞比例均显著增加,差异均具有统计学意义(P<0.01)。与模型组比较,芪参汤组TNF-α、IL-6、IL-1β、CXCL2的含量和CD86、iNOS阳性表达的巨噬细胞比例均显著降低,差异均具有统计学意义(P<0.01)。此外,与空白组相比,模型组巨噬细胞裂解液中NLRP3、Pro-IL-1β蛋白的表达和细胞上清液中Caspase-1 p10、p20及IL-1βp17蛋白的表达均显著增加,差异均具有统计学意义(P<0.01)。与模型组比较,芪参汤组巨噬细胞裂解液中NLRP3、Pro-IL-1β蛋白的表达和细胞上清液中Caspase-1 p10、p20及IL-1βp17蛋白的表达均显著降低,差异均具有统计学意义(P<0.01)。结论:参汤能够通过抑制TGR5/STAT1/STAT6信号通路抑制巨噬细胞NLRP3炎症小体的活化,从而抑制巨噬细胞M1型极化,改善炎症反应。

Mechanism of Qishen Decoction inhibition of macrophage M1 type polarization by targeting TGR5-mediated NLRP3 inflammasome

Objective:To observe the effect of Qishen decoction on TGR5-mediated activation of NLRP3 inflammasome,so as to clarify the molecular mechanism of its inhibition of macrophage M1-type polarisation to ameliorate non-alcoholic steatohepatitis;Methods:Mouse macrophage cell line RAW264.7 was randomly divided into a control group,model group,Qishen decoction group,TGR5 agonist group and Qishen decoction+TGR5 agonist group.Except for the control group,the remaining groups were constructed the macrophage NLRP3 activation model by palmitic acid induction,and the corresponding drugs were given to intervene.ELISA was used to detect the levels of TNF-α,IL-6,IL-1βand CXCL2 in macrophage supernatants,flow cytometry was used to detect the expression levels of macrophage polarisation marker molecules CD86 and iNOS,and Western blot was used to detect the expression of the TGR5/STAT1/STAT6 signaling pathway and the expression of NLRP3 inflammasome-associated proteins,respectively.Results:Compared with the control group,the contents of macrophages TNF-α,IL-6,IL-1β,CXCL2 and the proportion of macrophages with positive expression of CD86 and iNOS were significantly increased in the model group,and the differences were all statistically significant(P<0.01).Compared with the model group,the contents of TNF-α,IL-6,IL-1β,CXCL2 and the proportion of macrophages with positive expression of CD86 and iNOS were significantly decreased in the Qishen decoction group,and the differences were all statistically significant(P<0.01).In addition,the expression of NLRP3 and Pro-IL-1βproteins in the macrophage lysate and the expression of Caspase-1 p10,Caspase-1 p20 and IL-1βp17 proteins in the cell supernatant of the model group were significantly increased when compared with the control group,and the differences were all statistically significant(P<0.01).Compared with the model group,the expression of NLRP3 and Pro-IL-1βproteins in macrophage lysate and the expression of Caspase-1 p10,Caspase-1 p20 and IL-1βp17 proteins in cell supernatant of the Qishen decoction were significantly reduced,and the differences were all statistically significant(P<0.01);Conclusion:Qishen decoction can inhibit the activation of NLRP3 inflammasome in macrophages by inhibiting the TGR5/STAT1/STAT6 signaling pathway,thereby inhibiting macrophage M1 polarization and improving inflammatory response.

探讨中西医结合治疗糖尿病合并急性脑梗死的疗效观察

目的研究分析糖尿病合并急性脑梗死患者接受中西结合治疗方式的临床效果,为临床治疗提供参考。方法选取了2017年7月—2018年8月该院的糖尿病合并急性脑梗死患者进行分析,将患者分成了对照组和观察组,均有40例,对照组使用西医治疗,观察组使用中西结合治疗,比较两组的治疗效果。结果经过治疗,观察组患者的NIHSS评分平均是(8.3±0.9)分,对照组患者的NIHSS评分平均是(11.5±1.2)分,差异有统计学意义(P<0.05)。观察组患者的空腹血糖和餐后2h血糖治疗后比对照组,差异有统计学意义(P<0.05)。结论糖尿病合并急性脑梗死患者接受中西结合治疗方式的效果比较好,患者的神经功能概述明显,血糖指标得到了改善,预后效果有所改善,临床中可以进行推广使用。

脑心清片治疗缺血性脑卒中恢复期多中心随机双盲对照研究

目的 观察脑心清片治疗缺血性脑卒中恢复期的有效性及安全性。方法 本研究方案在中国临床试验中心注册(No.ChiCTR-TRC-13003968)。筛选721例受试者,选择缺血性脑卒中发病2周~3个月瘀血阻络证患者654例,采用随机对照方法,使用SAS软件区组随机分为试验组、对照组和安慰剂组,各218例,在西医常规治疗基础上,分别给予脑心清片+银杏叶片模拟剂(试验组)、脑心清片模拟剂+银杏叶片(对照组)和脑心清片模拟剂+银杏叶片模拟剂(安慰剂组),三组用法用量均为3片/次,3次/日,疗程12周。观察美国国立卫生研究院卒中量表NIHSS评分、卒中专门生命质量SSQOL量表评分、改良Rankin评分、日常生活能力Barthel指数、中医证候评分及不良反应发生率。结果 本研究共纳入分析654例,各组均为218例。治疗6周及12周后,试验组在改善NIHSS评分、SSQOL评分、改良Rankin量表评分、Barthel指数方面与对照组疗效相当(P>0.0167),但均优于安慰剂组(P<0.0167)。治疗12周后试验组中医证候有效率高于对照组、安慰剂组(P<0.0167);在治疗6周及12周后对失眠多梦的有效率试验组高于对照组(P<0.0167);在治疗12周后对下肢不遂、上肢不遂、口唇紫暗的有效率试验组高于安慰剂组(P<0.0167)。治疗12周后三组患者凝血四项(PT,APTT,TT,FIB)、血脂四项(TC,TG, LDL-C,HDL-C)、血糖及血液流变学指标疗后差值,组间差异均无统计学意义(P>0.05)。不良事件试验组9例(4.13%),对照组3例(1.38%),安慰剂组8例(3.67%,P=0.2021),不良反应安慰剂组1例,其他两组未发生不良反应。结论 脑心清片能改善缺血性脑卒中恢复期患者的神经功能缺损症状,提高患者生活质量和生活能力,降低致残率,改善中医证候,其疗效与银杏叶片相当,且安全性良好。