芍药苷抑制糖尿病视网膜病变基质金属蛋白酶9激活的研究

发布时间：2017-12-27 20:26

本文关键词：芍药苷抑制糖尿病视网膜病变基质金属蛋白酶9激活的研究　出处：《南京医科大学》2017年硕士论文　论文类型：学位论文

【摘要】：背景:糖尿病视网膜病变(DR)是糖尿病引起的并发症之一,是导致发达国家糖尿病患者后天视力丧失的主要原因。其病理表现为:视网膜细胞凋亡、视网膜神经功能紊乱、新生血管的大量生成等。尽管对这种病变的研究众多,但迄今对这种多因素疾病的确切机制还不清楚。小胶质细胞在DR疾病发展过程发挥着重要作用。基质金属蛋白酶(MMPs)能够降解细胞外基质,调节细胞内稳态平衡。基质金属蛋白9(MMP-9)损害血视网膜屏障(BRB),与DR的发展过程密切相关。芍药苷,属于单萜苷化合物,对小胶质细胞具有强大的免疫调节作用。本文我们研究芍药苷是否可通过抑制高糖刺激的小胶质细胞MMP-9的激活,从而改善DR。目的:研究芍药苷对糖尿病视网膜病变MMP-9激活的作用。实验方法:高糖刺激小胶质细胞,模拟体内糖尿病模型;腹腔注射STZ(链脲佐菌素)建立糖尿病小鼠模型;明胶酶谱检测MMP-9的活性;免疫印记法检测信号通路蛋白水平的表达;免疫荧光法检测NF-κB核转录情况;血糖仪测定小鼠血糖水平;HE染色观察小鼠视网膜形态结构。实验结果:高糖能够引起小胶质细胞MMP-9的激活,这种作用可被高迁移率族蛋白1(HMGB1)抑制剂、Toll样受体4(TLR-4)抑制剂、p38丝裂素活化蛋白激酶(p38MAPK)抑制剂、核转录因子-κB(NF-κB)抑制剂所取消。TLR-4抑制剂减少高糖诱导的小胶质细胞p38MAPK磷酸化水平。芍药苷增加热休克蛋白(HSP70)和细胞因子信号传导抑制蛋白-3(SOCS3)蛋白水平的表达,减少小胶质细胞MMP-9的表达。芍药苷诱发的SOCS3蛋白的表达被TLR-4抑制剂所取消。在STZ诱导的糖尿病鼠中,芍药苷能够诱发SOCS3蛋白表达,减少MMP-9激活。芍药苷抑制STZ诱导的IBA1,IL-1β表达,减少STZ诱导的糖尿病鼠血糖水平的升高,改善STZ诱导的糖尿病鼠视网膜的形态结构的改变。结论:高糖通过激活HMGB1-TLR4-NF-κB信号通路引起小胶质细胞MMP-9的激活,引起DR。芍药苷通过HSP70/TLR4/NF-κB信号通路诱发SOCS3表达,减少MMP-9激活,改善DR。
[Abstract]:Background: diabetic retinopathy (DR) is one of the complications caused by diabetes and is the main cause of the loss of acquired visual acuity in patients with diabetes in developed countries. The pathological features are retinal cell apoptosis, retinal nerve dysfunction, and a large number of neovascularization. Although there are many studies on this disease, the exact mechanism of this multifactor disease is not yet clear. Microglia play an important role in the development of DR disease. Matrix metalloproteinases (MMPs) can degrade extracellular matrix and regulate homeostasis in cells. Matrix metalloprotein 9 (MMP-9) damages the blood retinal barrier (BRB), which is closely related to the development of DR. Paeoniflorin, a monoterpene glycoside, has a strong immunomodulatory effect on microglia. In this article, we study whether paeoniflorin can improve the activation of MMP-9 in microglia stimulated by high glucose, thus improving DR. Objective: To study the effect of paeoniflorin on the activation of MMP-9 in diabetic retinopathy. Methods: high glucose stimulated microglia, simulated in vivo model of diabetes; intraperitoneal injection of STZ (STZ) to establish diabetic mouse model; gelatinase MMP-9 activity was detected; to detect the expression of signal transduction protein level by Western blot detection; NF- kappa B transcription by immunofluorescence method; Determination of blood glucose levels in mice blood glucose meter observation on the morphology of mouse retina; HE staining. Results: high glucose could induce the activation of microglia MMP-9, this effect was of high mobility group protein 1 (HMGB1) inhibitors, Toll like receptor 4 (TLR-4) inhibitor, p38 mitogen activated protein kinase (p38MAPK) inhibitor, nuclear factor kappa B (NF- K B) inhibitor of the cancelled. TLR-4 inhibitors reduce the level of p38MAPK phosphorylation in high glucose induced microglia. Paeoniflorin increases the expression of heat shock protein (HSP70) and cytokine signal transduction inhibitor protein -3 (SOCS3) protein, and reduces the expression of MMP-9 in microglia. The expression of paeoniflorin induced SOCS3 protein was cancelled by TLR-4 inhibitor. In STZ induced diabetic rats, paeoniflorin can induce the expression of SOCS3 protein and reduce the activation of MMP-9. Paeoniflorin inhibited the expression of IBA1 and IL-1 beta induced by STZ, reduced the increase of blood glucose level in diabetic rats induced by STZ, and improved the morphological structure of retina in STZ induced diabetic rats. Conclusion: high glucose induces the activation of MMP-9 in microglia by activating the HMGB1-TLR4-NF- kappa B signaling pathway, causing DR. Paeoniflorin induces SOCS3 expression through HSP70/TLR4/NF- kappa B signaling pathway, reducing MMP-9 activation and improving DR.
【学位授予单位】：南京医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285

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