TLR4在高糖加重缺血性肾小管损伤中的作用及机制

Recent epidemiological research has reported that diabetic patients showed higher injury than non-diabetic patient after acute kidney injury (AKI) induced by ischemia, but the mechanism is unknown. Our previous study suggested that ischemic AKI induced more severe kidney injury and higher tubules apoptosis in Akita diabetic mice than wild type non-diabetic mice. In addition, ischemic kidney injury in diabetic mice showed a correlation with their fasting blood glucose levels, which indicated that hyperglycemia was an important factor in the sensitivity changes in diabetes. However, the mechanism underlying the AKI sensitivity of diabetic patients is not clear. Following AKI, renal tubular epithelial cells express increased amounts of TLR4, which enhences inflammation and further promotes kidney injury. Reports also suggest that high glucose or diabetes activates TLR4 in kidney tubular cells. High glucose or diabetes might increase the kidney injury induced by ischemia by promoting TLR4 mediated inflammation during AKI, which needs further investigation. In this study, we will study the role of TLR4 mediated inflammation in sensitivity changes under diabetic condition, try to inhibit TLR4 mediated inflammation during AKI, and further reduce kidney injury level in diabetic patients.

临床研究证实：糖尿病患者发生缺血性肾损伤后，肾损伤程度远重于非糖尿病患者，但相关机制不明。申请人前期研究显示：糖尿病小鼠发生缺血性肾损伤后，血肌酐、尿素氮及肾小管上皮细胞凋亡率均远高于非糖尿病小鼠，且肾损伤程度与小鼠空腹血糖成正比，提示高血糖在此过程中起重要作用。但高糖加重缺血性肾损伤的机制尚不清楚。缺血性肾损伤时肾小管上皮细胞TLR4蛋白表达升高，TLR4可通过促进缺血性肾损伤过程中的炎症反应，加重肾损伤程度，提示TLR4炎症反应通路在缺血性肾损伤过程中起重要作用；同时研究提示高糖或糖尿病可增加肾小管细胞TLR4蛋白表达。高糖或糖尿病加重缺血性肾损伤是否也与TLR4蛋白上调、炎症反应增强相关呢？本课题拟在前期研究的基础上，重点研究TLR4炎症反应通路在高糖加重缺血性肾损伤中的作用，并寻找有效方法抑制TLR4炎症反应通路，从而减轻糖尿病患者发生缺血性肾损伤后肾脏损伤程度。

临床研究证实：糖尿病患者发生缺血性肾损伤后，肾损伤程度远重于非糖尿病患者，但相关机制不明。申请人前期研究显示：糖尿病小鼠发生缺血性肾损伤后，血肌酐、尿素氮及肾小管上皮细胞凋亡率均远高于非糖尿病小鼠，且肾损伤程度与小鼠空腹血糖成正比，提示高血糖在此过程中起重要作用。但高糖加重缺血性肾损伤的机制尚不清楚。本课题拟在前期研究的基础上，重点研究TLR4炎症反应通路在高糖加重缺血性肾损伤中的作用，并寻找有效方法抑制TLR4炎症反应通路，从而减轻糖尿病患者发生缺血性肾损伤后肾脏损伤程度。.体外研究实验结果显示：NRK-52E肾小管上皮细胞经低糖（5.5mM），高糖（30mM）培养基处理后，高糖组细胞TLR4表达水平升高，使用抗霉素A诱导小管细胞发生凋亡，高糖组NRK-52E肾小管上皮细胞凋亡水平及caspase活性水平要显著高于低糖组细胞；凋亡损伤后，NRK-52E细胞TLR4蛋白、TLR4 mRNA、MCP-1、MyD88水平均升高，高糖组细胞凋亡后的TLR4蛋白、TLR4 mRNA、MCP-1、MyD88水平显著高于低糖组细胞。TLR4抑制剂TAK242或TLR4 shRNA可显著降低高糖组细胞经抗霉素A诱导的细胞凋亡水平。.体内研究结果显示：Akita糖尿病小鼠及非糖尿病小鼠发生急性缺血性肾损伤后，糖尿病小鼠肌酐及尿素氮水平显著高于非糖尿病小鼠，肾组织TLR4、MCP-1、MyD88蛋白水平显著升高，缺血性肾损伤后Akita糖尿病小鼠TLR4蛋白表达水平显著高于非糖尿病小鼠，TLR4抑制剂TAK242可有效抑制糖尿病小鼠缺血性肾损伤后TLR4高表达，同时TAK242可有效降低Akita糖尿病小鼠缺血性肾损伤后的肌酐、尿素氮及肾小管细胞凋亡水平，并降低MCP-1、MyD88蛋白水平。.以上研究证实：高糖或糖尿病可通过激活TLR4蛋白，增强TLR4介导的炎症反应，从而加重缺血性肾小管损伤的程度，TLR4抑制剂可有效减轻糖尿病小鼠缺血性肾损伤后肾损伤程度。提示寻找有效方法抑制TLR4炎症反应通路，对于减轻糖尿病患者缺血性肾损伤的程度、延缓糖尿病患者进展至终末期肾病的速度具有重要的意义。