内皮素 A 受体（ETaR）下调通过增加一氧化氮产生来改善肾缺血再灌注损伤。
Downregulation of endothelin A receptor (ETaR) ameliorates renal ischemia reperfusion injury by increasing nitric oxide production.
机构信息
Department of Urology, Zhongshan Hospital, Fudan University, Shanghai 200032, China; Shanghai Key Laboratory of Organ Transplantation, Shanghai 200032, China; Department of Urology, Shanghai Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, China.
Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai 200030, China.
出版信息
Life Sci. 2019 Jul 1;228:295-304. doi: 10.1016/j.lfs.2019.05.013. Epub 2019 May 7.
AIMS
To investigate the protective effects of downregulating ETaR expression on renal ischemia reperfusion injury (IRI).
MAIN METHODS
The renal IRI model was generated by clamping the left renal artery for 60 min followed by nephrectomy of the right kidney. ETaR siRNA were perfused through the renal artery during ischemia. HE staining was performed to assess histological injury. PCR was performed to determine the expression of NF-κb, TNF-α, IFN-γ, IL-6 and TGF-β. ELISA was used to determine the levels of ET-1, TGF-β and eNOS. The level of nitric oxide (NO) was tested by the NO detection kit. The expression of PI3K, Akt, sGC and PKG were evaluated by western blot.
KEY FINDINGS
ETaR siRNA treatment reduced the levels of serum creatinine and urea nitrogen, decreased the number of apoptotic cells, and ameliorated histological damage after IRI. PCR results demonstrated that IRI increased mRNA levels of inflammatory factors, which were inhibited by ETaR siRNA treatment. ELISA result showed that ETaR siRNA decreased the levels of ET-1, TGF-β and eNOS in the renal tissues after IRI. Western blot results demonstrated that ETaR siRNA activated the PI3K/Akt and sGC/PKG signaling pathway. Conversely, the NOS inhibitor, L-NAME, reversed the effects of ETaR siRNA treatment.
SIGNIFICANCE
ETaR siRNA treatment inhibited inflammatory response and improved renal function after renal IRI. The underlying mechanisms of ETaR siRNA treatment may be through increasing eNOS activity through PI3K/Akt signaling, which subsequently increased NO production. The increased NO reduces the expression of ET-1 by inhibiting transcription of ET-1-associated genes via the sGC/PKG signaling pathway.
目的
研究下调内皮素 A 受体（ETaR）表达对肾缺血再灌注损伤（IRI）的保护作用。
主要方法
通过夹闭左肾动脉 60 分钟，然后切除右肾来建立肾 IRI 模型。在缺血期间通过肾动脉灌注 ETaR siRNA。通过 HE 染色评估组织学损伤。通过 PCR 确定 NF-κb、TNF-α、IFN-γ、IL-6 和 TGF-β 的表达。通过 ELISA 测定 ET-1、TGF-β 和 eNOS 的水平。通过 NO 检测试剂盒测定一氧化氮（NO）水平。通过 Western blot 评估 PI3K、Akt、sGC 和 PKG 的表达。
主要发现
ETaR siRNA 治疗降低了血清肌酐和尿素氮的水平，减少了细胞凋亡的数量，并改善了 IRI 后的组织学损伤。PCR 结果表明，IRI 增加了炎症因子的 mRNA 水平，而 ETaR siRNA 治疗抑制了这些水平。ELISA 结果表明，ETaR siRNA 降低了 IRI 后肾脏组织中 ET-1、TGF-β 和 eNOS 的水平。Western blot 结果表明，ETaR siRNA 激活了 PI3K/Akt 和 sGC/PKG 信号通路。相反，NOS 抑制剂 L-NAME 逆转了 ETaR siRNA 治疗的作用。
意义
ETaR siRNA 治疗抑制了肾 IRI 后的炎症反应并改善了肾功能。ETaR siRNA 治疗的潜在机制可能是通过 PI3K/Akt 信号增加 eNOS 活性，从而增加 NO 产生。增加的 NO 通过 sGC/PKG 信号通路抑制与 ET-1 相关基因的转录，从而降低 ET-1 的表达。
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