目的 探索腎康注射液對慢性腎衰竭大鼠腎損傷及腎纖維化的作用機(jī)制。方法 采用腺嘌呤法建立大鼠慢性腎衰竭模型，采用叔丁基過氧化氫（TBHP）誘導(dǎo)NRK-52E細(xì)胞，建立慢性腎衰竭腎損傷細(xì)胞模型。采用蘇木精–伊紅（HE）染色評(píng)估腎康注射液對慢性腎衰竭大鼠的腎臟病理學(xué)損傷，免疫組化法檢測模型大鼠腎損傷及纖維化的關(guān)鍵蛋白表達(dá)，免疫熒光法檢測NRK-52E細(xì)胞葡萄糖調(diào)節(jié)蛋白78（GRP78）蛋白表達(dá)，Western blotting法進(jìn)行腎組織和細(xì)胞中的關(guān)鍵蛋白定量，活性氧熒光探針檢測NRK-52E細(xì)胞中的活性氧（ROS）水平，采用JC-1檢測試劑盒檢測NRK-52E細(xì)胞中線粒體膜電位水平。結(jié)果 腎康注射液可以顯著緩解慢性腎衰竭大鼠的腎間質(zhì)纖維化狀態(tài)，減輕病理損傷；顯著抑制腎損傷分子1（KIM-1）、GRP78、內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)蛋白（CHOP）陽性表達(dá)表達(dá)，從而抑制內(nèi)質(zhì)網(wǎng)過度應(yīng)激狀態(tài)；可顯著抑制I型膠原蛋白（Collagen I）沉積，抑制α-平滑肌肌動(dòng)蛋白（α-SMA）表達(dá)，從而抑制腎間質(zhì)纖維化形成（P＜0.01）；腎康注射液能夠顯著抑制NRK-52E細(xì)胞內(nèi)ROS的水平，減輕NRK-52E線粒體氧化損傷，穩(wěn)定線粒體膜電位水平（P＜0.01）。結(jié)論 腎康注射液能夠減輕腎間質(zhì)纖維化、緩解腎損傷狀態(tài)，增強(qiáng)腎功能，其機(jī)制可能與抑制腎臟過度的內(nèi)質(zhì)網(wǎng)應(yīng)激，抗氧化損傷以及減少肌成纖維細(xì)胞的增殖和減少細(xì)胞外基質(zhì)的沉積實(shí)現(xiàn)有關(guān)。

Objective To explore the protective effect of Shenkang Injection on renal injury and renal fibrosis caused by chronic renal failure and its mechanism. Methods The rat model of chronic renal failure was established by adenine method, and NRK-52E cells were induced by TBHP. HE staining was used to evaluate the renal pathological injury of Shenkang Injection on rats with chronic renal failure. The expression of key proteins in renal injury and fibrosis was detected by immunohistochemical method. The expression of GRP78 in NRK-52E cells was detected by immunofluorescence method. Western Blotting method was used to quantify key proteins in tissues and cells, ROS levels in NRK-52E cells were detected by ROS fluorescent probes, and mitochondrial membrane potential levels in NRK-52E cells were detected by JC-1 detection kit. Results Shenkang Injection can significantly alleviate the renal interstitial fibrosis of chronic renal failure and alleviate pathological injury. It can significantly inhibit the positive expressions of KIM-1, GRP78, and CHOP, so as to inhibit the overstress state of ER. Collagen I deposition and α-SMA were significantly inhibited, thusinhibiting the formation of renal interstitial fibrosis (P < 0.01). Shenkang Injection could significantly inhibit the ROS level in NRK-52E cells, alleviate the mitochondrial oxidative damage of NRK-52E, and stabilize the mitochondrial membrane potential (P < 0.01). Conclusion Shenkang Injection can alleviate renal interstitial fibrosis, alleviate renal injury and enhance renal function, the mechanism of which may be related to inhibiting excessive endoplasmic reticulum stress, anti-oxidative injury, reducing proliferation of myofibroblasts and reducing ECM deposition.

R285

國家自然科學(xué)基金資助項(xiàng)目（82104515）;河南省高等學(xué)校重點(diǎn)科研項(xiàng)目（23A360018）