目的 在順鉑誘導的大鼠急性腎損傷模型中,系統(tǒng)評價短時間采尿下系列尿液腎毒性生物標志的診斷性能。方法 Wistar大鼠單次ip 6 mg·kg^-1順鉑構建急性腎損傷模型,在檢疫期、給藥后第3和6天分別收集給藥后5 h對照組和順鉑組動物尿液和血樣,使用日立7180全自動生化儀測定血清中丙氨酸氨基轉移酶(ALT)、天冬氨酸氨基轉移酶(AST)、總膽紅素(TBIL)、尿素氮(BUN)、肌酐(CRE)以及尿液樣本中尿葡萄糖苷酶(NAG)、尿總蛋白(uTP)、尿肌酐(uCr)水平;使用Luminex儀器測定尿液中叢生蛋白(CLU)、谷胱甘肽S轉移酶-α(GST-α)、干擾素誘導蛋白-10(IP-10)、腎損傷因子-1(KIM-1)、骨橋蛋白(OPN)、組織金屬蛋白酶抑制劑-1(TIMP-1)、血管內皮生長因子-A(VEGF-A)、酸性糖蛋白(AGP)、白蛋白(Alb)、β₂微球蛋白(β₂M)、半胱氨酸蛋白酶抑制劑C(CysC)、表皮生長因子(EGF)、中性粒細胞明膠酶相關脂質運載蛋白(NGAL)水平。結合動物腎臟組織病理學檢查,制作受試者操作特性曲線(ROC)進行腎毒性標志物的靈敏度和特異性分析。結果 順鉑組動物AST、ALT、TBIL值與各自對照組比較無統(tǒng)計學差異,且解剖大體檢查并未發(fā)現除腎臟以外其他組織器官病變,排除其他組織器官病變對腎生物標志物變化的影響。組織病理學結果證實順鉑誘導動物模型均出現典型急性腎損傷:外髓質外帶腎小管上皮細胞變性/壞死和髓質蛋白管型等。與對照組比較,傳統(tǒng)標志物血清BUN和CRE在順鉑給藥第6天、動物嚴重腎損傷時才出現顯著升高(P<0.05);而IP-10、KIM-1、Alb、β2M和CLU生物標志物則在給藥第3天已出現顯著增加(P<0.05),且持續(xù)升高,增幅明顯高于傳統(tǒng)指標。ROC分析結果表明,IP-10、CysC、KIM-1和Alb的曲線下面積(AUC)明顯優(yōu)于BUN和CRE,具有更高的靈敏度和特異性。結論 尿液IP-10、CysC、KIM-1和Alb的腎毒性診斷性能優(yōu)于傳統(tǒng)生物標志物BUN和CRE,建議可作為藥物致急性腎毒性早期診斷的生物標志物。

Objective To systematically evaluate the diagnostic performance of a series of newly reported urine nephrotoxicity biomarkers in a rat model of acute kidney injury induced by cisplatin. Methods A single ip injection with 6 mg·kg^-1 cisplatin in Wistar rats was used to construct an acute kidney injury (AKI) model. During the quarantine period and on the 3rd and 6th day following the administration, animal urine and blood samples in control and cisplatin group were collected five hours after administration. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), urea nitrogen (BUN), creatinine (CRE) in serum and urinary glucoside enzyme (NAG), total urinary protein (uTP), urinary creatinine (uCr) in urine samples were determined by Hitachi 7180 automatic biochemical analyzer. Clumping protein (CLU), glutathione S-transferase -α (GST- α), interferon inducible protein-10 (IP-10), kidney injury factor-1 (KIM-1), osteopontin (OPN), tissue metalloproteinase-1 (TIMP-1), vascular endothelial growth factor-A (VEGF-A), acid glycoprotein (AGP), albumin (Alb), β₂ microglobulin (β₂M), cysteine protease inhibitor C (CysC), epidermal growth factor (EGF), and neutrophil gelatinase-associated lipid carrier protein (NGAL) were determined by Luminex instrument. Combined with animal kidney histopathological examination, a receiver operating characteristic curve (ROC) were establised for sensitivity and specificity analysis of nephrotoxicity markers. Results Histopathological results confirmed that all animal models induced by cisplatin had typical acute kidney injury: Degeneration/ necrosis of renal tubule epithelial cells and myeloid protein tubule type. Compared with control group, the traditional markersserum BUN and CRE did not increase significantly until the 6th day following administration with cisplatin when the animals suffered severe kidney injury (P<0.05), while IP-10, KIM-1, Alb, β2M and CLU biomarkers were found to be significant increased on the third day of administration (P<0.05), and continues to increase. The degree of increase was significantly higher than the traditional biomarkes. ROC results showed that the area under the curve (AUC) of IP-10, CysC, KIM-1 and Alb were significantly better than BUN and CRE, suggesting these biomarkers had more high sensitivity and specificity. Conclusion Diagnostic performance of urine IP-10, CysC, KIM-1 and Alb were better than the traditional biomarkers BUN and CRE. It is suggested that they can be used as candidate biomarkers for the early diagnosis of acute nephrotoxicity caused by drugs

R965

國家自然科學基金項目(81603210);國家“重大新藥創(chuàng)制”科技重大專項(2018ZX09201017-001)