雷帕霉素影响肾缺血再灌注后组织损伤和修复的实验研究

目的研究雷帕霉素对肾缺血再灌注（IR）后不同时期肾小管上皮细胞凋亡及增殖、修复的影响。方法体质量为25～30 g BALB/c小鼠45只随机分为手术对照组、IR对照组和雷帕霉素组。建立阻断小鼠左侧肾蒂30 min后切除右侧肾模型。雷帕霉素组术前及术后每日给予0.5 mL雷帕霉素和生理盐水混悬液（1 mg∶10 mL）灌胃,IR对照组给予等量生理盐水灌胃。术后1、3和7 d每组各处死5只小鼠取全血及肾组织,检测血清肌酐水平,HE染色评价组织病理学改变。采用脱氧核糖核苷酸末端转移酶介导的缺口末端标记法检测肾小管上皮细胞凋亡,通过免疫组织化学检测肾小管上皮细胞增殖细胞核抗原以比较组间肾小管上皮细胞增殖修复差异。结果雷帕霉素组术后1 d肾功能好于IR对照组（P〈0.05）,但在7 d时却较IR对照组差（P〈0.05）。IR对照组和雷帕霉素组术后3个时期的肾组织病理学改变均较手术对照组严重（P〈0.05）,但前两组间损伤病理学评分均无明显差异（P〉0.05）。术后1 d和3 d时,雷帕霉素组凋亡细胞数明显少于IR对照组（P〈0.05）。术后1、3和7 d雷帕霉素组肾小管上皮细胞增殖细胞核抗原表达水平均显著低于IR对照组（P〈0.05）。结论肾缺血再灌注损伤早期以肾小管上皮细胞凋亡为主,后以细胞增殖修复为主。雷帕霉素因具有抗凋亡和抗增殖的双重作用,可以减轻缺血再灌注对肾功能损伤,但却不利于以后的组织修复。

The impact of rapamycin on renal ischemia reperfusion injury and repair in mice

Objective To investigate the effects of rapamycin on renal tubular epithelial cells during ischemia-reperfusion（IR） injury.Methods Forty-five male BALB/c mice（weighing 25-30 g） were divided into 3 groups： sham,IR,and rapamycin,15 in each.IR model was made by 30 minutes of left renal ischemia with non-traumatic vascular clamps and right nephrectomy after the clamp was released.Mice in the rapamycin group were intragastrically administered with 0.5 mL suspension of rapamycin and saline（1 mg ∶ 1 mL） and mice in the IR group with equivalent volume of saline.Five mice were sacrificed on each time point of day 1,3,and 7 after intervention to collect blood and renal tissue samples.Serum creatinine was detected.The tissue injuries were examined by HE staining assay.Apoptosis of tubular epithelium was observed by TUNEL assay.Tubular epithelial proliferation was evaluated by assessing the expression of PCNA with immunohistochemistry.Results Serum creatinine was lower in the rapamycin group on day 1 after reperfusion [（83.89±9.53）μmol/L,P0.05],but higher on day 7 [（19.44±1.15）μmol/L,P0.05],as compared to the IR group.The IR and rapamycin groups showed more severity of tissue injuries than the sham group did（both P0.05）,but no significant difference was seen between the 2 groups（P0.05）.Less TUNEL-positive cells were observed in the rapamycin group on day 1 and 3 after reperfusion（P0.05）.The expression of PCNA in the rapamycin group was significantly decreased at all time points（P0.05）.Conclusions The apoptosis of renal tubular epithelial cells took a major part at the early stage in IR injury.However,it was replaced by cell proliferation and repair in the late period.Rapamycin attenuated renal IR injury at the early stage but delayed tubular cells repair later owing to its anti-apoptosis and anti-proliferation properties.