衢枳壳提取物改善2型糖尿病db/db模型小鼠肾脏氧化损伤的作用研究

Quzhike, Extract of Citrus Changshan-Huyou, Ameliorates Renal Oxidative Damage in Type 2 Diabetic db/db Mice

目的观察衢枳壳提取物(QFAE)改善2型糖尿病db/db模型小鼠肾脏氧化损伤的作用及机制。方法 24只7~8周龄雄性C57BLKS/J db/db模型小鼠随机分为模型组,QFAE低、高剂量组,每组8只;同时以8只C57BLKS/J db野生型小鼠为对照组,对照组和模型组小鼠每天以蒸馏水灌胃,QFAE低、高剂量组小鼠每天按QFAE 100、300mg/kg灌胃,连续7周。记录小鼠体质量、血糖、肾质量;检测血清肌酐(Cr)和尿素氮(BUN)水平,肾组织超氧化物歧化酶(SOD)、过氧化氢酶(CAT)和还原型谷胱甘肽(GSH)活性;HE染色观察肾组织病理学形态;RT-PCR法检测各组小鼠肾组织抗氧化基因谷氨酸-半胱氨酸修饰亚基(Gclm)、醌氧化还原酶1(Nqo1)和谷氨酸-半胱氨酸连接酶催化亚基(Gclc)表达情况。结果与模型组比较,QFAE对db/db模型小鼠血糖[(31.30±1.88)mmol/L、(30.78±2.07)mmol/L比(32.78±0.66)mmol/L,P>0.05]、体质量[(44.76±3.55)g、(43.92±4.58)g比(45.26±2.74)g,P>0.05]无影响,但显著减轻小鼠肾质量[(0.54±0.04)g、(0.48±0.08)g比(0.66±0.05)g,P均<0.01],降低血清Cr[(38.16±2.47)μmol/L、(36.17±2.00)μmol/L比(45.76±3.70)μmol/L,P<0.05或P<0.01]和血清BUN [(7.72±1.22)mmol/L、(7.55±0.72)mmol/L比(11.31±0.63)mmol/L,P均<0.01]水平,提高肾组织SOD [(125.74±18.17)U/mgprot、(147.64±16.36)U/mgprot比(107.15±10.83)U/mgprot,P均<0.05]、GSH [(2.34±0.28)μmol/mgprot、(2.40±0.26)μmol/mgprot比(1.93±0.06)μmol/mgprot,P<0.05或P<0.01]和CAT [(38.20±6.53)U/mgprot、(42.50±10.90)U/mgprot比(28.99±1.67)U/mgprot,P<0.05或P<0.01]活性,且显著减轻肾组织病理损伤。RT-PCR结果显示,与模型组比较,QFAE能够促进抗氧化基因Gclm[(1.30±0.15)、(1.46±0.27)比(0.87±0.11),P<0.05或P<0.01]、Nqo1 [(1.23±0.06)、(1.31±0.18)比(0.82±0.16),P<0.05或P<0.01]、Gclc [(1.19±0.08)、(1.28±0.16)比(0.93±0.11),P<0.05或P<0.01]mRNA表达。结论 QFAE能够显著改善2型糖尿病db/db小鼠肾脏氧化损伤,其机制可能与促进抗氧化基因表达,提高肾组织抗氧化能力有关。

Objective To investigate the protective effect of Quzhou Fructus Aurantii extract(QFAE), i.e., extract of citrus changshan-huyou, on renal oxidative damage in type 2 diabetic db/db mice and its mechanism. Methods Twenty-four male C57 BLKS/J db/db model mice of 7 to 8 weeks old were randomly divided into model group,QFAE low and high dose group, with 8 mice in each group. Eight C57 BLKS/J db wild-type mice were as the control group. Mice in the control group and model group were fed with distilled water every day. Animals in QFAE low and high dose groups were administrated with 100 and 300 mg/kg QFAE every day for 7 weeks. The body mass, blood sugar and kidney mass were recorded. Levels of serum creatinine(Cr) and urea nitrogen(BUN)were measured, and the activities of superoxide dismutase(SOD), catalase(CAT) and reduced glutathione(GSH) in renal tissue were detected. HE staining was used to observe renal histopathology. RT-PCR was performed to detect the expression of glutamate-cysteine modified subunit(Gclm), quinone oxidoreductase 1(Nqo1) and glutamate-cysteine ligase catalyzed subunit(Gclc) in kidney tissues. Results Compared to model group, QFAE had no effect on blood sugar [(31.30 ±1.88) and(30.78 ±2.07) vs(32.78 ±0.66)mmol/L] and body mass [(44.76 ±3.55) and(43.92 ±4.58) vs(45.26±2.74)g], but reduced levels of renal body index [(0.54±0.04) and(0.48±0.08) vs(0.66±0.05)g,P<0.01], serum Cr[(38.16±2.47) and(36.17±2.00) vs(45.76±3.70)μmol/L, P<0.05 or P<0.01], serum BUN[(7.72±1.22) and(7.55±0.72) vs(11.31±0.63)mmol/L, P<0.01], and increased the activity of SOD [(125.74±18.17) and(147.64±16.36) vs(107.15±10.83)U/mgprot, P<0.05], GSH[(2.34±0.28) and(2.40±0.26) vs(1.93±0.06)μmol/mgprot, P<0.05 or P<0.01] and CAT[(38.20±6.53) and(42.50±10.90) vs(28.99±1.67)U/mgprot, P<0.05 or P<0.01] in kidney tissues. QFAE also relieved the pathological damage of renal tissues. Compared to model group, the results of RT-PCR showed that QFAE significantly up-regulated the mRNA expression of Gclm [(1.30±0.15) and(1.46±0.27) vs(0.87±0.11), P<0.05 or P<0.01], Nqo1[(1.23±0.06) and(1.31±0.18) vs(0.82±0.16), P<0.05 or P<0.01),and Gclc[(1.19±0.08) and(1.28±0.16) vs(0.93±0.11), P<0.05 or P<0.01]. Conclusion QFAE could ameliorates the renal oxidative damage in type 2 diabetic db/db mice significantly, and its mechanism is mainly related to the promotion of antioxidant gene expression, thereby improving the antioxidant capacity of renal tissues.