基于调控结肠组织中SCF和c-kit mRNA表达分析苍术燥性效应量效关系

Analysis on dose-effect relationship of Atractylodes Rhizoma dryness effect based on regulation of SCF and c-kit mRNA expressions in colon tissue

目的:基于结肠组织中干细胞因子(SCF)及其受体c-kit mRNA表达分析苍术燥性效应的量效关系,确定引起小鼠燥性反应的苍术基础剂量,为临床用药提供参考。方法:75只C57BL/6小鼠分为对照组和185、370、555及740 mg·kg^-1苍术挥发油组,每组15只,给药体积为0.015 mL·g^-1,灌胃给药14 d,对照组给予同体积1%吐温-20溶液。测定小鼠日均饮水量和日均尿量,采用免疫组织化学法检测小鼠肾组织中水通道蛋白2(AQP2)蛋白表达水平,计算小鼠脾脏和肾脏系数,采用RT-PCR法检测小鼠结肠组织中SCF和c-kit mRNA表达水平,根据以上指标考察各组小鼠燥性效应。结果:与对照组比较,185和370 mg·kg^-1苍术挥发油组小鼠日均饮水量、日均尿量、肾组织中AQP2蛋白表达水平、肾脏系数和脾脏系数差异均无统计学意义(P>0.05),小鼠结肠组织中SCF mRNA表达水平明显升高(t=4.859,P<0.01;t=6.651,P<0.05),185 mg·kg^-1苍术挥发油组小鼠结肠组织中c-kit mRNA表达水平明显升高(t=2.946,P<0.05),而370 mg·kg^-1苍术挥发油组差异无统计学意义(P>0.05);与对照组比较,555 mg·kg^-1苍术挥发油组小鼠日均尿量、结肠组织中c-kit mRNA表达水平明显降低(t=7.708,P<0.01;t=8.465,P<0.01),其他指标差异无统计学意义(P>0.05);与对照组比较,740 mg·kg^-1苍术挥发油组小鼠饮水量、肾组织中AQP2蛋白表达水平明显升高(t=3.554,P<0.01;t=4.636,P<0.01),日均尿量、脾脏系数、结肠组织中SCF和c-kit mRNA表达水平明显降低(t=7.708,P<0.01;t=4.985,P<0.01;t=7.314,P<0.01;t=28.45,P<0.01)。结论:低剂量(185和370 mg·kg^-1)苍术挥发油对健康小鼠无明显燥性作用,而高剂量(555 mg·kg^-1以上剂量)苍术挥发油可能通过影响SCF/c-kit信号通路对小鼠产生明显的燥性作用。

Objective:To analyze the dose-effect relationship of dryness effect of Atractylodes Rhizoma based on the expressions of stem cell factor(SCF)and its receptor c-kit mRNA in colon tissue and to ensure the basic dose of Atractylodes Rhizoma which caused the dryness reaction in the mice,and to provide the reference for clinical medicine.Methods:Seventy-five C57BL/6 mice were divided into control group,185,370,555,and 740 mg·kg^-1 Atractylodes oil groups,15 mice in each group,and the administration volume was 0.015 mL·g^-1;the mice were administered intragastrically for 14 d,and the mice in control group were given the same volume of 1%Tween-20 solution.The average daily water intake and daily average urine output of the mice were detected.Immunohistochemical method was used to detect the expression levels of aquaporin-2(AQP2)in the kidney tissue of the mice;the sleen and kidney coefficients of the mice were calculated,and RT-PCR method was used to determine the expression levels of SCF and c-kit mRNA in the colon tissue;the dryness effects of the mice in various groups were investigated according the obove indicatros.Results:Compared with control group,the average daily water intakes,daily urine outputs,the AQP2 expression levels in kidney tissue,the kidney coefficients and spleen coefficients of the mice in 185 and 370 mg·kg^-1 Atractylodes oil groups had no significant differences(P>0.05);the expression levels of SCF mRNA in colon tissue of the mice were significantly increased(t=4.859,P<0.01;t=6.651,P<0.05);the expression level of c-kit mRNA in colon tissue of the mice in 185 mg·kg^-1 Atractylodes oil group was significantly increased(t=2.946,P<0.05),but in 370 mg·kg^-1 Atractylodes oil group had no statistically significant difference(P<0.05).Compared with control group,the average daily urine output and the expression level of c-kit mRNA in colon tissue of the mice in 555 mg·kg^-1 Atractylodes oil group were significantly reduced(t=7.708,P<0.01;t=8.465,P<0.01),but there were no statistically significant differences in other indicators(P>0.05).Compared with control group,the water intake and the AQP2 protein expression level in kidney tissue of the mice in 740 mg·kg^-1 Atractylodes oil group were significantly increased(t=3.554,P<0.01;t=4.636,P<0.01),and the average daily urine volume,the spleen coefficient,and the expression levels of SCF and c-kit mRNA in colon tissue of the mice were decreased significantly(t=7.708,P<0.01;t=4.985,P<0.01;t=7.314,P<0.01;t=28.45,P<0.01).Conclusion:Low doses(185 and 370 mg·kg^-1)of Atractylodes oil show no obvious dryness effect in the healthy mice,and the high doses(555 mg·kg^-1 and above)of Atractylodes oil may show obvious dryness effect by influencing the SCF/c-kit signal pathway.