补肾健脾法对氢化可的松诱发的脾肾虚证小鼠肾上腺功能的影响

目的研究补肾健脾法对氢化可的松诱发的脾肾虚证小鼠肾上腺功能的影响。方法将60只ICR小鼠随机分为正常对照组、氢化可的松组、四君子汤组12.75 g(生药)/kg]、六味地黄汤组12.5 g(生药)/kg]、桂附地黄汤组13.5 g(生药)/kg],连续灌胃给药氢化可的松(1.65 mg/kg)14 d建立药源性证候模型;各中药复方组在灌胃氢化可的松的同时及停止氢化可的松灌胃后7 d,每日分别灌胃给药3个中药复方。每日称定小鼠体质量;在第14、21天,运用小鼠辨证论治实验方法学观测小鼠四诊指标;在第15、22天,计算脾脏、胸腺指数,ELISA检测血清皮质酮、促肾上腺皮质激素、肾上腺素和去甲肾上腺素水平,采用实时荧光定量PCR技术检测Star、Cyp11a1、Cyp21a1、Cyp11b1、Cyp11b2、Dbh、Ddc、Pnmt、Th mRNA表达,Western blot检测LDLR、SRBI、Star蛋白表达。结果氢化可的松干预后,小鼠体质量,抓力,尾部最低温度,脾脏、胸腺指数,血清皮质酮、促肾上腺皮质激素水平显著下降(P<0.05,P<0.01);显著抑制相关肾上腺类固醇激素合成酶(Star、Cyp11a1、Cyp21a1、Cyp11b1)mRNA表达(P<0.05,P<0.01),并抑制胆固醇转化为肾上腺类固醇激素过程中相关蛋白(LDLR、SRBI、StAR)表达。与氢化可的松组比较,桂附地黄汤组显著增加小鼠抓力(P<0.01),四君子汤组、六味地黄汤组和桂附地黄汤组显著升高血清皮质酮均水平(P<0.01),四君子汤组、桂附地黄汤组明显抑制Star、Cyp11a1、Cyp21a1、Cyp11b1 mRNA表达(P<0.05,P<0.01),3个复方组能改善LDLR、SRBI、StAR蛋白下降。结论桂附地黄汤对小鼠阳虚证候有所改善,3个中药复方对小鼠阴虚证候疗效不显著。3个中药复方对氢化可的松诱发的肾上腺皮质功能抑制均有缓解作用。

Effects of method of reinforcing kidney and invigorating spleen on adrenal function of hydrocortisone-induced spleen-kidney deficiency mice

AIM To study the effects of method of reinforcing kidney and invigorating spleen on adrenal func?tion of hydrocortisone?induced spleen?kidney deficiency mice.METHODS Sixty ICR mice were randomly divided into normal control group,hydrocortisone group,Sijunzi Decoction group12.75 g(raw drug)/kg],Liuwei Dihuang Decoction group12.5 g(raw drug)/kg],Guifu Dihuang Decoction group13.5 g(raw drug)/kg].Simultaneously,consective administration of hydrocortisone(1.65 mg/kg)was practiced to establish drug?in?duced syndrome models.Meanwhile,in the groups for Chinese herbal compounds intervention,the corresponding decoction was dosed by gastric gavage.After hydrocortisone gavage cessation,the administration of Chinese herbal compounds went on for another seven days.The body weight of the mice was weighed daily.On the 14th and 21st days,TCM four diagnostic indicators of mice were observed by using the experimental method of syndrome differen?tiation and treatment of mice.On the 15th and 22nd days,the spleen and thymus indexes were calculated,the levels of serum corticosterone,adrenocorticotropic hormone,adrenaline and noradrenaline were measured by ELISA,and the expressions of Star,Cyp11a1,Cyp21a1,Cyp11b1,Cyp11b2,Dbh,Ddc,Pnmt and Th mRNA were detected by real?time fluorescence quantitative PCR,and the expressions of LDLR,SRB1 and Star protein were detected by Western blot.RESULTS After mice were treated with hydrocortisone,the body weight,grasping power,tail temperature,spleen,level of thymus index,serum corticosterone and adrenocorticotropic hormone levels decreased significantly(P<0.05,P<0.01).The mRNA expression of related adrenal steroid hormone synthase(Star,Cyp11a1,Cyp21a1,Cyp11b1)(P<0.05,P<0.01),and expression of related proteins(LDLR,SRBI,STAR)during the conversion of adrenal cholesterol into hormones were inhibited.Compared with the hydrocortisone group,Guifu Dihuang Decoction group was observed with significantly increased grip of mice(P<0.01),Sijunzi Decoction group,Liuwei Dihuang Decoction group and Guifu Dihuang Decoction group shared significantly increased serum corticosterone level(P<0.01),Sijunzi Decoction group and Guifu Dihuang Decoction groups dis?played significantly inhibited expression of Star,Cyp11a1,Cyp21a1 and Cyp11b1 mRNA(P<0.05,P<0.01),and the three groups for Chinese herbal compounds intervention demonstrated improvement in LDLR,SRBI and StAR protein expression.CONCLUSION To mice models,Guifu Dihuang Decoction improves the syndrome of yang deficiency,but none of the three traditional Chinese medicine compounds displays significant efficacy on yin deficiency syndrome,and all the three traditional Chinese medicine compounds alleviate cortisol?induced adrenal function inhibition.