益肾填精中药对环境内分泌干扰物染毒大鼠睾酮合成关键酶及雄激素受体表达的影响

Effect of Chinese Herbs for Shen Invigorating Essence Replenishing on Expression of Restrict Enzymes of Testosterone Biosynthesis and Androgen Receptor of Rats Contaminated with Environmental Endocrine Disruptors

目的观察益肾填精中药对邻苯二甲酸二(2-乙基己基)酯(di-2-ethylhexylphthalate,DEHP)及氯氰菊酯(cypermethrin,CYP)染毒大鼠睾酮合成关键酶和雄激素受体(androgen receptor,AR)基因及蛋白表达水平的影响,探讨所用中药拮抗环境内分泌干扰物(environmental endocrine disruptors,EEDs)抗雄激素活性的可能作用机制。方法 70只21日龄雄性SD大鼠随机分为7组,每组10只。对照组(喂饲玉米油)、染毒A组(DEHP 500 mg/kg)、染毒B组(CYP 80 mg/kg)、染毒C组(DEHP 500 mg/kg+CYP 80 mg/kg)及治疗A组、治疗B组、治疗C组,治疗A、B、C组同时喂饲益肾填精中药合剂(40 m L/kg)。每日空腹灌胃给药,疗程为30天。采用实时荧光定量聚合酶链式反应(real time polymerase chain reaction,real time-PCR)及免疫印迹方法(Western blot),分别检测St AR、CYP11A1、CYP17A1、3β-HSD、17β-HSD、AR mRNA及蛋白表达水平。结果与对照组比较,染毒A、B、C组St AR、CYP11A1、CYP17A1、3β-HSD、17β-HSD、AR mRNA表达水平显著下调(P<0.05,P<0.01)。中药治疗干预可使上述基因表达水平明显上调(P<0.05,P<0.01)。DEHP、CYP单独及联合染毒可导致CYP11A1、CYP17A1、AR蛋白表达水平显著下调(P<0.05,P<0.01),而中药治疗干预可使上述蛋白表达水平显著上调(P<0.05,P<0.01)。结论益肾填精中药可通过使睾丸的AR以及睾酮合成关键酶的表达上调,促进染毒大鼠自身睾酮的生物合成,多水平、多靶点地发挥其对EEDs抗雄激素活性及生殖毒性的拮抗作用。

Objective To observe the effect of Chinese herbs for Shen invigorating essence replenishing （CHSIER） on restrict enzymes of testosterone biosynthesis and androgen receptor （AR） expression in testes of rats exposed to di-（2-ethylhexyl） phthalate （DEHP） andcypermethrin （CYP）, and to study its possible mechanism for antagonizing antiandrogen activity of environmental endocrine disruptors （EEDs）. Methods Seventy male Sprague-Dawley rats （21 days old） were randomly divided into seven groups （10 in each group）, i.e., the control group （fed with corn oil）, exposed group A （DEHP 500 mg/kg）, exposed group B （CYP 80 mg/kg）, exposed group C （DEHP 500 mg/kg ＋CYP 80 mg/kg）, and treatment group A, B, C （CHSIER 40 mL/kg）, respectively. All medication was administered by gastrogavage, once per day, 30 days as one therapeutic course, mRNA and protein expressions of acute regulatory protein （STAR）, CYP11A1, CYP17A1, 3β-hydroxysteroid dehydrogenase （3β-HSD）, 17β-HSD, androgen receptor （AR） in rat testes were detected by real time polymerase chain reaction （real time RT-PCR） and Western blot respectively. Results Compared with the control group, mRNA expres- sion levels of STAR, CYP11A1, CYP17A1,3β-HSD, 17β-HSD, and AR in rat testes were down-regulated in exposed group A, B, and C （P 〈0.05, P 〈0.01）. They could be up-regulated by CHSIER （P 〈0.05, P 〈0.01 ）. Protein expressions of CYP11A1, CYP17A1, and AR could be significantly down-regulated by DEHP, CYP, or their combination （P 〈 0.05, P 〈 0.01 ）, which could be significantly up-regulated by CHSIER （P 〈0.05, P 〈0.01 ）. Conclusions CHSIER could promote testosterone biosynthesis of expo- sure rats by up-regulating the expression of restrict enzymes of testosterone biosynthesis and AR, and significantly antagonize reproductive toxicity of EEDs at multiple levels and targets.