ObjectiveTo study the pathway of cis-dichlorodiamineplatinum (DDP) inhibiting the synthesis of steroid hormones in mice, and to observe the intervention effect of dehydroepiandrosterone (DHEA). MethodsSixty adult ICR mice were randomly divided into three groups: control group, DDP modeling group, and DHEA group, with 10 male and 10 female mice in each group. The DDP modeling group mice were intraperitoneally injected with DDP solution at a dose of 2.5 mg·kg-1·d-1, once every 3 days, a total of 7 times. On the same day of modeling, the control group mice were injected with an equal amount of physiological saline intraperitoneally. The DEHA treatment group mice were treated with DDP and given a dose of 8.3 mg·kg-1·d -1 of DHEA by gavage for 21 consecutive days. The changes of fatigue indexes of mice were observed by open field, grip and rod rotation tests. The morphology changes of adrenal gland, testicular and ovarian tissue were observed by pathological section and HE staining. The levels of serum steroid hormones were detected by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The mRNA and protein expression levels of the related genes of the hypothalamus, hypophysis, adrenal, testis and ovary were tested by real-time fluorescent quantitative PCR (RT-qPCR) and Western blotting. ResultsCompared with control group, both male and female mice in DDP modeling group were significantly losing weight (P＜0.05), their abilities in horizontal movement and vertical movement decreased (all P＜0.05), and the stay time and grip also significantly decreased (all P＜0.05) in female mice. Indexes of fatigue were improved after DHEA supplement (all P＜0.05). In the DDP modeling group, the arrangement of spermatogenic cells at all levels in the testicular tissue was disordered and the testicular interstitial edema was observed, and a large number of primordial follicles in the ovarian tissue were activated, the number of atresia follicles increased, and the number of granulosa cells in the follicles decreased; while in the DHEA group, the damaged phenotype of testicles and ovaries was significantly improved. Compared with control group, the levels of serum testosterone and dihydrotestosterone in both male and female DDP modeling mice significantly decreased (P＜0.01), the pregnenolone was down-regulated but corticosterone was up-regulated significantly (P＜0.05) in male mice, the corticosterone was down-regulated significantly (P＜0.05) in female mice. Compared with the DDP group, after DHEA supplement, the pregnenolone in male mice and the progesterone in female mice increased significantly (P＜0.05), but the pregnenolone in female mice and the progesterone in male mice decreased significantly (P＜0.05). Compared with control group, the expression levels of Cyp21a1 and Cyp11a1 genes in the adrenal gland and Gnrh gene in the hypothalamus of male and female mice in the DDP modeling group significantly decreased (all P＜0.05); the expression levels of Hsd3b2 gene in the adrenal gland, Star, Cyp11a1, and Lhr genes in the ovaries, Crh, Pomc, and Lhb genes in the hypothalamus, pituitary, and pituitary of female mice significantly decreased (all P＜0.05); the expression levels of Star gene and StAR protein in the testicles of male mice, as well as Fshb and Lhb genes in the pituitary gland, were significantly down-regulated (all P＜0.05). After DHEA supplement, compared with the DDP modeling group, the mRNA expression levels of Cyp17a1 in the adrenal gland of male mice and Cyp17a1, Lhr and Fshr genes in testis were down-regulated significantly (P＜0.05); the expression level of Cyp11a1 gene in the adrenal gland of female mice was also decreased (P＜0.05); while the expression levels of Hsd3b2 gene in the adrenal gland, Star, Cyp11a1, Hsd3b2 and Lhr gene in the ovary, and Lhb gene in the pituitary gland were all up-regulated ( P＜0.05). ConclusionThe function of hypothalamus-pituitary-adrenal/gonadal axis was inhibited by DDP intermittent injection, especially in female. Supplementation of DHEA can help regulate the homeostasis of steroid hormone levels.

目的 研究顺铂诱发药源性虚证性小鼠类固醇激素合成功能的改变.方法 72只雄性ICR小鼠分为对照组32只和顺铂组40只,顺铂组予10mg/kg顺铂腹腔注射,每周1次,共5次(d1、d8、d15、d22、d29).每次造模后1周各组取8只检测相应指标,以ELISA检测睾酮和皮质酮含量、RT-qPCR检测下丘脑、垂体、肾上腺和睾丸类固醇激素合成酶相关基因表达.Western Blot 检测 肾上腺 StAR、CYP11A1、CYP21A2、CYP11B1;睾丸 StAR、CYP11A1、CYP17A1、HSD3B2 蛋白表达.结果 ①顺铂给药4次(d28)后显著下降小鼠体重,胸腺与肾脏萎缩,脾肿大.②小鼠血清皮质酮在顺铂给药2次(d14)及4次(d28)显著上升,末次(d30)显著下降.血清睾酮含量在给药1次后(d7)明显上升,给药4次和末次(d28、d30)明显下降.③随顺铂累积肾上腺皮质束状带明亮细胞增多.④随顺铂累积,曲细精管改变、生精细胞紊乱加重,给药4次(d28)后曲细精管内改变显著,支持细胞减少,间质增厚,间质细胞水肿明显.⑤顺铂组肾上腺在给药1次(d7)后Cyp21a1基因表达和CYP11A1蛋白表达上调,但Cyp11b2基因表达下调;给药2次(d14)后Star、Cyp11a1、Cyp11b1、Cyp11b2和Cyp21a1基因表达均下调;给药3次(d21)Cyp11a1和Cyp21a1基因表达下调;给药4次(d28)Cyp11b1基因表达上调.⑥顺铂组睾丸在给药2次(d14)后Star、Cyp11a1、Cyp17a1、Hsd17b3和Ar基因表达均下调;给药3次(d21)Hsd3b2和Ar基因表达上调;给药4次(d28)Star、Cyp11a1、Cyp17a1、Hsd3b2、Hsd17b3和Ar基因表达均显著上调,StAR蛋白表达上调;末次给药(d30)Star、Cyp11a1、Cyp17a1、Hsd3b2、Hsd17b3和Ar基因表达均上调,StAR、CYP11A1 和 CYP17A1蛋白表达显著增强.⑦顺铂组下丘脑在给药2次(d14)GnRH基因表达增强,给药1次、4次和末次(d7、d28、d30)CRH基因表达均增强.⑧顺铂组垂体在给药1次(d7)Pou1f1基因表达上调而Fshb和Lhb基因表达均下调;给药4次(d28)Pomc基因表达上调.结论 顺铂可损害类固醇激素合成的肾上腺皮质轴与睾丸轴,尤其对睾丸各细胞的累积性损害可能是诱发肾精不足证的主要机制.

目的:研究顺铂短期给药诱发小鼠药源性证候的属性及对类固醇激素合成功能的影响.方法:40只雄性ICR小鼠随机分为对照组及顺铂不同剂量(3、5、10 mg·kg-1·d-1)组,每组10只.各药物干预组小鼠腹腔注射相应剂量的顺铂,连续5 d.分别于干预5 d后和停药5 d后,观察小鼠体征(体质量、摄食量、腋温、爪色泽),并进行行为学测试(抓力、旷场试验和转棒试验).观察期后,取血清;分离肾脏、心脏、胸腺和脾脏,计算脏器指数;收集肾上腺和睾丸.ELISA检测血清皮质酮和睾酮含量;HE染色后光镜下观察肾上腺与睾丸组织形态学变化;PCR、Western blot分别检测肾上腺皮质类固醇激素合成酶和睾丸睾酮合成酶相关的mRNA、蛋白表达.结果:①干预5 d后,顺铂10 mg·kg-1·d-1组小鼠体质量、摄食量、腋温、水平运动及垂直运动较对照组均显著降低(P＜0.05,P＜0.01);顺铂3、5mg·kg-1·d-1组小鼠腋温明显低于对照组,而爪色r值高于对照组(P＜0.05,P＜0.01).停药5 d后,顺铂10 mg·kg-1·d-1组小鼠体质量、摄食量、在棒时间、抓力、腋温、爪色r值、水平运动及垂直运动较对照组均显著降低(P＜0.05,P＜0.01);顺铂3、5 mg·kg-1·d-1组小鼠垂直运动较对照组亦明显减少(P＜0.01).②与对照组相比,10 mg·kg-1·d-1顺铂作用显著抑制小鼠脾脏、胸腺、心脏,脏器指数明显降低(P＜0.05,P＜0.01),而肾脏指数显著升高(P＜0.01);5 mg·kg-1·d-1顺铂作用亦抑制心脏(P＜0.05).③形态学观察显示,顺铂10mg·kg-1·d-1组小鼠肾上腺皮质球状带细胞肿胀明显,顺铂5、10mg·kg-1·d-1组小鼠可见各级生精细胞排列略紊乱,睾丸间质细胞收缩明显.④与对照组相比,顺铂10 mg·kg-1·d-1组小鼠血清皮质酮含量显著升高(P＜0.01),血清睾酮含量无明显变化.⑤与对照组相比,顺铂3 mg·kg-1·d-1组小鼠肾上腺Star、Cyp21a1的mRNA表达受到抑制(P＜0.05),而顺铂10mg·kg-1·d-1组小鼠肾上腺Star、Cyp11b1、Cyp21a1的mRNA表达及StAR、CYP21A2、CYP11B1蛋白表达显著上调(P＜0.05,P＜0.01).⑥与对照组相比,10 mg·kg-1·d-1顺铂作用显著抑制睾丸 StAR、CYP1 1A1、CYP17A1、HSD3B2的 mRNA 与蛋白表达(P＜0.05,P＜0.01);5 mg·kg-1·d-1 顺铂干预显著抑制 Star 和Hsd3b2的 mRNA 表达(P＜0.05,P＜0.01);3 mg·kg-1·d-1 顺铂作用亦显著抑制 Cyp17a1 和Hsd3b2的mRNA表达(P＜0.01).结论:顺铂短期给药可诱发小鼠虚证表现,以精气亏虚为主,其物质基础与类固醇激素合成及储备下降有关.

睾酮的合成与男子生殖功能及衰老相关,睾丸间质细胞是合成与分泌睾酮的主要场所,多种酶与转录因子参与睾酮合成过程.温补肾阳中药及其主要活性成分,包括淫羊藿、菟丝子和肉苁蓉等被证明具有调节生殖内分泌水平,改善氧化应激、抑制炎症发生、调节睾酮合成相关酶mRNA及蛋白表达,影响睾酮合成与分泌.结合文献就影响睾丸间质细胞睾酮合成与分泌的主要因素,及温补肾阳中药对睾酮合成与分泌的调节研究进行综述.

目的 研究顺铂不同给药方案对小鼠类固醇激素合成及储备功能的影响.方法 40只雌性ICR小鼠分为4组,顺铂连续和间隔造模组以及两个对照组.顺铂连续造模组给药7 d,每天注射3 mg/kg顺铂;间隔造模组每周注射1次10 mg/kg顺铂,共4次.末次造模前进行旷场实验、转棒疲劳实验、抓力测试;次日处死前称体重,摘取小鼠心脏、脾、胸腺和双侧肾并称重;RT-qPCR检测下丘脑GnRH和CRH、垂体Pomc、Fshb、Lhb、Pou1f1基因表达,以及肾上腺和卵巢类固醇合成酶(StAR、Cyp11a1、Cyp21a1、Cyp17a1、Cyp11b1、Hsd3b2、Cyp19a1、Hsd17b1)及受体(Esr1、Esr2和Gper1)基因表达;Western Blot检测肾上腺和卵巢StAR、CYP11A1、CYP21A2和CYP11B1蛋白表达.结果 与对照组比较:(1)不同频次顺铂造模均可抑制小鼠体重增长,顺铂累积后可以诱导小鼠气虚证表现,并抑制脏器质量.(2)不同频次顺铂造模均能加速激活原始卵泡分化,产生大量闭锁卵泡,诱导膜细胞及颗粒细胞凋亡,健康的成熟卵泡减少;但对肾上腺组织和细胞形态影响不大.(3)连续造模后肾上腺StAR蛋白表达下调;间隔造模后StAR升高,但下调CYP11B1蛋白表达.(4)连续造模后,卵巢Cyp11a1、Hsd3b2、Esr1和Gper1基因表达上调,Star和Cyp17a1基因表达下调;间隔造模后StAR蛋白表达和Hsd3b2基因表达下调.(5)顺铂连续造模后下丘脑GnRH和CRH基因表达明显下调.(6)顺铂连续造模后垂体Pomc、Fshb、Pou1f1基因表达升高,而间隔造模组Lhb表达下降.结论 顺铂能够诱发小鼠气虚、精气不足证的相关表现;合成类固醇激素的卵巢受到选择性损伤大于肾上腺,短期连续给药也进一步抑制HPA轴和HPG轴的高级中枢下丘脑和垂体功能.