去卵巢和雌激素预防性给药对大鼠甲状旁腺激素基因的表达调控研究

发布时间：2018-07-26 11:38

【摘要】：甲状旁腺是机体内分泌腺之一,其主要功能是分泌甲状旁腺激素(parathyroid hormone,PTH)。雌激素(estrogen)是由卵巢分泌的一种类固醇激素,雌激素的降低会打破钙磷平衡而导致骨质疏松,而PTH是人体骨转换率的主要决定因素,表明雌激素与PTH之间可能存在某种联系。然而,迄今为止尚缺乏雌激素调控PTH分泌的直接证据,雌激素调控PTH分泌的机理尚不清楚。本研究采用双侧卵巢切除术建立绝经后骨质疏松(postmenopausal osteoporosis,PMOP)大鼠模型,利用石蜡组织切片和HE染色对大鼠的股骨、胫骨、子宫、十二指肠、肾皮质和肾髓质进行组织形态学观察,利用酶联免疫吸附测定法(enzyme-linked immunosorbent assay,ELISA)检测血清中雌激素的变化,用实时定量PCR(real-time quantitative polymerase chain reaction,Real-time qPCR)技术检测SD大鼠甲状旁腺中PTH、雌激素受体(estrogen receptor,ER)表达变化。进一步对调控PTH表达的潜在通路中的关键基因,如钙敏感受体(calcium sensing receptor,CaSR)、维生素D受体(vitamin D receptor,VDR)、成纤维细胞生长因子-23(fibroblast growth factor-23,FGF-23)、FGF-23的受体1(fibroblast growth factor receptor 1,FGFR1)、FGF-23的辅助受体Klotho和早期生长反应基因1(early growth response gene 1,Egr1)的表达量进行分析,探讨去卵巢和预防性给药对PTH的调控机制。脏器和骨组织形态学观察结果表明,去卵巢(ovariectomized,OVX)组大鼠的股骨和胫骨均出现骨小梁断裂、间距变大、结构紊乱等骨质疏松症状,而去卵巢后预防性注射β-雌二醇(β-estradiol-treated OVX,OVX/E2)组并未出现明显的病理症状。相较于假手术(Sham-operated,Sham)组,OVX组和OVX/E2组的十二指肠、肾皮质和肾髓质区都未出现明显病变。相较于Sham组,OVX组大鼠子宫内膜固有层中的子宫腺数目增多、腺腔增大、子宫黏膜上皮明显增厚。OVX/E2组的子宫腺腺腔虽略有增大,但腺体形态较为规则,其子宫内膜也未发生明显的增厚现象。说明注射小剂量的β-雌二醇可以预防骨质疏松症的发生,且不会有子宫癌变的现象发生。ELISA检测结果表明三种不同处理组间大鼠的血清雌激素没有显著性的差异。Real-time qPCR检测结果表明,去卵巢后大鼠PTH的表达量显著下调,而预防性注射β-雌二醇后PTH的表达量显著上调。ER-α存在于甲状旁腺中,且去卵巢后ER-α的表达量上调,而预防性注射β-雌二醇后ER-α的表达量显著下调,说明甲状旁腺中ER-α能够响应雌激素的微弱变化,参与PTH的表达调控。去卵巢后大鼠甲状旁腺中VDR的表达量上调,而预防性注射β-雌二醇后大鼠甲状旁腺中VDR的表达量下调,表明VDR可能是抑制PTH的表达。去卵巢后血清中Ca~(2+)略有升高,甲状旁腺中CaSR的表达量显著下调,预防性注射β-雌二醇后Ca~(2+)略有升高,甲状旁腺中CaSR的表达量显著上调,表明Ca~(2+)/CaSR调控系统可能参与PTH的表达调控。去卵巢后大鼠甲状旁腺中FGF-23、FGFR1、Klotho及Egr1表达量显著下调,而预防性注射β-雌二醇后FGF-23、Klotho和Egr1的表达量下调,FGFR1的表达量显著上调,说明去卵巢后甲状旁腺中FGF-23/Klotho系统对PTH表达的抑制作用降低了,但是预防性注射β-雌二醇后这个受体系统的功能更低。这些结果表明,去卵巢和预防性给药可能会影响甲状旁腺中ER-α、VDR信号调控系统、Ca~(2+)/CaSR调控系统以及FGF-23/Klotho受体调控系统,进而调控PTH的表达。
[Abstract]:Parathyroid gland is one of the endocrine glands of the body, its main function is the secretion of parathyroid hormone (PTH). Estrogen (estrogen) is a steroid hormone secreted by the ovary. The decrease of estrogen will break the balance of calcium and phosphorus and cause osteoporosis, and PTH is the main determinant of bone turnover rate in human body, indicating that estrogen and P are the main factor. There may be some connection between TH. However, so far, there is no direct evidence that estrogen regulates the secretion of PTH. The mechanism of estrogen regulation of PTH secretion is not clear. This study uses bilateral oophorectomy to establish postmenopausal osteoporosis (postmenopausal osteoporosis, PMOP) rats model, using paraffin tissue section and HE staining to large The histomorphology of the femur, tibia, uterus, duodenum, kidney cortex and renal medulla were observed and the changes of estrogen in serum were detected by enzyme-linked immunosorbent assay (ELISA), and the real-time quantitative PCR (real-time quantitative polymerase chain reaction, Real-time qPCR) was used to detect the size of the rat. The changes in the expression of PTH and estrogen receptor (ER) in the parathyroid gland of the rat are the key genes in the potential pathways regulating the expression of PTH, such as the calcium sensitive receptor (calcium sensing receptor, CaSR), the vitamin D receptor (vitamin D), and fibroblast growth factor. The expression of the receptor 1 (fibroblast growth factor receptor 1, FGFR1), FGF-23 auxiliary receptor Klotho and the early growth response gene 1 (early growth response gene 1, Egr1) was analyzed to explore the regulation mechanism of the ovariectomized and preventive administration. In the femur and tibia of the rats, the fracture of the trabecular bone, the spacing and the disorder of the structure were observed. The prophylactic injection of beta estradiol (beta -estradiol-treated OVX, OVX/E2) after the ovariectomy had no obvious pathological symptoms. Compared to the sham operation (Sham-operated, Sham) group, the duodenum, the renal cortex and the renal cortex of group OVX and OVX/E2 group. There were no obvious lesions in the medullary region. Compared to group Sham, the number of uterine glands in the lamina propria of group OVX increased, the gland cavity increased, and the epithelium of the uterus was thickened in.OVX/E2 group, although the adeno gland of the uterus increased slightly, but the shape of the gland was more regular, and the endometrium had no obvious thickening. The amount of beta estradiol can prevent the occurrence of osteoporosis, and there is no.ELISA detection of the phenomenon of uterine carcinogenesis. The results of serum estrogen in the three different treatment groups showed no significant difference in serum estrogen. The results of.Real-time qPCR detection showed that the amount of PTH in the ovariectomized rats was significantly down, while the prophylactic injection of beta female was a prophylactic injection. After alcohol, the expression of PTH was significantly up-regulated in parathyroid glands, and the expression of ER- alpha was up-regulated after ovariectomized, while the expression of ER- alpha was significantly downregulated after the prophylactic injection of beta estradiol. It indicated that ER- alpha in parathyroid glands could respond to the weak changes in estrogen and participate in the expression and regulation of PTH. The expression of VDR in parathyroid glands of ovariectomized rats The amount of VDR was down regulated in the parathyroid parathyroid after prophylactic injection of beta estradiol, indicating that VDR may be the expression of inhibition of PTH. The serum Ca~ (2+) increased slightly after ovariectomized, and the expression of CaSR in parathyroid glands decreased significantly. The Ca~ (2+) increased slightly after the prophylactic injection of beta estradiol, and the expression of CaSR in parathyroid glands was significantly higher. The Ca~ (2+) /CaSR regulatory system may be involved in the regulation of PTH expression. The expression of FGF-23, FGFR1, Klotho and Egr1 in the parathyroid glands of the ovariectomized rats is significantly down, while the prophylactic injection of beta estradiol is down to FGF-23, Klotho and Egr1, and the expression of FGFR1 is up regulated, indicating the line in the parathyroid gland after ovariectomized. The inhibition of PTH expression is reduced, but the function of the receptor system after a prophylactic injection of beta estradiol is lower. These results suggest that ovariectomy and preventive administration may affect the ER- alpha, the VDR signal regulation system, the Ca~ (2+) /CaSR regulatory system and the FGF-23/Klotho receptor regulation system, and then regulate the table of PTH. Da.
【学位授予单位】：陕西理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R580

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