Dissertation > Excellent graduate degree dissertation topics show

The Effect and Molecular Mechanism of Bone-targeted Estrogen on Human Osteoblasts

Author: ZhuYaQin
Tutor: LiLingZhi；WangYue
School: Hebei Medical University
Course: Pharmacology
Keywords: The rhein piperazine estrone Postmenopausal osteoporosis OPG NF-κB activation receptor ligands Interleukin-6 Estrogen Receptor Estrogen response element Mitotic activator of protein kinase
CLC: R580
Type: Master's thesis
Year: 2010
Downloads: 62
Quote: 0
Read: Download Dissertation

Abstract

Objective: With the aging of the human society, post-menopausal osteoporosis (postmenopausal osteoporosis, PMOP) has become a frequently-occurring disease that affects older women's quality of life, the primary cause of the reduction of post-menopausal estrogen levels. A bone targeting estrogen drugs in the treatment of the PMOP the while avoiding the side effects of estrogen on breast, ovarian, and other non-target organs, therefore, these drugs have been known as one of the hot spots of the current anti-osteoporosis drug research . The room to a bone targeting molecules rhein as a carrier for the mother, estrone synthesized the bone targeting estrogen Rhein piperazine estrone (rhein-piperizinyl-estrone, named LC), early studies have demonstrated that LC- ovariectomized rats has significant anti-osteoporosis activity, and can significantly promote the proliferation and differentiation of primary cultured rat osteoblasts. The topics for further study LC on human osteoblast-like cells MG-63 proliferation, differentiation, function, OPG (osteoprotegerin, OPG) / NF-kappa B activation receptor ligand (receptor activator of NF-kappa B of ligand, RANKL) / NF-κB is activated receptor (receptor activator of NF-κB, RANK) systems as well as interleukin (IL) -6 (interleukin-6, IL-6) and its role in receptor expression, and its possible mechanism of action was discussed. : 1.MTT method detection MG-63 cell proliferative capacity. 2. Flow cytometry cell cycle. 3. The disodium phosphate benzene was detected by alkaline phosphatase (of alkaline phosphatase, ALP) activity. 4. Reverse transcriptase polymerase chain reaction (reverse transcription polymerase chain reaction, RT-PCR) detection of OPG, RANKL, IL-6 and its receptor (IL-6Rα, gp130) mRNA expression levels. 5 enzyme-linked immunosorbent assay (enzyme-linked immnosorbent assay, ELISA) to detect the level of IL-6 protein expression. Immunoblotting (Western Blot) technical testing of OPG, RANKL, IL-6Rα and gp130 protein expression levels. 7 using RNA interference (RNA interference, RNAi), respectively, of the MG-63 cells estrogen receptor (estrogen receptor, ER) α and ERβ gene silencing, G418 screened positive monoclonal expand after culture with Western Blot respectively, identified ERα or ERβ high, and in inhibiting LC affinity of two subtypes of ER expression clones used to study whether there are differences. Application luciferase reporter gene assay and three kinds of mitogen-activated protein kinase (mitogen activated protein kinase, MAPK)-specific signal blocker LC role of ER signaling pathway. Results: LC on human osteoblast-like cells MG-63 biological properties of LC can significantly promote human osteoblast-like cells MG-63 proliferation in a dose-dependent manner, and make the G1 phase of the cell cycle by regulating the MG-63 reduced proportion of cells that the G2 S phase cells were increased in order to promote cell growth. The LC through enhanced stimulation of ALP activity of MG-63 cells into bone cells differentiation and maturation. LC in a dose-dependent manner significantly raised the MG-63 cell OPG mRNA and protein expression levels, significantly down-regulated RANKL mRNA and protein expression levels of 100 nM LC inhibition is the most significant. In addition, LC still capable of significant inhibition of MG-63 cells of IL-6 transcription level and protein level of expression; LC inhibited the expression levels of IL-6RαmRNA, while no effect on the protein level; LC gp130 transcriptional level expression and protein levels had no effect. LC molecular mechanisms of osteoblast role ER blocker ICI182780 (the trade name Fwy Division group, Fulvestrant) was completely blocked by the LC into bone cells, indicating that the LC ER pathway play a role. LC for ERα or ERβ high inhibitory expression cells were no significant differences, suggesting LC similar to the affinity for ER both subtypes. LC significantly enhanced transcription of the ER gene promoter activity, suggesting that the LC can by classical genomic mechanisms, i.e. the estrogen response element (Estrogen Response Element the ERE) gene transcription pathways play a role, while the MAPK-specific signal blocker PD98059, (ERK inhibition agent), SP600125, (JNK inhibitor), SB202190 (p38MAPK inhibitor) can be partially blocked by LC role that the role of the LC dependent non-genomic mechanisms, namely the MAPK signaling pathway. Conclusion: 1.LC can stimulate osteoblast proliferation, differentiation, and thus contributing to the growth of osteoblasts by regulating cell cycle. 2.LC can promote bone cells OPG expression, inhibition of RANKL, and IL-6 The expression. 3.LC ER ways osteoblasts play a role, and the two subtypes of ER ERα, ERβ is not selective. 4.LC osteoblasts through activation of ERE gene transcription pathway and MAPK signaling pathways play a role.

Related Dissertations

CLC: > Medicine, health > Internal Medicine > Endocrine diseases and metabolic diseases
© 2012 www.DissertationTopic.Net Mobile