The Research On The Protective Function And Signal Mechanism Of 17β-E₂ Homocysteine Induced To Produce The Oxidative Injury To Osteoblast

Objective:The purpose of this research was to investigate the protective and signal mechanism of 17β-Estradiol(17β-E2)in the homocysteine(Hcy)induced to produce oxidative injury model of the MG63(human)osteoblast-like cells.Methods:MG63(human)osteoblast-like cells were cultured and treated in different time in vitro. During the experiments,MG63(human)osteoblast-like cells were randomly divided into four groups, including the blank control group, the Hcy treatment group, the 17β-E2 treatment group and U73122,an specific inhibitor of PLC-yl;In treatment group, quantification of cell death was observed by using Hoechst33342/PI nuclear staining and fluorescence microscopy. Intracelluar ROS were measured by using the fluorescent probe,H2DCFDA,with spectrofluoro-photometer. The signal protein expression of Phospholipase C-yl(PLC-yl), extracellular signal-regulated kinase(ERK),p38 mitogen-activated protein kinases (p38MAPK),c-Jun N-terminal kinases(JNK),cysteine-aspartic acid protease 3 (Caspase-3)and bcl-2-Associated X Protein(Bax) were measured with Western Blotting.Results:The production of intracelluar ROS induced 1-5mM Hcy in human bone cell line MG63 was inhibited by 1μM 17β-E2 in a concentration-dependent and time-dependent manner.3mM Hcy could induce obviously human bone cell line MG63 apoptosis and necrosis in a time-dependent. More strikingly,1μM 17β-E2 could inhibit 3mM Hcy-induced apoptosis and necrosis.1μM 17P-E2 could inhibit obviously 3mM Hcy-induced down-regulation of PLC-yl and ERK phosphorylation, p38 MAPK and JNK phosphorylation,up-regulation Caspase-3 and Bax protein expression in human osteoblast-like cell line MG63. 1μM 17β-E2 and 7μM U73122 co-pretreated human osteoblast-like cell line MG63 could inhibit obviously down-regulation of PLC-yl and ERK phosphorylation,up-regulation of p38 MAPK phosphorylation,up-regulation of Caspase-3 and Bax protein expression,but JNK phosphorylation has not changed.Conclusions:1.High concentration of Hcy could induce the increase of intracelluar ROS and the number of necrosis in human osteoblast-like cell line MG63.17β-E2 had obvious role in protection and could suppress Hcy-induced necrosis of cells.2.High concentration of Hcy could regulate ERK,p38MAPK and JNK phosphorylation,and activation of Caspase-3 and Bax in human osteoblast-like cell line MG63.17β-E2 inhibited high concentration of Hcy-induced the change of above-mentioned signal transduction associated protein in a PLC-yl-dependent manner.Accordingly,it brought into full protective function.