The Pathogenesis Study Of Steroid Induced Osteonecrosis Of Femoral Head

Background:Glucocorticoids usage is the most common cause of non-traumatic osteonecrosis of femoral head (ONFH). Despite the intense reseach, the molecular pathogenesis remains obscure. Osteogenesis-angiogenesis coupling is the most important factor of bone remodelling, osteogenesis and angiogenesis inhibition both are evident in the pathology of ONFH.Purpose:To explore the mechanism of Osteogenesis-angiogenesis coupling inhibition in steroid induced ONFH, to observe the dexamethasone effect on proliferation and viablity of murine pre-osteoblast MC3T3-E1and Human Umbilical Vein Endothelial Cells (HUVEC).To investigate the molecular mechanism of this effect.Methods:Murine pre-osteoblast MC3T3-E1and HUVEC were adopted to study the effect of dexamethasone on proliferation, viability, autophagy, apoptosis and cell cycle of these two cell lines. Combined using bioinformatics analysis and biochemical experiment, the molecular mechanism of apoptosis and cell cycle arrest of MC3T3-E1cells investigated.Results:Part I dexamethasone at concentration of10-6M significantly inhibited the proliferation of MC3T3-E1, and lead to an increase of the proportion of dead cells. Further experiments found that dexamethasone inhibited the proliferation of MC3T3-E1cells through induction of apoptosis and G1phase cell cycle arrest, and this inhibitory effect were dependent of glucocorticoid receptor a (GRa) activation. By in silico protein-protein interaction prediction, P53was found to likely an effector molecule of GRa activation. Western blot confirmed that P53was upregulated by dexamethasone challenge and the upregulation was abolished by GRa inactivation through RU486pretreatment. P53downstream protein such as NOXA, cyclin D1were also upregulated by dexamethasone and reversed by RU486. But the non-specific P53inhibitor pifithrin-α can not reverse dex-induced apoptosis and cell cycle arrest of MC3T3-E1.PART Ⅱ HUVEC were challenged by10-9,10-8,10-7,10-6,10-5M dexamethasone for24h and48h,repectively, It was found that the proliferation of HUVEC was not affected by various concentration gradient of dexamethasone by CCK-8test. Autophagy, apoptosis and cell cycle arrest ware also absent in HUVEC stimulated by dexamethasone.Conclusion:dexamethasone significantly induced proliferation inhibition and apoptosis on MC3T3-E1, but had no effect on HUVEC's proliferation and viability. Osteogenesis and angiogenesis coupling inhibition by dexamethasone may through direct osteoblast inhibition but not endothelial cells.