Shan Su1, Di Zhang1, Jinjin Liu2, Haiyan Zhao3, Xulei Tang2, Hongxia Che4, Qiangmei Wang1, Wanna Ren5, Donghu Zhen6. 1. The First Clinical Medical College of Lanzhou University, Lanzhou, 730000, Gansu Province, China. 2. Department of Endocrinology, The First Hospital of Lanzhou University, 1 Donggang West Road, Lanzhou, 730000, Gansu Province, China. 3. Department of Paediatrics, Gansu Province People's Hospital, Lanzhou, 730000, Gansu Province, China. 4. Department of Endocrinology, The Third People's Hospital, Lanzhou, 730000, Gansu Province, China. 5. Department of Opthalmology, The Second Hospital of Lanzhou University, Lanzhou, 730000, Gansu Province, China. 6. Department of Endocrinology, The First Hospital of Lanzhou University, 1 Donggang West Road, Lanzhou, 730000, Gansu Province, China. zhdh8279@163.com.
Abstract
INTRODUCTION: Homocysteine (Hcy) is considered a newly identified risk factor for osteoporosis. Nevertheless, the underlying mechanism of folate (FA), a key factor in the metabolism of Hcy, in protection against osteoblast dysfunction remains unclear. The purpose of this study was to investigate the mechanism by which FA attenuates Hcy-induced osteoblast damage. MATERIALS AND METHODS: The Hcy-induced MC3T3-E1 cells were treated with different concentrations of FA. Cell morphology, cell density, cell proliferation ability, alkaline phosphatase (ALP) activity and mineralization capacity were observed and determined; the gene expression of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (BAX) and ERS-associated factors, including glucose-regulated protein 78 (GRP-78), activating transcription factor 4 (ATF-4) and growth arrest and DNA damage inducible gene 153 (CHOP/GADD153), were assessed by RT-PCR; and protein levels of GRP-78 and ATF-4 were analyzed by western blotting. RESULTS: Hcy suppressed the proliferation, differentiation and mineralization ability of MC3T3-E1 cells in a concentration-dependent manner and activated the ERS signaling pathway. After intervention with different concentrations of FA, the cell viability and density, ALP activity, number of mineralized nodules, calcium content and Bcl-2 gene expression were all significantly increased, whereas the gene expression of GRP-78, CHOP/GADD153, ATF-4 and Bax was markedly downregulated, and protein levels of GRP-78 and ATF-4 were also markedly decreased. CONCLUSION: The adverse effects of Hcy on osteoblast differentiation are dose dependent. FA not only protects against osteoblasts apoptosis but also has a direct osteogenic effect on Hcy-induced osteoblasts, which could be partially mediated by inhibition of the PERK-activated ERS pathway.
INTRODUCTION: Homocysteine (Hcy) is considered a newly identified risk factor for osteoporosis. Nevertheless, the underlying mechanism of folate (FA), a key factor in the metabolism of Hcy, in protection against osteoblast dysfunction remains unclear. The purpose of this study was to investigate the mechanism by which FA attenuates Hcy-induced osteoblast damage. MATERIALS AND METHODS: The Hcy-induced MC3T3-E1 cells were treated with different concentrations of FA. Cell morphology, cell density, cell proliferation ability, alkaline phosphatase (ALP) activity and mineralization capacity were observed and determined; the gene expression of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (BAX) and ERS-associated factors, including glucose-regulated protein 78 (GRP-78), activating transcription factor 4 (ATF-4) and growth arrest and DNA damage inducible gene 153 (CHOP/GADD153), were assessed by RT-PCR; and protein levels of GRP-78 and ATF-4 were analyzed by western blotting. RESULTS: Hcy suppressed the proliferation, differentiation and mineralization ability of MC3T3-E1 cells in a concentration-dependent manner and activated the ERS signaling pathway. After intervention with different concentrations of FA, the cell viability and density, ALP activity, number of mineralized nodules, calcium content and Bcl-2 gene expression were all significantly increased, whereas the gene expression of GRP-78, CHOP/GADD153, ATF-4 and Bax was markedly downregulated, and protein levels of GRP-78 and ATF-4 were also markedly decreased. CONCLUSION: The adverse effects of Hcy on osteoblast differentiation are dose dependent. FA not only protects against osteoblasts apoptosis but also has a direct osteogenic effect on Hcy-induced osteoblasts, which could be partially mediated by inhibition of the PERK-activated ERS pathway.