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Qiu Y.Y.,The 2Nd Shenzhen Peoples Hospital | Chen Y.,The 2Nd Shenzhen Peoples Hospital | Zeng T.H.,The 2Nd Shenzhen Peoples Hospital | Guo W.H.,The 2Nd Shenzhen Peoples Hospital | And 2 more authors.
Genetics and Molecular Research | Year: 2016

Chondrocytes, which are embedded within the growth-plate or the intervertebral disc, are sensitive to environmental stresses, such as inflammation and hypoxia. However, little is known about the molecular signaling pathways underlying hypoxia-induced mitochondrial dysfunction and apoptosis in chondrocytes. We first examined the apoptosis, caspase-3 activity, and apoptosis-associated markers in human chondrocyte cell line C28/I2 under normoxia or hypoxia. We then investigated mitochondrial dysfunction and the activation of cyclic adenosine monophosphate response element-binding protein (CREB) signaling in the same human chondrocyte cell line. Our results indicated that hypoxia induced apoptosis and reduced CREB phosphorylation in chondrocytes. Upregulated mitochondrial superoxide and reactive oxygen species levels, and reduced mitochondrial membrane potential and complex IV activity were observed in hypoxia-treated C28/I2 cells. In conclusion, the present study confirmed reduced CREB phosphorylation, apoptosis induction, and mitochondrial dysfunction in the hypoxia-treated chondrocyte cells. This implies the key role played by CREB signaling in hypoxiainduced mitochondrial dysfunction and apoptosis in chondrocytes. © FUNPEC-RP.


Qiu Y.,The 2Nd Shenzhen Peoples Hospital | Chen Y.,The 2Nd Shenzhen Peoples Hospital | Zeng T.,The 2Nd Shenzhen Peoples Hospital | Guo W.,The 2Nd Shenzhen Peoples Hospital | And 2 more authors.
Molecular Biology Reports | Year: 2016

The healing process of fractured bone is affected by the multiple factors regulating the growth and differentiation of osteoblasts and bone mesenchymal stem cells (MSCs), however, such markers and molecular events need to be orchestrated in details. This study investigated the effect of polyphenol(-)-epigallocatechin-3-gallate (EGCG) on the hypoxia-induced apoptosis and osteogenic differentiation of human bone marrow-derived MSCs, examined the miR-210 induction by EGCG, explored the target inhibition of the expression of receptor tyrosine kinase ligand ephrin-A3 (EFNA3) by miR-210, and then determined the association of the miR-210 promotion with the hypoxia-induced apoptosis and osteogenic differentiation. Results demonstrated that EGCG treatment significantly inhibited the hypoxia-induced apoptosis in MSCs and promoted the level of alkaline phosphatase (ALP), bone morphogenetic protein 2 (BMP-2), propeptide of type I procollagen I (PINP) and runt-related transcription factor 2 (RUNX2) in MSCs under either normoxia or hypoxia. Moreover, the EGCG treatment upregulated the miR-210 expression, in an association with EFNA3 downregulation; and the miR-210 upregulation significantly downregulated the expression of EFNA3 via the specific binding to the 3′ UTR of EFNA3. In addition, the manipulated miR-210 upregulation exerted amelioration on the hypoxia-induced apoptosis and on the hypoxia-reduced expression of ALP, BMP-2, PINP and RUNX2 in MSCs. In summary, our study indicated the protective role of EGCG in response to hypoxia and promontory role to osteogenic differentiation in MSCs via upregulating miR-210 and downregulating the expression of miR-210-targeted EFNA3. Our study implies the protective role of EGCG in the hypoxia-induced impairment in MSCs. © 2016, Springer Science+Business Media Dordrecht.


Qiu Y.,The 2nd Shenzhen Peoples Hospital | Chen Y.,The 2nd Shenzhen Peoples Hospital | Zeng T.,The 2nd Shenzhen Peoples Hospital | Guo W.,The 2nd Shenzhen Peoples Hospital | And 2 more authors.
Cell Biology International | Year: 2016

High-mobility group box 1 (HMGB1) is a nuclear protein that involves the binding with DNA and influences chromatin regulation and transcription. HMGB1 activates monocytes and neutrophils, which are involved in inflammation during wounding. In this study, we investigated the promotion of HMGB1 under hypoxia and determined the regulatory role of HMGB1 on the fibrosis of mouse osteoblast-like MC3T3-E1 cells or of human osteoblast MG-63 cells. Results demonstrated that HMGB1 expression was significantly upregulated in MC3T3-E1 or MG-63 cells under hypoxia. We also found that treatment with 10 and 100 ng/mL of HMGB1 significantly promoted the fibrosis-associated markers such as Collagen I, α-SMA, whereas downregulated the E-cadherin, indicating the differentiation of MC3T3-E1 or MG-63 cells into fibroblast cells. Further investigation indicated that the HMGB1 treatment markedly activated the mitogen-activated protein kinases (MAPKs), including extracellular signal-related kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (p38) phosphorylation, as well as nuclear factor (NF)-κB nuclear translocation. On the other side, using specific inhibitors and shRNAs of protein kinases, we observed that repression of ERK, JNK, p38, and NF-κB all inhibited HMGB1-induced cellular differentiation and migration of MC3T3-E1 cells. In addition, knocking down of advanced glycation end products (RAGE) but not Toll-like receptor (TLR)2 and TLR4 by shRNAs attenuated HMGB1-induced myofibroblast differentiation and migration. In conclusion, our study demonstrated that HMGB1 induced the fibrosis of osteoblasts in vitro via activating the RAGE-MAPK and NF-κB interaction signaling pathways. © 2016 International Federation for Cell Biology


PubMed | The 2nd Shenzhen Peoples Hospital
Type: Journal Article | Journal: Cell biology international | Year: 2016

High-mobility group box 1 (HMGB1) is a nuclear protein that involves the binding with DNA and influences chromatin regulation and transcription. HMGB1 activates monocytes and neutrophils, which are involved in inflammation during wounding. In this study, we investigated the promotion of HMGB1 under hypoxia and determined the regulatory role of HMGB1 on the fibrosis of mouse osteoblast-like MC3T3-E1 cells or of human osteoblast MG-63 cells. Results demonstrated that HMGB1 expression was significantly upregulated in MC3T3-E1 or MG-63 cells under hypoxia. We also found that treatment with 10 and 100ng/mL of HMGB1 significantly promoted the fibrosis-associated markers such as Collagen I, -SMA, whereas downregulated the E-cadherin, indicating the differentiation of MC3T3-E1 or MG-63 cells into fibroblast cells. Further investigation indicated that the HMGB1 treatment markedly activated the mitogen-activated protein kinases (MAPKs), including extracellular signal-related kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (p38) phosphorylation, as well as nuclear factor (NF)-B nuclear translocation. On the other side, using specific inhibitors and shRNAs of protein kinases, we observed that repression of ERK, JNK, p38, and NF-B all inhibited HMGB1-induced cellular differentiation and migration of MC3T3-E1 cells. In addition, knocking down of advanced glycation end products (RAGE) but not Toll-like receptor (TLR)2 and TLR4 by shRNAs attenuated HMGB1-induced myofibroblast differentiation and migration. In conclusion, our study demonstrated that HMGB1 induced the fibrosis of osteoblasts in vitro via activating the RAGE-MAPK and NF-B interaction signaling pathways.


PubMed | The 2nd Shenzhen Peoples Hospital
Type: Journal Article | Journal: Genetics and molecular research : GMR | Year: 2016

Chondrocytes, which are embedded within the growth-plate or the intervertebral disc, are sensitive to environmental stresses, such as inflammation and hypoxia. However, little is known about the molecular signaling pathways underlying hypoxia-induced mitochondrial dysfunction and apoptosis in chondrocytes. We first examined the apoptosis, caspase-3 activity, and apoptosis-associated markers in human chondrocyte cell line C28/I2 under normoxia or hypoxia. We then investigated mitochondrial dysfunction and the activation of cyclic adenosine monophosphate response element-binding protein (CREB) signaling in the same human chondrocyte cell line. Our results indicated that hypoxia induced apoptosis and reduced CREB phosphorylation in chondrocytes. Upregulated mitochondrial superoxide and reactive oxygen species levels, and reduced mitochondrial membrane potential and complex IV activity were observed in hypoxia-treated C28/I2 cells. In conclusion, the present study confirmed reduced CREB phosphorylation, apoptosis induction, and mitochondrial dysfunction in the hypoxia-treated chondrocyte cells. This implies the key role played by CREB signaling in hypoxia-induced mitochondrial dysfunction and apoptosis in chondrocytes.


PubMed | The 2nd Shenzhen Peoples Hospital
Type: Journal Article | Journal: Molecular biology reports | Year: 2016

The healing process of fractured bone is affected by the multiple factors regulating the growth and differentiation of osteoblasts and bone mesenchymal stem cells (MSCs), however, such markers and molecular events need to be orchestrated in details. This study investigated the effect of polyphenol(-)-epigallocatechin-3-gallate (EGCG) on the hypoxia-induced apoptosis and osteogenic differentiation of human bone marrow-derived MSCs, examined the miR-210 induction by EGCG, explored the target inhibition of the expression of receptor tyrosine kinase ligand ephrin-A3 (EFNA3) by miR-210, and then determined the association of the miR-210 promotion with the hypoxia-induced apoptosis and osteogenic differentiation. Results demonstrated that EGCG treatment significantly inhibited the hypoxia-induced apoptosis in MSCs and promoted the level of alkaline phosphatase (ALP), bone morphogenetic protein 2 (BMP-2), propeptide of type I procollagen I (PINP) and runt-related transcription factor 2 (RUNX2) in MSCs under either normoxia or hypoxia. Moreover, the EGCG treatment upregulated the miR-210 expression, in an association with EFNA3 downregulation; and the miR-210 upregulation significantly downregulated the expression of EFNA3 via the specific binding to the 3 UTR of EFNA3. In addition, the manipulated miR-210 upregulation exerted amelioration on the hypoxia-induced apoptosis and on the hypoxia-reduced expression of ALP, BMP-2, PINP and RUNX2 in MSCs. In summary, our study indicated the protective role of EGCG in response to hypoxia and promontory role to osteogenic differentiation in MSCs via upregulating miR-210 and downregulating the expression of miR-210-targeted EFNA3. Our study implies the protective role of EGCG in the hypoxia-induced impairment in MSCs.

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