PMID- 29763908 OWN - NLM STAT- MEDLINE DCOM- 20180821 LR - 20191224 IS - 1421-9778 (Electronic) IS - 1015-8987 (Linking) VI - 47 IP - 1 DP - 2018 TI - Connexin 43 Modulates Osteogenic Differentiation of Bone Marrow Stromal Cells Through GSK-3beta/Beta-Catenin Signaling Pathways. PG - 161-175 LID - 10.1159/000489763 [doi] AB - BACKGROUND/AIMS: Bone marrow stromal cells (BMSCs) are multipotent precursors that give rise to osteoblasts, and contribute directly to bone formation. Connexin 43 (Cx43) is the most ubiquitous gap junction protein expressed in bone cell types, and plays crucial roles in regulating intercellular signal transmission for bone development, differentiation and pathology. However, the precise role and mechanism of Cx43 in BMSCs are less known. Here, we investigate the function of Cx43 in osteogenic differentiation of BMSCs in vitro. METHODS: BMSCs were isolated by whole bone marrow adherent culture. Knock down of Cx43 was performed by using lentiviral transduction of Cx43 shRNA. BMSCs were induced to differentiate by culturing in a-MEM, 10% FBS, 50 microM ascorbic acid, 10 mM beta-glycerophosphate, and 100 nM dexamethasone. Alkaline phosphatase (ALP) activity and alizarin red S staining were used to evaluate osteogenic differentiation in calcium nodules. Target mRNAs and proteins were analyzed by using real-time quantitative PCR (qPCR) and western blotting. RESULTS: Cx43 expression markedly increased during osteogenic differentiation. Osteogenic differentiation was suppressed following lentiviral-mediated knockdown of Cx43 expression, as judged by decreased levels of Runt-related transcription factor 2 (Runx2), bone sialoprotein (BSP), osteocalcin (Bglap), Osterix (Osx), alkaline phosphatase (ALP) activity and the number of calcium nodules in response to osteogenic differentiation stimuli. Knock down of Cx43 reduced the level of phosphorylation of GSK-3beta at Ser9 (p-GSK-3beta), resulting in decreased beta-catenin expression and activation. Furthermore, treatment of Cx43-knockdown cells with lithium chloride (LiCl), a GSK-3beta inhibitor, reduced osteogenic differentiation and decreased GSK-3beta levels, as well as partially rescued levels of both total and activated beta-catenin. CONCLUSION: These findings indicate that Cx43 positively modulates osteogenic differentiation of BMSCs by up-regulating GSK-3beta/beta-catenin signaling pathways, suggesting a potential role for Cx43 in determining bone mass and bone mineral density by modulating osteogenesis. CI - (c) 2018 The Author(s). Published by S. Karger AG, Basel. FAU - Lin, Fei-Xiang AU - Lin FX AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. AD - Department of Spine Surgery, The Affiliated Ganzhou Hospital of Nanchang University (Ganzhou People's Hospital), Ganzhou, China. FAU - Zheng, Gui-Zhou AU - Zheng GZ AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. FAU - Chang, Bo AU - Chang B AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. AD - Department of Neurological Surgery, The First Affiliated Hospital of Shantou University Medical College, Shantou, China. FAU - Chen, Rong-Chun AU - Chen RC AD - Department of Spine Surgery, The Affiliated Ganzhou Hospital of Nanchang University (Ganzhou People's Hospital), Ganzhou, China. FAU - Zhang, Qi-Hao AU - Zhang QH AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. AD - Department of Neurological Surgery, The First Affiliated Hospital of Shantou University Medical College, Shantou, China. FAU - Xie, Peng AU - Xie P AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. FAU - Xie, Da AU - Xie D AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. AD - Department of Neurological Surgery, The First Affiliated Hospital of Shantou University Medical College, Shantou, China. FAU - Yu, Guo-Yong AU - Yu GY AD - Department of Neurological Surgery, The First Affiliated Hospital of Shantou University Medical College, Shantou, China. FAU - Hu, Qin-Xiao AU - Hu QX AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. AD - Department of Neurological Surgery, The First Affiliated Hospital of Shantou University Medical College, Shantou, China. FAU - Liu, De-Zhong AU - Liu DZ AD - Department of Neurological Surgery, The First Affiliated Hospital of Shantou University Medical College, Shantou, China. FAU - Du, Shi-Xin AU - Du SX AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. FAU - Li, Xue-Dong AU - Li XD AD - Department of Orthopedics, The Third Affiliated Hospital (The Affiliated Luohu Hospital) of Shenzhen University, Shenzhen, China. LA - eng PT - Journal Article DEP - 20180510 PL - Germany TA - Cell Physiol Biochem JT - Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology JID - 9113221 RN - 0 (Connexin 43) RN - 0 (Gja1 protein, rat) RN - 0 (beta Catenin) RN - EC 2.7.11.1 (Glycogen Synthase Kinase 3 beta) SB - IM MH - Animals MH - Cell Differentiation MH - Cells, Cultured MH - Connexin 43/*metabolism MH - Glycogen Synthase Kinase 3 beta/*metabolism MH - Mesenchymal Stem Cells/*cytology/metabolism MH - *Osteogenesis MH - Rats, Sprague-Dawley MH - *Signal Transduction MH - beta Catenin/*metabolism OTO - NOTNLM OT - Beta-catenin OT - Bone marrow stromal cells OT - Connexin 43 OT - GSK-3beta OT - Osteogenic differentiation EDAT- 2018/05/16 06:00 MHDA- 2018/08/22 06:00 CRDT- 2018/05/16 06:00 PHST- 2017/10/10 00:00 [received] PHST- 2018/03/05 00:00 [accepted] PHST- 2018/05/16 06:00 [pubmed] PHST- 2018/08/22 06:00 [medline] PHST- 2018/05/16 06:00 [entrez] AID - 000489763 [pii] AID - 10.1159/000489763 [doi] PST - ppublish SO - Cell Physiol Biochem. 2018;47(1):161-175. doi: 10.1159/000489763. Epub 2018 May 10.