Journal articles on the topic 'Western Alp'
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1
Feng, Gaoke, Chaoshi Qin, Fei Sha, Yongnan Lyu, Jinggang Xia, and Xuejun Jiang. "Evaluation of Inflammatory and Calcification after Implantation of Bioabsorbable Poly-L-Lactic Acid/Amorphous Calcium Phosphate Scaffolds in Porcine Coronary Arteries." Journal of Nanomaterials 2021 (January 23, 2021): 1–8. http://dx.doi.org/10.1155/2021/6652648.
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Purpose. Our previous research has confirmed that the addition of nano-amorphous calcium phosphate (ACP) materials can improve the support of poly-L-lactic acid (PLLA) vascular scaffolds. Based on this, we continued to explore the effect of novel bioresorbable scaffold composed of PLLA and ACP nanoparticles on inflammation and calcification of surrounding tissues after scaffold implantation in porcine coronary artery. Methods. PLLA/ACP scaffolds in the experimental group and PLLA scaffolds in the control group were implanted into the coronary arteries of small pigs. Serum levels of C-reactive protein (CRP), calcium (Ca), and alkaline phosphatase (ALP) were measured before implantation and at 1, 6, 12, and 24 months after operation. Intravascular ultrasonography (IVUS) was performed to evaluate the vascular calcification score. The scaffold and surrounding tissues were hematoxylin-eosin staining for inflammation score. The scaffold and surrounding tissues were stained with NF-κB and ALP, and the positive expression index was calculated. Western blot was used to detect the expression of IL-6 and BMP-2 in the tissues around the scaffold. Results. There was no statistically significant difference between the two groups in CRP, calcium, and ALP at preimplant, 1 month, 6 months, 12 months, and 24 months ( P > 0.05 ). The inflammation score, NF-κB positive expression index, and calcification score in the PLLA/ACP group were lower than that in the PLLA group at 12 months and 24 months ( P < 0 05 ). The ALP positive expression index in the PLLA/ACP group was lower than that in the PLLA group at 6 months, 12 months, and 24 months ( P < 0 05 ). Western blot results showed that the IL-6 expression level in the PLLA/ACP group was significantly lower than that in the control group at 6 months, 12 months, and 24 months ( P < 0.05 ). The expression of BMP-2 in the PLLA/ACP group was significantly lower than that in the control group at 12 months and 24 months ( P < 0.05 ). Conclusion. The PLLA/ACP composite scaffold has good biocompatibility. The incorporation of nanoscale ACP can reduce the inflammatory response caused by the acid metabolites of PLLA scaffolds, reduce the expression of procalcification factors in the body, and inhibit tissue calcification. The PLLA/ACP composite scaffold provides reliable guidance for the application and development of degradable vascular scaffold.
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Xuan, Qiang, Xiaoli Yang, Linjian Mo, Fengyu Huang, Youhong Pang, Min Qin, Zhiqiang Chen, Min He, Qi Wang, and Zeng-Nan Mo. "Expression of the Serine Protease Kallikrein 7 and Its Inhibitor Antileukoprotease Is Decreased in Prostate Cancer." Archives of Pathology & Laboratory Medicine 132, no. 11 (November 1, 2008): 1796–801. http://dx.doi.org/10.5858/132.11.1796.
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Abstract Context.—Kallikreins are a subgroup of serine proteases with diverse physiologic functions. It has been confirmed that kallikrein 7 (KLK7) is differentially expressed in ovarian and breast cancer. Antileukoprotease (ALP) has been shown to be a specific inhibitor of human kallikrein 7 (hK7). Antileukoprotease overexpression is commonly associated with aggressive, high-risk, or metastatic cancer originating from various organs. Objective.—To investigate the expression and potential role of hK7 and its inhibitor ALP in prostate cancer. Design.—The mRNA expression of KLK7 and ALP transcript in benign prostate epithelial cells and prostate cancers was evaluated by semiquantitative reverse transcription–polymerase chain reaction. We examined hK7 and ALP protein expression by immunohistochemistry in 20 normal prostate tissues, 50 benign prostatic hyperplasia tissues, and 103 prostate cancers. Western blot examination showed protein expression of hK7 and ALP in benign prostate epithelial cells and prostate cancer cell lines. Results.—Semiquantitative polymerase chain reaction examination revealed that the mRNA level of KLK7 and ALP was significantly decreased in prostate cancers compared with that in benign prostate epithelial cells (P < .001). Immunohistochemical expression of hK7 was observed in prostate epithelial cells, whereas little or no staining was observed in prostate cancer. Western blot analysis revealed that hK7 and ALP were decreased in malignant prostate epithelium. Conclusions.—Like hK7, ALP is down-regulated in prostate cancers, which begs the question of whether it remains an effective inhibitor of hK7 or whether it is discordant in time or space and is ineffective as an inhibitor of hK7. The function of KLK7 and ALP in prostate cancer should be further studied.
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Xiao, Wen-lin, Dai-zun Zhang, Cun-hui Fan, and Bao-jun Yu. "Intermittent Stretching and Osteogenic Differentiation of Bone Marrow Derived Mesenchymal Stem Cells via the p38MAPK-Osterix Signaling Pathway." Cellular Physiology and Biochemistry 36, no. 3 (2015): 1015–25. http://dx.doi.org/10.1159/000430275.
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Aims: The relationship between the p38MAPK signaling pathway and osterix in osteogenic differentiation of BMMSCs subjected to intermittent stretching was investigated. Methods: BMMSCs derived from C57BL/6J mice were divided into the following groups: 1) control, 2) stretch, and 3) SB203580+stretch (SB203580 is a p38MAPK signal pathway inhibitor). BMMSCs were exposed to an intermittent mechanical strain of 0.8% (8000μ strain) at 0.5 Hz, twice a day for 30 min each application. BMMSCs were harvested on days 1, 3, and 5 post-treatment. The expression of ALP, COL I, OCN, and osterix mRNA was assessed utilizing RT-PCR while the expression of P-p38MAPK and osterix protein was assessed by Western blot analysis. The osterix gene in mouse BMMSCs was knocked down using RNAi technology and its protein expression was also assessed by Western blot. RT-PCR was used to detect ALP, COL I, and OCN mRNA expression. Results: Intermittent stretching was found to promote expression of ALP, COL I, OCN, and osterix mRNA. Silencing the osterix gene was found to reduce levels of ALP, COL I, and OCN mRNA. Western blot analysis demonstrated that the levels of osterix and P-p38MAPK proteins in the stretch group were significantly higher than in the control group (P<0.05). There was less expression of ALP, COL I, OCN, and osterix mRNA in the SB203580+stretch group than in the control and stretch groups. Conclusions: Data demonstrate that intermittent stretching promotes osteogenic differentiation of BMMSCs, and the p38MAPK-osterix pathway has an important role in the control of osteogenesis-related gene expression.
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Schmidt, Yvonne, Martin Biniossek, G. Björn Stark, Günter Finkenzeller, and Filip Simunovic. "Osteoblastic alkaline phosphatase mRNA is stabilized by binding to vimentin intermediary filaments." Biological Chemistry 396, no. 3 (March 1, 2015): 253–60. http://dx.doi.org/10.1515/hsz-2014-0274.
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Abstract Vascularization is essential in bone tissue engineering and recent research has focused on interactions between osteoblasts (hOBs) and endothelial cells (ECs). It was shown that cocultivation increases the stability of osteoblastic alkaline phosphatase (ALP) mRNA. We investigated the mechanisms behind this observation, focusing on mRNA binding proteins. Using a luciferase reporter assay, we found that the 3′-untranslated region (UTR) of ALP mRNA is necessary for human umbilical vein endothelial cells (HUVEC)-mediated stabilization of osteoblastic ALP mRNA. Using pulldown experiments and nanoflow-HPLC mass spectrometry, vimentin was identified to bind to the 3′-UTR of ALP mRNA. Validation was performed by Western blotting. Functional experiments inhibiting intermediate filaments with iminodipropionitrile and specific inhibition of vimentin by siRNA transfection showed reduced levels of ALP mRNA and protein. Therefore, ALP mRNA binds to and is stabilized by vimentin. This data add to the understanding of intracellular trafficking of ALP mRNA, its function, and have possible implications in tissue engineering applications.
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Wundersitz, Sebastian, Cristina Pablo Tortola, Sibylle Schmidt, Ramon Oliveira Vidal, Melanie Kny, Alexander Hahn, Lukas Zanders, et al. "The Transcription Factor EB (TFEB) Sensitizes the Heart to Chronic Pressure Overload." International Journal of Molecular Sciences 23, no. 11 (May 25, 2022): 5943. http://dx.doi.org/10.3390/ijms23115943.
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The transcription factor EB (TFEB) promotes protein degradation by the autophagy and lysosomal pathway (ALP) and overexpression of TFEB was suggested for the treatment of ALP-related diseases that often affect the heart. However, TFEB-mediated ALP induction may perturb cardiac stress response. We used adeno-associated viral vectors type 9 (AAV9) to overexpress TFEB (AAV9-Tfeb) or Luciferase-control (AAV9-Luc) in cardiomyocytes of 12-week-old male mice. Mice were subjected to transverse aortic constriction (TAC, 27G; AAV9-Luc: n = 9; AAV9-Tfeb: n = 14) or sham (AAV9-Luc: n = 9; AAV9-Tfeb: n = 9) surgery for 28 days. Heart morphology, echocardiography, gene expression, and protein levels were monitored. AAV9-Tfeb had no effect on cardiac structure and function in sham animals. TAC resulted in compensated left ventricular hypertrophy in AAV9-Luc mice. AAV9-Tfeb TAC mice showed a reduced LV ejection fraction and increased left ventricular diameters. Morphological, histological, and real-time PCR analyses showed increased heart weights, exaggerated fibrosis, and higher expression of stress markers and remodeling genes in AAV9-Tfeb TAC compared to AAV9-Luc TAC. RNA-sequencing, real-time PCR and Western Blot revealed a stronger ALP activation in the hearts of AAV9-Tfeb TAC mice. Cardiomyocyte-specific TFEB-overexpression promoted ALP gene expression during TAC, which was associated with heart failure. Treatment of ALP-related diseases by overexpression of TFEB warrants careful consideration.
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Wang, Yirui, Zhixiong Liang, Liang Chen, Guosheng Yang, Jing Xu, Chunmei Deng, Chun Wang, and Changjiang Lei. "Protective Effect of Iron Oxide Nanoparticles on Periodontal Injury in Rats by Inhibiting Collagenase-1 and Alkaline Phosphatase Expression." Journal of Biomedical Nanotechnology 18, no. 4 (April 1, 2022): 1131–37. http://dx.doi.org/10.1166/jbn.2022.3322.
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This study intends to assess whether iron oxide nanoparticles affect periodontal injury and collagenase-1 (COL-1), and alkaline phosphatase (ALP) in rats. In this study, the ALP activity and Col-1 concentration in rats with periodontal injury were determined.We detected the periodontal histopathological changes and expression of periodontal pocket depth (PD) and attachment loss (AL) by Hematoxylin and eosin (HE) staining.We also detected Col-1 and ALP proteins in periodontal tissues by Western blot. Real-time reverse transcription–polymerase chain reaction (RT-PCR) detected Col-1 and ALP mRNA level in periodontal tissues of rats in each group, while ALP activity and Col-1 concentration in gingival crevicular fluid in model group increased compared to sham group (P < 0.05). After intervention by iron oxide nanoparticles, ALP activity and Col-1 concentration in the gingival crevicular fluid of model rats decreased greatly (P < 0.05). The gingival atrophy was more serious in model group, and many inflammatory cells infiltrated into the tissue and destroyed the alveolar tissue. Meanwhile, the periodontal tissue from rats in intervention group was greatly improved, and the degree of alveolar bone destruction was also significantly reduced, while the PD and AL periodontal indexes were significantly inhibited (P < 0.05). The protein and relative expression showed that the protein and mRNA expressions of ALP and Col-1 in periodontal tissue from model group were lower than those in sham group (P < 0.05). After intervention by iron oxide nanoparticles, the protein and mRNA expressions of ALP and Col-1 in the periodontal tissues in intervention group increased (P < 0.05). Iron oxide nanoparticles can thus inhibit the expression of ALP and COL-1 in periodontal injury rats, and improve the periodontal injury tissue.
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Guo, Meiling, Fen Liu, Wenjuan Wang, Zhirong Liu, Zhipeng Zhu, Yiyu Liu, and Zhen Huang. "Naringin Promotes Osteogenic/Odontogenic Differentiation of Dental Pulp Stem Cells via Wnt/β-Catenin." Evidence-Based Complementary and Alternative Medicine 2022 (May 29, 2022): 1–8. http://dx.doi.org/10.1155/2022/4505471.
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Purpose. This investigation intended to unravel the effect and mechanism of naringin on the proliferation and osteogenic differentiation of human dental pulp stem cells (hDPSCs). Methods. hDPSCs were induced to differentiate, and the degree of cell differentiation was observed by alizarin red staining, Oil Red O staining, and Alcian blue staining. hDPSCs were treated with 0, 20, 40, and 80 μmol/L naringin for 48 h, respectively. The proliferation rate and chemotaxis of the cells were measured by MTT and transwell assay, alkaline phosphatase (ALP) activity and osteogenic differentiation degree by ALP staining and alizarin red staining, and gene expression of osteogenic markers by qRT-PCR. Additionally, western blot was performed to test the levels of Wnt/β-catenin signaling-related proteins in hDPSCs. Results. The isolated hDPSCs with spindle-shaped morphology had good differentiation capability. Further experiments confirmed naringin-caused increases in the proliferation rate and migration ability of hDPSCs. In addition, compared with the control group, naringin-treated cells had strong ALP activity and ossification levels and higher expression of Runx2, OPN, DSPP, and DMP1. The western blot results showed that naringin significantly activated Wnt/β-catenin signaling in hDPSCs. Conclusion. Taken together, naringin enhances the proliferation, migration, and osteogenesis of hDPSCs through stimulating Wnt/β-catenin signaling pathway.
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Hao, Yan-Jie, Xin Jiang, Wei Zhou, Yong Wang, Lan Gao, Yu Wang, Guang-Tao Li, et al. "Connective tissue disease-associated pulmonary arterial hypertension in Chinese patients." European Respiratory Journal 44, no. 4 (May 2, 2014): 963–72. http://dx.doi.org/10.1183/09031936.00182813.
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We sought to investigate the characteristics, survival and risk factors for mortality in Chinese patients with connective tissue disease (CTD)-associated pulmonary arterial hypertension (APAH) in modern therapy era.129 consecutive adult patients who visited one of three referral centres in China with a diagnosis of CTD-APAH confirmed by right heart catheterisation during the previous 5 years were enrolled. The end-point was all-cause death or data censoring.Systemic lupus erythematosus was the most common underlying CTD (49%) and systemic sclerosis just accounted for 6% in this cohort. The overall survival at 1 and 3 years was 92% and 80%, respectively. Pericardial effusion, a shorter 6-min walk distance, lower mixed venous oxygen saturation, higher pulmonary vascular resistance (PVR) and alkaline phosphatase (ALP), and lower total cholesterol levels were all associated with a higher risk of death among the study population. Higher PVR and ALP were independent predictors of mortality.In conclusion, unlike in western patients, systemic lupus erythematosus is the most common underlying disease in Chinese patients with CTD-APAH. The survival of Chinese patients with CTD-APAH in the modern treatment era is similar to that in western countries. Elevated PVR and ALP are independent risk factors for poor outcomes.
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Zhou, Shuzuo, Gang Zhang, Kun Wang, Zhong Yang, and Yinghui Tan. "miR-141-3p Targeted SIRT1 to Inhibit Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells." Stem Cells International 2023 (February 3, 2023): 1–7. http://dx.doi.org/10.1155/2023/9094092.
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Purpose. To explore the expression of miR-141-3p during the osteogenic differentiation of human bone marrow mesenchymal stem cells (BMSCs) and its regulatory effect. Methods. Differentiation of BMSCs was induced by dexamethasone. The mRNA expression of miR-141-3p, ALP, RUNX2, and OCN was measured using RT-qPCR. The protein expression was detected via western blot. The target of miR-141-3p was predicted through the TargetScan website and confirmed using luciferase reporter assay. Results. miR-141-3p expression declined during osteogenic differentiation. The relative ALP activities and the mRNA expression of ALP, RUNX2, and OCN were markedly reduced in the miR-141-3p mimic group while increased in the inhibitor group. Cell viability was suppressed in the miR-141-3p mimic group and promoted in the inhibitor group. SIRT1 was predicted to be a downstream gene of miR-141-3p, and this prediction was confirmed via the luciferase reporter assay. The results of the western blot assay demonstrated that SIRT1 expression was decreased in the miR-141-3p mimic group. SIRT1 reversed the inhibitory influence of miR-141-3p on the osteogenic differentiation ability of BMSCs. Conclusion. miR-141-3p targeted SIRT1 to inhibit osteogenic differentiation of BMSCs via the Wnt/β-catenin signaling pathway.
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Gao, Bingjun, Yarong Wu, Lijian Zhou, and Xin Chen. "MicroRNA-595 promotes osteogenic differentiation of bone marrow mesenchymal stem cells by targeting HMGA2." Tropical Journal of Pharmaceutical Research 21, no. 3 (May 28, 2022): 457–63. http://dx.doi.org/10.4314/tjpr.v21i3.1.
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Purpose: To investigate the effect of miR-595 on osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs).Methods: Human BMSCs were osteogenically differentiated, and protein expression of alkaline phosphatase (ALP), osteocalcin (OCN), and Runt-related transcription factor 2 (RUNX2) were evaluated by western blot. Expression of miR-595 was measured by quantitative reverse transcription (qRT-PCR). The effect of miR-595 on viability of BMSCs was determined by MTT assay. Osteogenic differentiation of BMSCs was assessed by ALP and Alizarin red S (ARS) staining. The target gene of miR-595 was predicted by TargetScan analysis and validated by luciferase activity assay.Results: MiR-595 expression was higher in osteogenically differentiated BMSCs than in undifferentiated BMSCs (p < 0.01). Osteogenic ALP, OCN, and RUNX2 were also upregulated (p < 0.01). MiR-595 expression increased the viability of BMSCs, mineralized bone matrix formation, and ALP activity. High mobility group AT-hook 2 (HMGA2) expression was lower in osteogenically differentiated BMSCs and was found to be a target of miR-595. Overexpression of HMGA2 attenuated the miR-595-induced increase in cell viability, ALP activity, mineralized bone matrix formation, and osteogenic gene expression in BMSCs.Conclusion: The miR-595/HMGA2 axis is involved in osteogenic differentiation of BMSCs suggesting that it is a promising therapeutic target for osteoporosis.
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Wang, Yanping, Yanqiu Wang, Yadie Lu, and Jinhua Yu. "High Glucose Enhances the Odonto/Osteogenic Differentiation of Stem Cells from Apical Papilla via NF-KappaB Signaling Pathway." BioMed Research International 2019 (April 8, 2019): 1–10. http://dx.doi.org/10.1155/2019/5068258.
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Objective. The transport and metabolism of glucose are important during mammalian development. High glucose can mediate the biological characteristics of mesenchymal stem cells (MSCs). However, the role of high glucose in the odonto/osteogenic differentiation of stem cells from apical papilla (SCAPs) is unclear. Materials and Methods. SCAPs were isolated and identified in vitro. Then, SCAPs were cultured in normal α-MEM and high glucose α-MEM separately. MTT assay was applied to observe the proliferation of SCAPs. ALP activity, alizarin red staining, real-time RT-PCR, and western blot were used to detect the odonto/osteogenic capacity of SCAPs as well as the participation of NF-κB pathway. Results. SCAPs in 25mmol/L glucose group expressed the maximum proteins of RUNX2 and ALP as compared with those in 5, 10, and 15 mmol/L groups. MTT assay showed that 25 mmol/L glucose suppressed the proliferation of SCAPs. ALP assay, alizarin red staining, real-time RT-PCR, and western blot showed 25 mmol/L high glucose can obviously enhance the odonto/osteogenic capacity of SCAPs. Moreover, the NF-κB pathway was activated in 25mmol/L glucose-treated SCAPs and the odonto/osteogenic differentiation was inhibited following the inhibition of NF-κB signaling pathway. Conclusions. High glucose can enhance the odonto/osteogenic capacity of SCAPs via NF-κB pathway.
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Zhang, Wei, Yuanbo Wang, Song Jin, Hui Xin, and Changxin Wang. "Overexpression of Guanine Nucleotide Binding Protein, Alpha Stimulating (GNAS) Regulates Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells in High Glucose Environment by Regulating ERK/P38 Signaling Pathway." Journal of Biomaterials and Tissue Engineering 10, no. 2 (February 1, 2020): 259–64. http://dx.doi.org/10.1166/jbt.2020.2300.
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Bone marrow mesenchymal stem cells (BMSCs) can treat osteoporosis. Whether GNAS affects BMSCs osteogenic differentiation under high glucose condition is unknown. Rat BMSCs were isolated and randomly divided into control group, high glucose group and GNAS group. The BMSCs were transfected with GNAS plasmid in high glucose environment followed by analysis of GNAS expression by Real time PCR and Western blot, BMSCs proliferation by MTT assay, Caspase 3 activity, ALP activity, formation of calcified nodules by alizarin red staining, OC and BMP-2 expression by Real time PCR and expression of ERK/P38 signaling pathway protein by Western blot. In high glucose environment, GNAS expression was significantly decreased, cell proliferation was inhibited, Caspase 3 activity was increased, along with decreased ALP activity, calcified nodules formation and expression of OC, BMP-2, p-ERK1/2 and p-P38 (P < 0.05). GNAS plasmid transfected into high glucose environment BMSCs can significantly promote GNAS expression and cell proliferation, decrease Caspase 3 activity, increase p-ERK1/2 and p-P38 expression, ALP activity and calcified nodules formation as well as increase OC and BMP-2 expression (P < 0.05). GNAS1 expression is decreased in BMSCs cells in a high glucose environment. Overexpression of GNAS1 can inhibit the apoptosis of BMSCs by regulating the ERK/P38 signaling pathway, promote its proliferation and differentiation into osteogenic direction.
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Liu, Lin, Kun Liu, Yanzhe Yan, Zhuangzhuang Chu, Yi Tang, and Chunbo Tang. "Two Transcripts of FBXO5 Promote Migration and Osteogenic Differentiation of Human Periodontal Ligament Mesenchymal Stem Cells." BioMed Research International 2018 (2018): 1–12. http://dx.doi.org/10.1155/2018/7849294.
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Objectives. Enhanced migration and osteogenic differentiation of mesenchymal stem cells (MSCs) are beneficial for MSC-mediated periodontal tissue regeneration, a promising method for periodontitis treatment. FBXO5, a member of the F-box protein family, is involved in the osteogenic differentiation of MSCs. Here, we investigated the effect of FBXO5 on human periodontal ligament stem cells (hPDLSCs). Materials and Methods. hPDLSCs were isolated from periodontal ligament tissue. Lentivirus FBXO5 shRNA was used to silence FBXO5 expression. Two transcripts of FBXO5 were overexpressed and transduced into hPDLSCs via retroviral infection. Migration and osteogenic differentiation of hPDLSCs were evaluated using the scratch migration assay, alkaline phosphatase (ALP) activity, ALP staining, alizarin red staining, western blotting, and real-time polymerase chain reaction. Results. The expression of FBXO5 was upregulated after osteogenic induction in hPDLSCs. FBXO5 knockdown attenuated migration, inhibited ALP activity and mineralization, and decreased RUNX2, OSX, and OCN expression, while the overexpression of two transcript isoforms significantly accelerated migration, enhanced ALP activity and mineralization, and increased RUNX2, OSX, and OCN expression in hPDLSCs. Conclusions. Both isoforms of FBXO5 promoted the migration and osteogenic differentiation potential of hPDLSCs, which identified a potential target for improving periodontal tissue regeneration.
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Zhang, Hui, Jie Song, and Xianjie Zhou. "Long Noncoding RNA P53 Upregulated Regulator of P53 Levels Promotes Osteogenic Differentiation in Osteoporosis Progression Through Sponging miR-135a-5p." Journal of Biomaterials and Tissue Engineering 12, no. 10 (October 1, 2022): 2085–91. http://dx.doi.org/10.1166/jbt.2022.3125.
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This study aimed to explore the expression of long noncoding RNA p53 upregulated regulator of P53 levels (lncRNA PURPL) and microRNA (miR)-135a-5p in osteoporosis and their role in osteogenic differentiation. The relationship between lncRNA PURPL and miR-135a-5p was confirmed by Star-Base and luciferase reporter assay. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay was used to detect lncRNA PURPL and miR-135a-5p expression. RT-qPCR and western blot analysis were used to measure osteogenic markers expression. Alkaline phosphatase (ALP) activity was also determined. Results indicated that lncRNA PURPL binds to miR-135a-5p. lncRNA PURPL expression was decreased and miR-135a-5p expression was increased in patients with osteoporosis. In the process of osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs), the expression levels of osteoblast markers including RUNX family transcription factor 2 (Runx2), ALP and Osterix, and ALP activity were significantly increased. Besides, lncRNA PURPL was improved, while miR-135a-5p was down-regulated during the osteogenic differentiation of hBMSCs. Moreover, lncRNA PURPL-siRNA significantly decreased the expression of ALP, Runx2 and Osterix, and reduced ALP activity in hBMSCs subjected to osteogenic induction, while all of these effects were reversed by miR-135a-5p inhibitor. In conclusion, lncRNA PURPL/miR-135a-5p may be a new axis for osteoporosis treatment.
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Huang, Z., X. Huang, Y. Huang, Z. Li, Q. Huang, and T. Li. "POS0044 T315 SUPPRESSES OSTEOGENIC DIFFERENTIATION IN SAOS-2 CELLS BY INHIBITING PHOSPHORYLATION OF AKT." Annals of the Rheumatic Diseases 80, Suppl 1 (May 19, 2021): 227.1–227. http://dx.doi.org/10.1136/annrheumdis-2021-eular.2428.
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Background:New bone formation is common in the late stage of various inflammatory arthritis, while osteoblasts play a vital role in this process. Activation of PI3K/ Akt pathway promotes the differentiation and enhances the function of osteoblasts [1]. T315 is a novel small molecule drug, which may induce apoptosis and suppress the expression of cellular markers of chronic lymphocytic leukemia cells by disrupting PI3K/ Akt pathway [2]. However, the lack of study focuses on the influence of T31T on the other cells, except tumor cell lines.Objectives:We aimed to assess the effect of T315 on human osteoblast-like Saos-2 cells, while its potential mechanism in PI3K/ Akt pathway was evaluated as well.Methods:(1) Saos-2 was stimulated with an osteogenic reagent which contained L-ascorbic acid, β-glycerophosphoric acid, and dexamethasone. The concentration of T315 was adjusted to 0μg/ml, 1μg/ml, and 2μg/ml in the culture medium. (2) Alizarin red stain and alkaline phosphatase (ALP) stain were performed at d0, d7, d14, and d21 after being treated with T315. (3) Cellular protein was extracted at d0, d3, and d6 after being treated with T315, then ALP activity was tested based on a recommendation from the manufacturer of the kit. (4) Collagen type 1α2 Chain (Col1α2) and osteocalcin (OCN), two osteogenic markers, were measured through western blot, with glyceraldehyde 3-phosphate dehydrogenase (GAPDH) as an endogenous control. (5) Phospho-phosphoinositide 3-kinase (pPI3K), phospho-protein kinase B (pAkt), and runt-related transcription factor 2 (Runx2) was tested through western blot as well. GAPDH or protein kinase (Akt) was chosen as an internal reference as appropriate. (6) Analysis of variance with the least significant difference was used to analyze the data. A P<0.05 was considered statistically significant.Results:(1) The higher concentration of T315 related to the less relative mineralized area and the positive area of ALP, while longer incubation time with T315 decreased these regions as well (Figure 1A-C). (2) T315 reduced the activity of ALP accordingly (Figure 1D). (3) T315 suppressed the protein expression of Col1α2 and OCN in a dose-dependent and time-dependent manner (Figure 1E, F). (4) T315 did not alter pPI3K, but it inhibited the phosphorylation of Akt (Figure 1G, H). (5) Runx2 was reduced because of the greater dose or longer incubation time with T315 (Figure 1).Conclusion:T315 inhibits the differentiation of osteoblasts through inhibiting the phosphorylation of Akt. Surprisingly, pPI3K seldom changes in this process, so its detail mechanism should be investigated in further.References:[1]Exp Biol Med (Maywood) 2020;245(6):552-561.[2]Blood 2015;125(2):284-295.Figure 1.Effect of T315 on Saos-2 cells and PI3K/Akt pathway. (A) Alizarin red stain and ALP stain at d21. (B) Relative mineralized area in Alizarin red stain. (C) Positive area in ALP stain. (D) ALP activity. Western blot analysis and its bands at d6 for Col1α2 (E), OCN (F), pPI3K (G), pAkt (H), and Runx2 (I). Results were normalized by GAPDH or Akt. ALP: Alkaline phosphatase; Col1α2: Collagen type 1α2 Chain; GAPDH: Glyceraldehyde 3-phosphate dehydrogenase; OCN: Osteocalcin; pPI3K: Phospho-phosphoinositide 3-kinase; pAkt: Phospho-protein kinase B; Akt: Protein kinase; Runx2: Runt-related transcription factor 2. a Compared with d3 in the same concentration of T315, P<0.05. b Compared with d0 in the same concentration of T315, P<0.05. c Compared with 1μg/ml T315 in the same incubation time, P<0.05. d Compared with 2μg/ml T315 in the same incubation time, P<0.05.Disclosure of Interests:None declared.
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Li, Haijian, Jincheng Jincheng, Lin Wang, Yiting Fu, and Chunzhen Zhao. "The Temporal Profile of OPG Expression and Regulatory Role on Ischemic Brain Injury in Rats After MCAo." Medical Research 3, no. 1 (March 31, 2021): 1–5. http://dx.doi.org/10.6913/mrhk.202103_3(1).0001.
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Objective To investigate the temporal profile of osteoprotegerin (OPG) in middle cerebral artery occlusion (MCAo) rats and the serum level of RANKL and ALP in OPG deficient and wild type rats. Methods Rats was anesthetized and subjected to MCAo by transient occlusion of middle cerebral artery occlusion. The serum level of OPG, RANKL and ALP in rats after MCAo was examined by ELISA assay. The protein expression of OPG in ischemic brain was determined by Western blot analysis. Results The level of OPG in the rat serum was significantly increased from 6 h after MCAo and peaked at 12-24 and 72-168 h. The protein expression of OPG was upregulated from 12 h after MCAo and peaked at 72 h. The level of RANKL and ALP was significantly decreased in OPG deficient rats after MCAo. Conclusion OPG/RANKL signaling is associated with brain injury after MCAo, indicating a potential therapeutic target for ischemic stroke.
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Prieto-Sánchez, María T., José E. Blanco-Carnero, María Ruiz-Palacios, Ana Pagán, Antonio J. Ruiz-Alcaraz, and Elvira Larqué. "Increased Alkaline Phosphatase in Cord Blood of Obese Diabetic Mothers Is Associated to Polyunstaurated Fatty Acid Levels." Annals of Nutrition and Metabolism 75, no. 3 (2019): 153–62. http://dx.doi.org/10.1159/000504404.
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Introduction: Recent studies indicate that alkaline phosphatase (ALP) may affect expression and activity of fatty acid (FA) transport proteins in placenta and other tissues. Objective: To evaluate if disturbed FA profile in offspring of gestational diabetes mellitus (GDM) with different maternal pregestational weight could be related to maternal or neonatal ALP. Methods: Prospective observational study of pregnant women recruited in the third trimester (25 controls, 23 lean-GDM, 20 obese-GDM). Fetal ultrasound was performed. At delivery, FAs were analyzed in placenta, maternal, and venous cord blood. Western blotting analysis of lipid carriers was performed in placenta. Results: Newborns from obese-GDM tended to higher birthweight (p = 0.059) than those from both lean-GDM and controls. ALP in maternal blood tended to be lower in GDM (p = 0.170) while increased significantly in cord blood of obese-GDM with respect to controls (p = 0.039). Saturated FA percentages in cord blood were significantly higher (p < 0.000), while polyunsaturated FA (PUFA) percentages were lower (p = 0.003) in both GDM, which could be due to a lower expression of major family domain 2a receptor (MFSD2a) in the placenta. Plasma ALP in the offspring of obese-GDM was inversely associated to cord essential PUFAs (β = –6.18, p = 0.005) and to placental MFSD2a (β = –38.46, p = 0.014). Conclusions: Cord PUFA and placental MFSD2a are decreased in both lean and obese-GDM pregnancies. Higher ALP in cord blood of obese-GDM could play a role in the FA levels in these pregnancies
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Liu, Xinwei, and Yue Zhou. "Downregulation of lncRNA ANRIL Inhibits Osteogenic Differentiation of Periodontal Ligament Cells via Sponging miR-7 through NF-κB Pathway." Analytical Cellular Pathology 2021 (November 24, 2021): 1–11. http://dx.doi.org/10.1155/2021/7890674.
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Background. Long noncoding RNAs (lncRNAs) are dysregulated in periodontitis development and involved in osteogenesis. The current study was aimed at investigating the function of lncRNA ANRIL in periodontal ligament cells (PDLCs) and potential molecular mechanisms. Methods. Firstly, the level of ANRIL was tested by qPCR. Then, PDLCs were treated with a mineralizing solution to induce osteogenic differentiation. ALP activity was measured, and protein levels of BMP2, Osterix, and OCN were measured by Western blot. A target of ANRIL was verified using dual-luciferase reporter assay. miR-7 level was measured by qPCR, and the signals of the NF-κB pathway were tested by Western blot. Results. ANRIL expression was downregulated in PDL tissues. Next, ALP activity and protein levels of BMP2, Osterix, and OCN were increased to show that PDLCs were differentiated. ANRIL level was increased in differential PDLCs, in which knockdown inhibited osteogenic differentiation. Then, miR-7 was found as a target of ANRIL. The miR-7 level was upregulated in PDL tissues and reduced in differential PDLCs. Inhibition of miR-7 suppressed ALP activity and BMP2, Osterix, and OCN expression. Moreover, inhibition of miR-7 reversed the effects on the osteogenic differentiation induced by knockdown of ANRIL. Besides, the levels of p-P65 and p-IκBα were elevated by ANRIL downregulation and were rescued by suppressing miR-7. Conclusions. Knockdown of ANRIL inhibited osteogenic differentiation via sponging miR-7 through the NF-κB pathway, suggesting that ANRIL might be a therapeutic target for periodontitis.
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Pang, Xin-Gang, Yu Cong, Ni-Rong Bao, Yong-Gang Li, and Jian-Ning Zhao. "Quercetin Stimulates Bone Marrow Mesenchymal Stem Cell Differentiation through an Estrogen Receptor-Mediated Pathway." BioMed Research International 2018 (2018): 1–11. http://dx.doi.org/10.1155/2018/4178021.
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Objectives. The present study aimed to investigate the overall effect of quercetin on mouse bone marrow mesenchymal stem cell (BMSC) proliferation and osteogenic differentiation in vitro. Materials and Methods. BMSCs were treated with different concentrations of quercetin for 6 days. The effects of quercetin on cell proliferation were assessed at predetermined times using Cell Counting Kit-8 (CCK-8) assay. The cells were then treated with quercetin, estrogen, or an estrogen receptor (ER) antagonist (which was also administered in the presence of quercetin or estrogen) for 7 or 21 days. The effects of quercetin on BMSC osteogenic differentiation were analyzed by an alkaline phosphatase (ALP) assay kit, Alizarin Red S staining (ARS), quantitative real-time PCR (qPCR), and western blotting. Results. The CCK-8 and ALP assays and ARS staining showed that quercetin significantly enhanced BMSC proliferation, ALP activity, and extracellular matrix production and mineralization, respectively. The qPCR results indicated that quercetin promoted osterix (OSX), runt-related transcription factor 2 (RUNX2), and osteopontin (OPN) transcription in the presence of osteoinduction medium, and the western blotting results indicated that quercetin enhanced bone morphogenetic protein 2 (BMP2), Smad1, Smad4, RUNX2, OSX, and OPN expression and Smad1 phosphorylation. Treatment with the ER inhibitor ICI182780 blocked the effects of quercetin. Conclusions. Our data demonstrated that quercetin promotes BMSC proliferation and osteogenic differentiation. Quercetin enhances BMP signaling pathway activation and upregulates the expression of downstream genes, such as OSX, RUNX2, and OPN, via the ER.
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Zeng, Kangrui, Qiongyi Kang, Yutong Li, Weiping Li, Qing Cheng, and Wenwei Xia. "EVL Promotes Osteo-/Odontogenic Differentiation of Dental Pulp Stem Cells via Activating JNK Signaling Pathway." Stem Cells International 2023 (January 12, 2023): 1–14. http://dx.doi.org/10.1155/2023/7585111.
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Objective. Human dental pulp stem cells (hDPSCs) were recognized as a suitable and promising source of stem cells in dental pulp regeneration. However, the mechanism by which hDPSCs differentiation into osteo-/odontogenic lineage remains unclear. Ena/VASP-like protein (EVL) has been found to be involved in diverse biological processes. In this study, we explored the role and underlying mechanism of EVL in osteo-/odontogenic differentiation of hDPSCs. Methods. Expression of EVL was detected in hDPSCs by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot (WB) analyses during osteo-/odontogenic differentiation. The function of EVL in osteo-/odontogenic differentiation and involvement of MAPK signaling pathways were evaluated by alkaline phosphatase (ALP) staining and activity, alizarin red staining (ARS), and qRT-PCR and western blot analyses. Results. The expression of EVL was upregulated during osteo-/odontogenic differentiation of hDPSCs. Overexpression of EVL significantly increased osteo-/odontogenic capacity of hDPSCs, which was reflected in increased alkaline phosphatase (ALP) staining, ALP activity, mineralized nodule formation, and the expressions of genes related to osteo-/odontogenic differentiation, while downregulation of EVL inhibited it. In addition, EVL activated the JNK pathway and phosphorylation of p38 MAPK during differentiation procedure of hDPSCs. The EVL-enhanced differentiation of DPSCs was suppressed by blocking the JNK pathway, rather than the p38 MAPK pathway. Conclusion. EVL promotes the osteo-/odontogenic differentiation of hDPSCs by activating the JNK pathway, providing a future target for osteo-/odontogenic differentiation and dental pulp regeneration.
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Nan, Hai, and Yun Zhang. "Interleukin-6 Promotes Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells by Activating Wnt Signaling Pathway." Journal of Biomaterials and Tissue Engineering 9, no. 9 (September 1, 2019): 1261–65. http://dx.doi.org/10.1166/jbt.2019.2121.
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Bone marrow mesenchymal stem cells (BMSCs) can differentiate into adipocytes, osteoblasts. Osteoporosis is closely related to BMSCs osteogenic differentiation, and IL-6 is closely related to osteoporosis. This study assessed the effect of IL-6 on BMSCs osteogenic differentiation. Rat BMSCs were cultured and osteogenic induction of BMSCs was performed in the presence of different concentrations (0, 10, 100 ng/ml) of IL-6 followed by analysis of IL-6 level by ELISA, ALP activity by the instructions of the alkaline phosphatase (ALP) detection kit, IL-6, Runx2 and OCN mRNA level, and level of β-catenin by Western blot. Compared with 0 d, IL-6 protein content and IL-6 mRNA expression in cell culture medium was increased significantly on day 7 d, 14 d and 21 d. Compared with 0 ng/ml group, 10, 100 ng/ml IL-6 group showed significantly increased ALP activity and Runx2 and OCN mRNA level in a dose-response relationship. β-catenin was increased significantly in 100 ng/ml IL-6 group. No difference of ALP activity and the expression of osteogenic differentiationmarkers was found between blocking group and control group, which was significantly lower than those in experimental group. IL-6 can promote BMSCs osteogenic differentiation through Wnt signaling.
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Chou, LongHang, YaTing Chang, KaiWen Lan, Meng Liu, YuKun Lu, XiaoLei Li, PeiRu Li, and Yue Xu. "CCK regulates osteogenic differentiation through TNFα/NF-κB in peri-implantitis." Journal of International Medical Research 50, no. 12 (December 2022): 030006052211413. http://dx.doi.org/10.1177/03000605221141312.
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Objective Peri-implantitis is characterized by peri-implant mucositis and alveolar bone resorption. This study investigated cholecystokinin ( CCK) expression and the mechanism underlying its involvement in peri-implantitis. Methods mRNA sequencing was performed using the Gene Expression Omnibus database GSE106090. Human bone marrow mesenchymal stem cells (hBMSCs) were pretreated with various concentrations of CCK (0, 10, 30, or 100 nM) for 1 hour before induction in osteogenic differentiation medium for 2 weeks. Alkaline phosphatase (ALP) activity was determined, and the cells were stained with alizarin red. The expression levels of TNFα and the osteogenic markers ALP, RUNX2, and OCN were measured using quantitative real-time PCR. TNFα, phosphorylated P65, and total P65 levels were determined by western blot. Results Compared with healthy individuals, 262 and 215 genes were up- and down-regulated, respectively, in the periodontal tissues of patients with peri-implantitis. CCK expression was significantly upregulated in patients with peri-implantitis. CCK reduced ALP activity, osteogenic differentiation, and levels of the osteogenic markers ALP, RUNX2, and OCN. Moreover, CCK promoted levels of TNFα and phosphorylated P65, which is a marker of activation for the NF-κB inflammatory pathway. Conclusions CCK regulates osteogenic differentiation through the TNFα/NF-κB axis in peri-implantitis.
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Cui, Yi, Sheng Lu, Hongbo Tan, Jun Li, Min Zhu, and Yongqing Xu. "Silencing of Long Non-Coding RNA NONHSAT009968 Ameliorates the Staphylococcal Protein A-Inhibited Osteogenic Differentiation in Human Bone Mesenchymal Stem Cells." Cellular Physiology and Biochemistry 39, no. 4 (2016): 1347–59. http://dx.doi.org/10.1159/000447839.
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Background/Aims: Osteomyelitis is defined as an inflammation of the bones and bone marrow. The inflammatory microenvironment attenuates the osteogenic differentiation capacity of stem cells and inhibits osteoblast-mediated bone formation, leading to net bone loss. However, the whole expression profile, function and side effect of long non-coding RNAs (lncRNAs) on osteogenic differentiation of stem cells in an inflammatory microenvironment of osteomyelitis are not known. Methods: In the present study, human bone mesenchymal stem cells (hBMSCs) were treated with different concentrations of Staphylococcal protein A (SpA) to trigger an inflammatory microenvironment in vitro to partly duplicate the inflammatory microenvironment of osteomyelitis, which was confirmed using ELISA for detecting the inflammatory cytokines. The complete expression profiles of lncRNAs and mRNA during osteogenic differentiation of hBMSCs in an inflammatory microenvironment triggered by SpA were analyzed using a lncRNA microarray. LncRNA expression levels were verified by quantitative reverse transcription PCR analysis (qRT-PCR). The expression of NONHSAT009968 in hB-MSCs was silenced by infection with lentivirus expressing NONHSAT009968-shRNA. The expression of Runx2, OCN, OPN, COL1A1, and alkaline phosphatase (ALP) activity was detected by western blot. Alizarin red staining and ALP activity detection were carried out. Results: The results of ELISA showed that SpA treatment induced secretion of inflammatory cytokines IL-1A, IL-6, and TNFA. The results of alizarin red staining and ALP detection showed that SpA treatment suppressed the osteogenic differentiation of hBMSCs. A total of 2033 lncRNAs were found with aberrant expression in SpA-treated hBMSCs compared to controls. Among these lncRNAs, 641 were down-regulated and 1392 were up-regulated. Based on the results of qRT-PCR, lncRNA NONHSAT009968 was chosen for further investigation. The results of alizarin red staining, ALP activity detection, and western blot detection of Runx2, OCN, OPN, COL1A1, and ALP indicated that NONHSAT009968 silencing ameliorates SpA-inhibited osteogenic differentiation in hBMSCs. Conclusion: Our present study provides a basis for future analyses of the role of lncRNAs in osteoblastic differentiation in an inflammatory environment triggered by SpA, and lncRNA NONHSAT009968 might be a new target for promoting osteoblast formation.
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Wang, Wen-Ling, Shi-Yuan Sheu, Yueh-Sheng Chen, Shung-Te Kao, Yuan-Tsung Fu, Tzong-Fu Kuo, Kuo-Yu Chen, and Chun-Hsu Yao. "Evaluating the Bone Tissue Regeneration Capability of the Chinese Herbal DecoctionDanggui Buxue Tangfrom a Molecular Biology Perspective." BioMed Research International 2014 (2014): 1–10. http://dx.doi.org/10.1155/2014/853234.
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Large bone defects are a considerable challenge to reconstructive surgeons. Numerous traditional Chinese herbal medicines have been used to repair and regenerate bone tissue. This study investigated the bone regeneration potential ofDanggui Buxue Tang(DBT), a Chinese herbal decoction prepared fromRadix Astragali(RA) andRadix Angelicae Sinensis(RAS), from a molecular biology perspective. The optimal ratio ofRAandRASused inDBTfor osteoblast culture was obtained by colorimetric and alkaline phosphatase (ALP) activity assays. Moreover, the optimal concentration ofDBTfor bone cell culture was also determined by colorimetric, ALP activity, nodule formation, Western blotting, wound-healing, and tartrate-resistant acid phosphatase activity assays. Consequently, the most appropriate weight ratio ofRAtoRASfor the proliferation and differentiation of osteoblasts was 5 : 1. Moreover, the most effective concentration ofDBTwas 1,000 μg/mL, which significantly increased the number of osteoblasts, intracellular ALP levels, and nodule numbers, while inhibiting osteoclast activity. Additionally, 1,000 μg/mL ofDBTwas able to stimulate p-ERK and p-JNK signal pathway. Therefore,DBTis highly promising for use in accelerating fracture healing in the middle or late healing periods.
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Fang, Ming, Xingwu Wang, Yongli Wei, Wuliang Yu, and Jianmeng Lu. "Tanshinone IIA Regulates Osteoblast Differentiation and Promotes Fracture Healing via Mammalian Target of Rapamycin Complex 1 Signaling Pathway." Journal of Biomaterials and Tissue Engineering 11, no. 9 (September 1, 2021): 1737–43. http://dx.doi.org/10.1166/jbt.2021.2587.
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This study assessed the effect and potential molecular mechanism of tanshinone IIA on fracture healing. Mice model with fracture were established. Digital radiographic photographic system was used to detect callus formation after treatment with tanshinone II A (Tan IIA) and alkaline phosphatase (ALP) staining analyzed ALP activity. Osteoblast proliferation was also measured. Western blot and Quantitative real-time PCR (qRT-PCR) measured osteogenic markers expression. Compared with control group, Tan IIA treatment could increase callus formation, stimulate osteoblast proliferation, osteogenic proteins and genes expression, and activate mTORC1 signaling pathway. However, Tan IIA’s effects were significantly inhibited after rapamycin treatment. Tan IIA regulates osteoblast differentiation by mTORC1 signaling and promotes intramembranous ossification in the process of callus formation, which accelerates bone healing.
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Shen, Mei-jie, Ge-ge Wang, Yu-zhen Wang, Jing Xie, and Xi Ding. "Nell-1 Enhances Osteogenic Differentiation of Pre-Osteoblasts on Titanium Surfaces via the MAPK-ERK Signaling Pathway." Cellular Physiology and Biochemistry 50, no. 4 (2018): 1522–34. http://dx.doi.org/10.1159/000494651.
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Background/Aims: This study aimed to investigate the effect of Nell-1 on the osteogenic behaviors of pre-osteoblasts on titanium (Ti) surfaces and to identify the underlying signaling pathway. Methods: Nell-1 at different concentrations was added to culture medium to stimulate MC3T3-E1 subclone 14 on Ti surfaces. A CCK-8 colorimetric assay was used to detect cell proliferation. Alkaline phosphatase activity (ALP) assay and enzyme-linked immunosorbent assay (ELISA) were used to evaluate ALP activity and the osteocalcin (OCN) secretion, respectively. Indicators of osteoblastic differentiation were assessed using real-time polymerase chain reaction analysis (RT-PCR). Western blot (WB) assay was used to analyze the expression changes of the osteogenic proteins and the mitogen-activated protein kinase (MAPK) pathway. Results: Nell-1 significantly increased the osteogenic gene and protein expression levels of ALP, OCN, Runx2, osteoprotegerin (OPG), collagen type I (Col-I), and Osterix (Osx) in pre-osteoblasts on Ti surfaces. The optimal concentration of Nell-1 was 100 ng/ ml. In addition, Nell-1 activated ERK and JNK, but not P38, in MC3T3-E1 cells on the Ti surface. Except for ALP and Col-I, the promotive effects of Nell-1 on the expression of osteogenic markers were suppressed by ERK inhibitor U0126. Conclusion: Certain concentrations of Nell-1 can promote the osteogenic differentiation of pre-osteoblasts on Ti surfaces by activating the MAPK/ERK signaling pathway.
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Li, Zhengqiang, Junfa Zheng, Di Wan, and Xiaoqin Yang. "Uniaxial Static Strain Promotes Osteoblast Proliferation and Bone Matrix Formation in Distraction Osteogenesis In Vitro." BioMed Research International 2020 (August 13, 2020): 1–12. http://dx.doi.org/10.1155/2020/3906426.
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Objective. We aimed at investigating the effects of uniaxial static strain on osteoblasts in distraction osteogenesis (DO). Methods. To simulate the mechanical stimulation of osteoblasts during DO, 10% uniaxial static strain was applied to osteoblasts using a homemade multiunit cell stretching and compressing device. Before and after applying strain stimulation, the morphological changes of osteoblasts were observed by inverted phase-contrast microscopy, Coomassie blue staining, and immunofluorescence. Alkaline phosphatase (ALP) activity, mRNA levels (proliferating cell nuclear antigen [PCNA], ALP, Runx2, osteocalcin [OCN], collagen type I, hypoxia-inducible factor- [HIF-] 1α, and vascular endothelial growth factor [VEGF]), and protein levels (Runx2, OCN, collagen type I, HIF-1α, and VEGF) were evaluated by using ALP kit, real-time quantitative reverse transcription-polymerase chain reaction, western blot, and enzyme-linked immunosorbent assay. Results. After the mechanical stimulation, the cytoskeleton microfilaments were rearranged, and the cell growth direction of the osteoblasts became ordered, with their direction being at an angle of about 45° from the direction of strain. The proliferation of osteoblasts and the expression levels of mRNA and protein of ALP, Runx2, OCN, collagen type I, HIF-1α, and VEGF were significantly higher than in the nonstretch control groups. Conclusion. Our homemade device can exert uniaxial static strain and promote the proliferation of osteoblasts and bone matrix formation. It can be used to simulate the mechanical stimulation of osteoblasts during DO.
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Yang, Nan, Xiao Zhang, Lingfeng Li, Tongtong Xu, Meihui Li, Qi Zhao, Jinling Yu, Jue Wang, and Zhihui Liu. "Ginsenoside Rc Promotes Bone Formation in Ovariectomy-Induced Osteoporosis In Vivo and Osteogenic Differentiation In Vitro." International Journal of Molecular Sciences 23, no. 11 (May 31, 2022): 6187. http://dx.doi.org/10.3390/ijms23116187.
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Ginsenoside Rc is one of the active components used in traditional Chinese medicine. We aim to explore how ginsenoside Rc can be used in the treatment of osteoporosis. Micro-CT demonstrated that the treatment of ovariectomized (OVX) mice with ginsenoside Rc significantly inhibited the decrease in bone mineral density, bone volumetric fraction, and trabecular number, and the increase in trabecular separation. Histological staining, qRT-PCR, and Western blot demonstrated that ginsenoside Rc enhances the microstructure of trabecular bone, and promotes the expression of bone formation-related genes. Alkaline phosphatase (ALP) staining, Alizarin Red staining, qRT-PCR, and Western blotting demonstrated that ginsenoside Rc dose-dependently promoted the osteogenic differentiation of MC3T3-E1 cells. A ginsenoside Rc-induced increase in the expression of β-catenin, p-GSK-3β, collagen-1, ALP, and RUNX-2 family transcription factor-2 was significantly attenuated upon 10 μM XAV-939 treatment, while the decrease in the expression of GSK-3β and p-β-catenin was significantly enhanced. Ginsenoside Rc promotes bone formation in ovariectomy-induced osteoporosis in vivo and promotes osteogenic differentiation in vitro via the Wnt/β-catenin signaling pathway.
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Yin, Heng, Jianwei Wang, Mao Wu, Yong Ma, Shanfu Wang, and Qiuju Su. "Preventive Effects of Evodiamine on Dexamethasone-Induced Osteoporosis in Zebrafish." BioMed Research International 2019 (January 22, 2019): 1–6. http://dx.doi.org/10.1155/2019/5859641.
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The aim of this study was to investigate the effect of evodiamine (EV) on dexamethasone-induced osteoporosis in zebrafish. Zebrafish larvae were exposed to different concentrations of dexamethasone to obtain the osteoporosis in zebrafish. Calcium, phosphorus, and alizarin red staining determination were performed to evaluate the effects of EV on bone mineralization. Alkaline phosphatase (ALP), hydroxyproline (HP), and tartrate resistant acid phosphatase (TRAP) were also measured by commercial kits. The expression of MMP3-OPN-MAPK pathway in zebrafish was measured by Western blot. RT-PCR was used to determine mRNA levels of MMP3, OPN, and MAPK. EV could significantly increase the content of calcium and phosphorus. The results of alizarin red staining showed that EV could significantly increase the calcium sink of horse fish, increasing the area of bone formation. EV could increase the content of hydroxyproline in zebrafish. EV also increased ALP and TRAP in zebrafish. Western blot and RT-PCR results showed that EV restored the MMP3-OPN-MAPK pathway in zebrafish. In conclusion, we found that EV can alleviate dexamethasone-induced osteoporosis in zebrafish. The mechanism is related to activating MMP3-OPN-MAPK pathway and then activating bone remodeling.
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Kunimatsu, Ryo, Tomoka Hiraki, Kodai Rikitake, Kengo Nakajima, Nurul Aisyah Rizky Putranti, Takaharu Abe, Kazuyo Ando, Ayaka Nakatani, Shuzo Sakata, and Kotaro Tanimoto. "Effects of Human Deciduous Dental Pulp-Derived Mesenchymal Stem Cell-Derived Conditioned Medium on the Metabolism of HUVECs, Osteoblasts, and BMSCs." Cells 11, no. 20 (October 14, 2022): 3222. http://dx.doi.org/10.3390/cells11203222.
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In this study, we assessed the effects of human deciduous dental pulp-derived mesenchymal stem cell-derived conditioned medium (SHED-CM) on the properties of various cell types. The effects of vascular endothelial growth factor (VEGF) in SHED-CM on the luminal architecture, proliferative ability, and angiogenic potential of human umbilical vein endothelial cells (HUVECs) were determined. We also investigated the effects of SHED-CM on the proliferation of human-bone-marrow mesenchymal stem cells (hBMSCs) and mouse calvarial osteoblastic cells (MC3T3-E1) as well as the expression of ALP, OCN, and RUNX2. The protein levels of ALP were examined using Western blot analysis. VEGF blockade in SHED-CM suppressed the proliferative ability and angiogenic potential of HUVECs, indicating that VEGF in SHED-CM contributes to angiogenesis. The culturing of hBMSCs and MC3T3-E1 cells with SHED-CM accelerated cell growth and enhanced mRNA expression of bone differentiation markers. The addition of SHED-CM enhanced ALP protein expression in hBMSCs and MT3T3-E1 cells compared with that of the 0% FBS group. Furthermore, SHED-CM promoted the metabolism of HUVECs, MC3T3-E1 cells, and hBMSCs. These findings indicate the potential benefits of SHED-CM in bone tissue regeneration.
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Zhou, Yonghuan, Guotang Lan, Yan Zhou, Tianhao Qu, and Qing Xiong. "Effect of Yes Associated Protein on Osteogenic/Adipogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Under Oxidative Stress." Journal of Biomaterials and Tissue Engineering 11, no. 8 (August 1, 2021): 1636–42. http://dx.doi.org/10.1166/jbt.2021.2724.
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Oxidative stress affects bone marrow mesenchymal stem cells (BMSCs). YAP is an effector in Hippo signaling, but its’ role in BMSCs osteogenesis/adipogenesis under oxidative stress has not been reported. Mice BMSCs were isolated and assigned into 3 groups, normal control group; oxidative stress group; and YAP group (transfected with YAP plasmid) followed by analysis of YAP expression by Real time PCR. After 14 days of osteogenesis or adipogenic induction, RUNX2, OPN, FABP4 and PPARγ2 mRNA level was measured along with ROS and SOD activities, ALP activity and Wnt5 expression by western blot. Under oxidative stress, YAP expression significantly decreased, RUNX2 and OPN mRNA expression decreased, ROS expression increased, SOD activity decreased, FABP4 and PPARγ2 protein expression increased, ALP activity and Wnt5 expression decreased (P <0.05). YAP plasmid transfection could significantly up-regulate YAP, RUNX2 and OPN mRNA level, decrease ROS, increase SOD and ALP activity, reduce FABP4 and PPARγ2 mRNA expression and increase Wnt5 expression (P <0.05). YAP level in BMSCs is decreased under oxidative stress. Up-regulating YAP can improve the redox balance, promote BMSCs osteogenic differentiation under oxidative stress and inhibit their differentiation to adipocytes.
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Wu, Bin, Fenghua Bai, Jianping Lin, Guangji Wang, Wentao Cai, and Mingxia Lin. "Effect of Phosphatase and Tensin Homolog 12 on Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells During Aging by Regulating Transforming Growth Factor Beta/Smad Signaling Pathway." Journal of Biomaterials and Tissue Engineering 11, no. 8 (August 1, 2021): 1630–35. http://dx.doi.org/10.1166/jbt.2021.2723.
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Aging affects bone marrow mesenchymal stem cells (BMSC) differentiation. PTEN12 regulates cell proliferation and apoptosis. However, whether PTEN12 affects BMSCs osteogenic differentiation during aging is unknown. Two BMSCs derived from Zempster24−/− (senescence) and Zempster24+/+ (normal) mice were cultured in vitro. Real-time PCR analysis was used to analyze PTEN12 expression. PTEN12 siRNA was transfected into senescent Zempster24-/-BMSCs and after 14 days of osteogenic induction, cell proliferation was analyzed by MTT method along with measuring expression of osteocalcin, type I collagen, RUNX2 and OPN by Real time PCR, ALP activity, and TGFβ/smad signaling protein expression by Western blot. Compared to normal BMSCs, PTEN12 level in aging BMSCs was significantly elevated, osteocalcin, type I collagen, RUNX2 and OPN mRNA level was decreased along with reduced ALP activity and TGFβ1 and Smad2 expression (P < 0.05). PTEN12 siRNA transfection into senescent BMSCs significantly down-regulated PTEN12, upregulated osteocalcin, type I collagen, RUNX2 and OPN mRNA, increased ALP activity and TGFβ1 and Smad2 expression (P <0.05). Aging increases PTEN12 level and inhibits BMSCs osteogenic differentiation. Down-regulation of PTEN12 in BMSCs during aging can promote BMSCs osteogenic differentiation by regulating TGFβ/smad signaling pathway.
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Yang, Qing, Cheng Li, Manli Yan, and Chunhua Fang. "Effect of SOX9 on Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Through WNTβ/Catenin Pathway." Journal of Biomaterials and Tissue Engineering 9, no. 10 (December 1, 2019): 1429–34. http://dx.doi.org/10.1166/jbt.2019.2140.
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Bone marrow mesenchymal stem cells (BMSCs) can be differentiated into different types of cells. SOX9 involves in the development and progression of various diseases. Our study aims to assess SOX9's effect on osteogenic differentiation of BMSCs and its related regulatory mechanisms. Rat BMSCs were isolated and randomly divided into control group, SOX9 group and SOX9 siRNA group, which was transfected with pcDNA-SOX9 plasmid or SOX9 siRNA respectively followed by analysis of SOX9 expression by Real time PCR, cell proliferation by MTT assay, Caspase3 and ALP activity, GSK-3β expression and Wntβ/Catenin Signaling pathway protein expression by Western blot, and expression of osteogenic genes Runx2 and BMP-2 by Real time PCR. Transfection of pcDNA-SOX9 plasmid into BMSCs significantly inhibited cell proliferation, promoted Caspase3 activity, decreased ALP activity and downregulated Runx2 and BMP-2, increased GSK-3β expression and decreased Wntβ/Catenin expression protein expression (P< 0.05). SOX9 siRNA transfection significantly promoted cell proliferation, inhibited Caspase3 activity, increased ALP activity and upregulated Runx2 and BMP-2, downregulated GSK-3β and increased Wntβ/Catenin expression. SOX9 regulates BMSCs proliferation and osteogenic differentiation through Wntβ/Catenin signaling pathway.
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Li, Xiaoliang, Guofeng Xia, Hongmei Xin, Chunsheng Tao, Weiwei Lai, and Peifeng Sun. "lncRNA MALAT1 Inhibits Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells by Down-Regulating WNT5A." Journal of Biomaterials and Tissue Engineering 9, no. 11 (November 1, 2019): 1520–27. http://dx.doi.org/10.1166/jbt.2019.2167.
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ncRNA involves in osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). WNT5A participates in the growth and development of osteogenic differentiation. This study aims to investigate whether lncRNA MALAT1 regulates BMSCs osteogenesis through WNT5A. qRT-PCR was done to detect the lncRNA MALAT1 level and osteogenic markers in osteoporosis patients and control groups. The above markers and WNT5A protein levels were detected by Western blot. The degree of osteogenic differentiation was detected by ALP activity assay and ALP staining. The differentiation ability of BMSCs after lncRNA MALAT1 overexpression was detected by ARS staining. The binding site of lncRNA MALAT1 to WNT5A was determined by dual luciferase reporter assay. lncRNA MALAT1 expression was higher in patients with osteoporosis, and decreased significantly with increased osteogenic induction. Overexpression of lncRNA MALAT1 in BMSCS reduced WNT5A level, while interference with lncRNA MALAT1 increased WNT5A levels. In cells with overexpression of lncRNA MALAT1, transfection of si-WNT5A can significantly downregulate the RUNX2, OSX, ALP, OCN, OPN, and COL1A1, thereby inhibiting osteogenic differentiation, interfering with the regulation of WNT signaling pathway and regulating BMSCs osteogenic differentiation. lncRNA MALAT1 and WNT5A can regulate BMSCs osteogenesis, thus accelerating the progression of osteoporosis.
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Gu, Mingyong, and Runquan Zheng. "Apolipoprotein E Inhibits Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells by Inhibiting β-Catenin Expression." Journal of Biomaterials and Tissue Engineering 9, no. 12 (December 1, 2019): 1739–44. http://dx.doi.org/10.1166/jbt.2019.2194.
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Bone marrow mesenchymal stem cells (BMSCs) can differentiate into adipocytes, osteoblasts. Apolipoprotein E (ApoE) is closely related to bone metabolism and its effect on bone marrow mesenchymal stem cells is unclear. Therefore, this study investigated ApoE's effect on BMSCs osteogenic differentiation. BMSCs were isolated from ApoE – and WT mouse and cultured to induce osteogenic induction followed by analysis of expression of osteogenic differentiation marker genes by Real-time PCR, calcium nodules formation by ARS staining, ALP activity and -catenin protein level by Western blot. The number of bone differentiation markers, ALP activity and calcium nodules formation as well as β-catenin protein level in ApoE– group were significantly elevated compared with WT (P < 0 05). After treatment with DKK-1, β-catenin expression was significantly reduced (P < 0 05) without difference between ApoE– + DKK-1 group and WT group (P > 0 05). WT+ DKK1 group showed significantly reduced osteogenic differentiation marker expression, ALP activity and calcium nodule number compared to WT (P < 0 05) without difference between ApoE– + DKK1 group and WT group (P > 0 05). ApoE inhibits BMSCs osteogenic differentiation by inhibiting β-catenin expression.
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Zhang, Shouping, Ying Wang, and Lili Sun. "Progranulin Reverses Decitabine-Induced Adipogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Through Downregulation of NF-κB Signaling Pathway." Journal of Biomaterials and Tissue Engineering 10, no. 1 (January 1, 2020): 87–92. http://dx.doi.org/10.1166/jbt.2020.2217.
Full textAbstract:
Decitabine can induce BMSCs adipogenic differentiation. Progranulin (PGRN) is a chondrogenic factor. However, the effect of Progranulin on the adipogenic differentiation of BMSCs induced by decitabine remains unclear. Rat BMSCs were isolated and divided into control group, Decitabine group, and Decitabine+PGRN group followed by analysis of survival rate of BMSCs cells by MTT assay, Caspase 3 activity, ALP activity, Runx2, OP and PPARγ2 expression by Real time PCR, lipids formation by Oil red O staining and the expression of NF-κB by Western blot. Decitabine treatment can significantly inhibit the proliferation of BMSCs, promote the increase of Caspase 3 activity, decrease ALP activity and the expression of Runx2 and OP, increase PPARγ2 expression, the ability of adipogenesis and NF-κB expression (P < 0005). Progranulin addition significantly promoted BMSCs proliferation, inhibited Caspase 3 activity, increased ALP activity and Runx2, OP expression, decreased PPARγ2 expression, adipogenic capacity and NF-κB expression, compared to Decitabine group (P < 0005). Decitabine inhibits BMSCs proliferation, promotes apoptosis, induces adipogenic differentiation, and inhibits osteogenic differentiation. Progranulin reverses the effect of defercitin on the induction of adipogenic differentiation of BMSCs by down-regulating the NF-κB signaling pathway.
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Wang, Guoliang, Yujuan Dong, and Hua Yan. "Effect of Insulin-Like Growth Factor-1 on Bone Formation of Bone Marrow Mesenchymal Stem Cells Under High Glucose Environment by Regulating Insulin Like Growth Factor Binding Protein 2(IGFBP-2)/p38 Pathway." Journal of Biomaterials and Tissue Engineering 10, no. 8 (August 1, 2020): 1242–47. http://dx.doi.org/10.1166/jbt.2020.2372.
Full textAbstract:
Diabetes easily affects the biological properties of bone marrow mesenchymal stem cells (BMSCs). Insulin-like growth factor-1 (IGF-1) promotes bone healing during osteoporotic fractures. However, IGF-1's effect on BMSCs high glucose is unclear. Rat BMSCs were assigned into control group, high glucose group, and IGF-1 group in which BMSCs were transfected with pc-DNA 3.1-IGF-1 plasmid on the basis of high glucose followed by analysis of IGF-1, RUNX2 and OPN mRNA level by real time PCR, cell proliferation by MTT assay, alkaline phosphatase (ALP) activity, IGFBP-2 level by ELISA, and p38 phosphorylation by Western blot. High glucose group showed significantly decreased IGF-1, RUNX2 and OPN mRNA level, reduced ALP activity, IGFBP-2 expression and p38 phosphorylation compared to control group (P < 0 05). Transfection of IGF-1 plasmid under high-glucose environment significantly upregulated IGF-1, RUNX2 and OPN mRNA, increased ALP activity, IGFBP-2 expression and p38 phosphorylation compared to high-glucose group (P < 0 05). IGF-1 expression is reduced in high glucose environment. Up-regulation of IGF-1 can promote BMSCs osteogenic differentiation through the IGFBP-2/p38 pathway.
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Liu, Yanyi, Zekun Gan, and Fei Hu. "Effect of hydroxyapatite bioceramics on the growth of osteoblasts and HIF-α/VEGF signal axis in partial hypoxia environment in vitro." Technology and Health Care 30 (February 25, 2022): 363–69. http://dx.doi.org/10.3233/thc-thc228033.
Full textAbstract:
BACKGROUND: Hydroxyapatite bioceramic is a kind of bone implant commonly used in oral clinic treatment. In the early stage of tissue repair, cells will suffer hypoxic due to the interruption of blood supply. OBJECTIVE: Studying the expression of osteoblasts in hypoxic environment will help us to understand the expression and response mechanism of osteoblasts at the implantation site of hydroxyapatite in the early stage of hypoxia. METHODS: MG63 osteoblast cell line was used in this study. The cells of normal group were incubated under normal oxygen and hydroxyapatite ceramics condition. The cells of hypoxia group were incubated under hypoxia (37∘C, 8% CO2, 8% O2, 86% N2) and hydroxyapatite ceramics condition. Cell proliferation was measured by CCK8 assay. Apoptosis was measured by flow cytometry. Serum alkaline phosphatase (ALP) activity was measured by ALP kit. Hypoxia inducible factor (HIF-α) and vascular endothelial growth factor (VEGF) were detected by Western blot. RESULTS: Compared to the normal group, the cells of hypoxia group showed a dramatically higher proliferation ability, especially at 48 h (P< 0.05). Due to hypoxia, cell apoptosis was induced, but there is no difference between these two groups. Interestingly, the ALP activity of hypoxia group was higher than that of normal group at 24 h and 48 h (P< 0.05). Mechanically, western blot result showed that the protein level of both HIF-α and VEGF were up-regulated in hypoxia group. CONCLUSIONS: Under hypoxia condition, hydroxyapatite bioceramics can promote the proliferation of MG63 osteoblasts, elevate the activity of alkaline phosphatase and upregulate HIF-α and VEGF expression without effect on apoptosis.
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Ma’arif, Burhan, Iffatul Abada, Anisah Mahardiani, Abdul Hakim, Novia Maulina, Neny Purwitasari, Khoirul Hidayah, and Seow Lay Jing. "A Systematic Review: Comparison of Immunocytochemistry, ELISA, and Western Blot Methods in Alkaline phosphatase Measurement at Genistein-induced Osteoblast Cell." Biomedical and Pharmacology Journal 15, no. 4 (December 20, 2022): 1853–65. http://dx.doi.org/10.13005/bpj/2523.
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Osteoporosis is a bone disorder characterized by the decrease of bone mass along with bone micro-architecture damage and has a risk become a fracture. One of the causes of osteoporosis is estrogen deficiency. Genistein is a phytoestrogen compound in the isoflavone group containing a similar structure compared to 17β-estradiol, thus it can bind to estrogen receptors and produce an estrogenic effect. Genistein induction can stimulate bone formation and promote the increase of alkaline phosphate (ALP) activities in osteoblast cells which can be observed by immunocytochemistry or Enzyme-linked Immunosorbent Assay (ELISA) or Western blot method. Using the PRISMA guideline technique, choose and strategize article searches by reading the title, abstract, and then the whole text of the article. Articles with the keywords "genistein or osteoblast cells or alkaline phosphate or immunocytochemistry or immunofluorescence or ELISA or western blot" were retrieved from databases including Google Scholar, PubMed, Researchgate, and Sciencedirect. 24 relevant research articles were uncovered as a result of this systematic review. Comparison of immunocytochemistry and ELISA methods in order to analyze the activities of ALP in osteoblast induced by genistein includes selectivity, sensitivity, processing time, and cost efficiency parameters. The immunocytochemistry method has a higher level of sensitivity and a faster processing time, whereas the ELISA method has a higher level of selectivity and less cost efficiency. The western blot method has selectivity for detecting complex-level protein expression.
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Mizerska-Kowalska, Magdalena, Adrianna Sławińska-Brych, Katarzyna Kaławaj, Aleksandra Żurek, Beata Pawińska, Wojciech Rzeski, and Barbara Zdzisińska. "Betulin Promotes Differentiation of Human Osteoblasts In Vitro and Exerts an Osteoinductive Effect on the hFOB 1.19 Cell Line Through Activation of JNK, ERK1/2, and mTOR Kinases." Molecules 24, no. 14 (July 19, 2019): 2637. http://dx.doi.org/10.3390/molecules24142637.
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Although betulin (BET), a naturally occurring pentacyclic triterpene, has a variety of biological activities, its osteogenic potential has not been investigated so far. The aim of this study was to assess the effect of BET on differentiation of human osteoblasts (hFOB 1.19 and Saos-2 cells) in vitro in osteogenic (with ascorbic acid as an osteogenic supplement) and osteoinductive (without an additional osteogenic supplement) conditions. Osteoblast differentiation was evaluated based on the mRNA expression (RT-qPCR) of Runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), type I collagen-α1 (COL1A1), and osteopontin (OPN). Additionally, ALP activity and production of COL1A1 (western blot analysis) and OPN (ELISA) were evaluated. The level of mineralization (calcium accumulation) was determined with Alizarin red S staining. BET upregulated the mRNA level of RUNX2 and the expression of other osteoblast differentiation markers in both cell lines (except the influence of BET on ALP expression/activity in the Saos-2 cells). Moreover, it increased mineralization in both cell lines in the osteogenic conditions. BET also increased the mRNA level of osteoblast differentiation markers in both cell lines (except for ALP in the Saos-2 cells) in the osteoinductive conditions, which was accompanied with increased matrix mineralization. The osteoinductive activity of BET in the hFOB 1.19 cells was probably mediated via activation of MAPKs (JNK and ERK1/2) and mTOR, as the specific inhibitors of these kinases abolished the BET-induced osteoblast differentiation. Our results suggest that BET has the potential to enhance osteogenesis.
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Hao, Wei, Hongzhi Liu, Lugang Zhou, Yujie Sun, Hao Su, Jianqiang Ni, Tian He, Peng Shi, and Xin Wang. "MiR-145 regulates osteogenic differentiation of human adipose-derived mesenchymal stem cells through targeting FoxO1." Experimental Biology and Medicine 243, no. 4 (December 17, 2017): 386–93. http://dx.doi.org/10.1177/1535370217746611.
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In this study, we aimed to investigate the expression of miR-145 before and after hASCs osteogenic differentiation. We also intended to explore the influence of the target relationship between miR-145 and FoxO1 on osteogenic differentiation. Dual-luciferase reporter gene assay and real-time PCR were used to confirm the target relationship between miR-145 and FoxO1. Furthermore, the modulatory effects of miR-145 and FoxO1 on hASCs osteoinductive differentiation were measured by real-time PCR , Western blot, ALP staining, ARS staining, and cell immunofluorescence assay. After osteogenic differentiation, miR-145 was gradually down-regulated, while FoxO1 was up-regulated in hASCs. MiR-145 could directly target FoxO1 3′UTR. FoxO1 was negatively regulated by miR-145. After osteoinductive differentiation, BSP, Ocn, and OPN expression was lowered with the overexpression of miR-145 or the knockdown of FoxO1. Furthermore, ALP and ARS staining assay results showed weakened ALP activity and extracellular matrix calcification. When overexpressing miR-145 and FoxO1 simultaneously, no obvious change in ALP activity and extracellular matrix calcification was seen. MiR-145 could suppress hASCs osteoinductive differentiation by suppressing FoxO1 directly. Impact statement Researching on ASCs was a promising strategy to study osteogenic differentiation. The regulatory role of miR-145 on hASCs osteogenic differentiation remained partially explored. Our study revealed a novel mechanism of the osteogenic differentiation process and suggested that miR-145 and its target gene FoxO1 may be potential targets for the therapy of human osteogenic-related disorders.
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Qi, Chen, Xu Xiaofeng, and Wang Xiaoguang. "Effects of Toll-Like Receptors 3 and 4 in the Osteogenesis of Stem Cells." Stem Cells International 2014 (2014): 1–7. http://dx.doi.org/10.1155/2014/917168.
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Objective. To investigate the effects of Toll-like receptors in stem cell osteogenesis.Methods. Bone marrow mesenchymal stem cells (BMSCs) were divided into the blank group, the TLR-3 activated group, and the TLR-4 activated group. After 10 days’ osteogenic-promoting culture, expression of type I collagen and osteocalcin was determined by Western blot. Osteoblasts (OBs) were also divided into three groups mentioned above. Alkaline phosphatase (ALP) and alizarin red staining were performed after 10 days’ ossification-inducing culture. The expression ofβ-catenin was investigated by Western blot.Results. Both the TLR-3 and TLR-4 activated groups had increased expression of type I collagen and osteocalcin; the effect of TLR-4 was stronger. The intensity of alizarin red and ALP staining was strongest in the TLR-3 activated group and weakest in the TLR-4 activated group. Activation of TLR-4 decreased the expression ofβ-catenin, whilst activation of TLR-3 did not affect the expression ofβ-catenin.Discussion. This study suggested that both TLR-3 and -4 promoted differentiation of BMSCs to OBs. TLR-3 had an inducing effect on the ossification of OBs to osteocytes, whilst the effect of TLR-4 was the opposite because of its inhibitory effect on the Wnt signaling pathway.
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You, Chuanfei, Jun Liu, Ruoyu Qiu, Leijun Xu, Furen Dai, Qianzhao Ni, and Weisheng Qiu. "MiR-141 Modulates Bone Marrow Mesenchymal Stem Cells (BMSCs) Osteogenic/Adipogenic Differentiation Under Oxidative Stress." Journal of Biomaterials and Tissue Engineering 12, no. 7 (July 1, 2022): 1466–71. http://dx.doi.org/10.1166/jbt.2022.3055.
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BMSCs Osteogenic differentiation is beneficial to the construction of bone tissue engineering. Oxidative stress can affect BMSCs differentiation. MiR-141 regulates BMSCs proliferation. However, MiR-141’s role in BMSCs osteogenic/adipogenic differentiation under oxidative stress is unclear. Mice BMSCs were assigned into control group; oxidative stress group; and si-MiR-141 group followed by detecting miR-141 level. After 14 days of osteogenesis or adipogenesis induction, RUNX2, OPN and FABP4 mRNA level was analyzed together with analysis of ROS and SOD content, ALP activity and TGFβ/smad signaling protein level by Western blot. Under oxidative stress, MiR-141 was significantly upregulated and RUNX2 and OPN level was decreased, along with increased ROS content and FABP4 level, reduced SOD and ALP activity and expression of TGFβ1 and smad2 (P < 0.05). Transfection of MiR-141 siRNA into BMSCs under oxidative stress down-regulated MiR-141, significantly upregulated RUNX2 and OPN, reduced ROS, elevated SOD activity, downregulated FABP4, and increased ALP activity and TGFβ1 and smad2 expression (P < 0.05). In conclusion, MiR-141 expression is increased in BMSCs under oxidative stress. Down-regulating MiR-141 improves the redox imbalance through TGFβ/smad signaling pathway, promotes osteogenic differentiation of BMSCs and inhibits differentiation to adipocytes.
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Xu, Jiali, Lei He, and Haibing Yang. "Triiodothyronine participates in odontoblast differentiation of apical papilla stem cells through regulation of ERK and p38MAPK signaling pathways." Tropical Journal of Pharmaceutical Research 21, no. 9 (October 13, 2022): 1867–72. http://dx.doi.org/10.4314/tjpr.v21i9.8.
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Purpose: To investigate the effect of triiodothyronine (T3) in odontoblast differentiation of apical papilla stem cells, and the mechanism of action involved.
Methods: Apical unclosed permanent molars extracted from patients due to orthodontics and impaction were selected. The extracted teeth were cultured in the isolation stage of SCAP cells. The cells were exposed to different concentrations of T3. The effects of ERK and p38 MAPK signaling pathways on activity of alkaline phosphatase (ALP) were determined. Calcium deposition was measured using a calcium determination kit, while the expression of BMP - 2 protein by T3 was determined by Western blot assay. Fluorescence quantitative polymerase chain reaction (FqPCR) method was used to determine the mRNA expression of BMP.
Results: The ALP activities were significantly higher in T3 groups than in control group. Relative to control, there were marked differences in ALP activity and calcium deposition in T3 group, T3 + PD group and T3 + SB group (p < 0.05). Relative to control, the mRNA and protein expressions of BMP-2 in T3 group were increased significantly (p < 0.05).
Conclusion: Triiodothyronine regulates the differentiation of apical papilla stem cells into dentin through ERK and p38MAPK signaling pathways. This provides the mechanism underlying odontoblast differentiation of apical papilla stem cells.
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Liu, Jiannan, Mingjiang Jin, Zhen Zhang, Lihuang Wu, Xuejun Jin, Chenping Zhang, and Yue Xing. "Effects of Titanium Micro-Nanopermeable Structures on Osteogenic Differentiation." Journal of Nanomaterials 2018 (December 17, 2018): 1–11. http://dx.doi.org/10.1155/2018/6468246.
Full textAbstract:
To evaluate the effects of different Ti surface micro-nanopermeable structures on osteoblast proliferation and differentiation and explore related mechanisms, hybrid technology of sandblast, acid etching, and hydrothermal (HT) was used to form the micro-nanopermeable surface of Ti. Scanning electron microscopy (SEM), surface profiler, and contact angle meter were utilized to assess the surface morphology, roughness, and hydrophilicity. MTT, SEM, alkaline phosphatase (ALP) activity assay, and real-time PCR were performed to investigate proliferation, adhesion and spreading, and differentiation of MC3T3-E1 cells grown on polished Ti (control), sandblast + acid etching- (SLA-) treated Ti, and SLA + HT-treated Ti. MAPK signal pathway activity was evaluated by Western blotting. The results showed that SLA + HT could result in not only formation of microscale groove containing submicroscale and nanoscale porous structures in Ti samples but also rough and hydrophilic surface. SLA + HT treatment has the best effects on cell adhesion and spreading. Significantly increased levels of ALP activity and osteogenic genes including Alp, Ocn, Opn, Runx2, and Bsp, as well as p38 but not ERK phosphorylation, were found in the SLA + HT group. In conclusion, sandblast, acid etching, and hydrothermal treatment on Ti regulates osteoblast differentiation, while activation of the MAPK p38 signaling pathway served as the mechanism.
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Shao, Xinyu, Xiaojun Cao, Ge Song, Yuan Zhao, and Bimin Shi. "Metformin Rescues the MG63 Osteoblasts against the Effect of High Glucose on Proliferation." Journal of Diabetes Research 2014 (2014): 1–6. http://dx.doi.org/10.1155/2014/453940.
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Aims. To study the proliferation of osteoblasts and genes expression under normal glucose, high glucose, and metformin (Met).Methods. MG63 osteoblast-like cells were cultured in osteogenic medium supplemented with normal glucose (glucose 5.5 mmol/L) or high glucose (glucose 16.7 mmol/L) and metformin + high glucose (Met 300 μmol/L + glucose 16.7 mmol/L). Proliferation was detected with CCK-8 assay at days 1, 3, and 7. Real-time PCR and Western blot were performed to compare the expression of collagen I (Col I), osteocalcin (OCN), osteoprotegerin (OPG), receptor activator for NF-κB ligand (RANKL), and metal matrix proteinases 1 and 2 (MMP1, MMP2). Alkaline phosphatase (ALP) activity was also detected at days 6, 12, and 18.Results. Exposure to high glucose inhibited the proliferation of osteoblasts (P<0.05), with suppressed OCN and OPG. Meanwhile, Col I, RANKL, MMP1, and MMP2 were unaffected. Metformin attenuated the suppression on proliferation with increased expression of Col I, OCN, and OPG, meanwhile suppressing MMP1 and MMP2. High glucose lowered the intracellular ALP, while metformin raised it. Metformin attenuated the downregulation of ALP completely at day 6, partly at day 12, but not at day 18.Conclusions. Metformin attenuated the suppression effect of high glucose to the osteoblast proliferation and gene expression, more prominently in earlier stage.
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Yang, Gui-Cun, You-Hua Xu, Hong-Xia Chen, and Xiao-Jing Wang. "Acute Lymphoblastic Leukemia Cells Inhibit the Differentiation of Bone Mesenchymal Stem Cells into Osteoblasts In Vitro by Activating Notch Signaling." Stem Cells International 2015 (2015): 1–11. http://dx.doi.org/10.1155/2015/162410.
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The disruption of normal hematopoiesis has been observed in leukemia, but the mechanism is unclear. Osteoblasts originate from bone mesenchymal stem cells (BMSCs) and can maintain normal hematopoiesis. To investigate how leukemic cells inhibit the osteogenic differentiation of BMSCs and the role of Notch signaling in this process, we cocultured BMSCs with acute lymphoblastic leukemia (ALL) cells in osteogenic induction medium. The expression levels of Notch1, Hes1, and the osteogenic markers Runx2, Osteopontin (OPN), and Osteocalcin (OCN) were assessed by real-time RT-PCR and western blotting on day 3. Alkaline phosphatase (ALP) activity was analyzed using an ALP kit, and mineralization deposits were detected by Alizarin red S staining on day 14. And then we treated BMSCs with Jagged1 and anti-Jagged1 neutralizing Ab. The expression of Notch1, Hes1, and the abovementioned osteogenic differentiation markers was measured. Inhibition of the expression of Runx2, OPN, and OCN and reduction of ALP activity and mineralization deposits were observed in BMSCs cocultured with ALL cells, while Notch signal inhibiting rescued these effects. All these results indicated that ALL cells could inhibit the osteogenic differentiation of BMSCs by activating Notch signaling, resulting in a decreased number of osteoblastic cells, which may impair normal hematopoiesis.
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Su, Shu-Jem, Yao-Tsung Yeh, Shu-Hui Su, Kee-Lung Chang, Huey-Wen Shyu, Kuan-Ming Chen, and Hua Yeh. "Biochanin A Promotes Osteogenic but Inhibits Adipogenic Differentiation: Evidence with Primary Adipose-Derived Stem Cells." Evidence-Based Complementary and Alternative Medicine 2013 (2013): 1–12. http://dx.doi.org/10.1155/2013/846039.
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Biochanin A has promising effects on bone formationin vivo, although the underlying mechanism remains unclear yet. This study therefore aimed to investigate whether biochanin A regulates osteogenic and adipogenic differentiation using primary adipose-derived stem cells. The effects of biochanin A (at a physiologically relevant concentration of 0.1–1 μM) were assessedin vitrousing various approaches, including Oil red O staining, Nile red staining, alizarin red S staining, alkaline phosphatase (ALP) activity, flow cytometry, RT-PCR, and western blotting. The results showed that biochanin A significantly suppressed adipocyte differentiation, as demonstrated by the inhibition of cytoplasmic lipid droplet accumulation, along with the inhibition of peroxisome proliferator-activated receptor gamma (PPARγ), lipoprotein lipase (LPL), and leptin and osteopontin (OPN) mRNA expression, in a dose-dependent manner. On the other hand, treatment of cells with 0.3 μM biochanin A increased the mineralization and ALP activity, and stimulated the expression of the osteogenic marker genes ALP and osteocalcin (OCN). Furthermore, biochanin A induced the expression of runt-related transcription factor 2 (Runx2), osteoprotegerin (OPG), and Ras homolog gene family, member A (RhoA) proteins. These observations suggest that biochanin A prevents adipogenesis, enhances osteoblast differentiation in mesenchymal stem cells, and has beneficial regulatory effects in bone formation.
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Fang, Hongsong, Shuang Deng, Zhihui Jin, Hao Peng, and Sen Chen. "The Role of MiR-339 in the Teriparatide-Induced Proliferation and Differentiation of Bone Marrow Mesenchymal Stem Cells." Journal of Biomaterials and Tissue Engineering 9, no. 8 (August 1, 2019): 1148–53. http://dx.doi.org/10.1166/jbt.2019.2104.
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Osteoporosis is closely related to BMMSCs differentiation. Teriparatide promotes BMMSCs proliferation and differentiation with unclear mechanism. Both miR-339 and DLX5 are closely related to the differentiation of BMMSCs. Our study intends to assess the mechanism by how teripeptide promotes the proliferation and differentiation of BMMSCs. Rat BMMSCs were cultured and transfected with miR-339 inhibitor/NC and then treated with tripeptide (0, 10, 20, 50 nmol/L) followed by analysis of cell proliferation by CCK8 assay, alkaline phosphatase (ALP) activity, expression of miR-339, DLX5, Runx2 and OCN by real-time PCR, and DLX5 protein level by Western blot. miR-339 inhibitor transfection significantly decreased miR-339 expression, increased DLX-5 protein level, cell number, ALP activity and expression of osteogenic genes. Compared with 0 nmol/L group, 10, 20, 50 nmol/L group presented significantly increased cell number. With increased teriparatide concentration, cell number, ALP activity and expression levels of Runx2 and OCN was increased gradually and miR-339 and DLX5 expression was reduced. The luciferase activity in miR339 inhibitor and pmirGLO-DLX5-3′ UTR-wt-transfected cells was higher than cells transfected with miR-339 NC and pmirGLO-DLX5-3′ UTR-wt. Teriparatide promotes BMMSCs osteogenic differentiation by down-regulating miR-339 which targets DLX5 expression.
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Yang, Qing, Lei Wu, Yang Liu, and Bing Yuan. "Effect of Chordin-Like 1 on Differentiation of Bone Marrow Mesenchymal Stem Cells Through Wnt5/TGF-β Signaling Pathway." Journal of Biomaterials and Tissue Engineering 9, no. 9 (September 1, 2019): 1304–10. http://dx.doi.org/10.1166/jbt.2019.2138.
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Chordin-like 1 (CHRDL1) functions in multiple tissues and organs. However, whether CHRDL1 affects bone marrow mesenchymal stem cells (BMSCs) differentiation remain unclear. Rat BMSCs were isolated and divided into control group, CHRDL1 group and CHRDL1 siRNA group followed by analysis of CHRDL1 level by real time PCR and ELISA, cell proliferation by MTT assay, Caspase 3 activity, ALP activity, expression of o Runx2, OC and PPARγ2 by Real time PCR, TGF-β secretion by ELIS, and Wnt5 protein expression by Western blot. CHRDL1 expression was significantly increased in CHRDL1 group, along with significantly promoted cell proliferation, decreased Caspase 3 activity, increased ALP activity and expression of Runx2 and OC, decreased PPARγ2 expression, increased TGF-β secretion and Wnt5 expression compared to control group (P < 0.05). However, CHRDL1 siRNA transfection significantly decreased CHRDL1 expression, inhibited cell proliferation, increased Caspase 3 activity, decreased ALP activity and Runx2 and OC expression, increased PPARγ2 expression, decreased TGF-β secretion and Wnt5 expression. (P < 0.05). Down-regulation of CHRDL1 expression in BMSCs promotes Wnt5/TGF-β signaling transduction, which in turn increases BMSCs proliferation and osteogenic differentiation. Up-regulation of CHRDL1 expression in BMSCs inhibited the activation of Wnt5/TGF-β signaling pathway, promoted BMSCs apoptosis, and inhibited BMSCs proliferation and osteogenic differentiation.