Viburnum opulus L. Juice Phenolic Compounds Influence Osteogenic Differentiation in Human Osteosarcoma Saos-2 Cells

Bone mass loss occurs with a decrease in osteoblast proliferation and differentiation, or the enhancement of bone resorption, which further leads to the impairment of bone mineral density and increase in bone fracture. Recent studies suggest that some phenolic compounds found in food play positive role in bone metabolism. High content of phenolic compounds with potential beneficial effects on bone metabolism have been identified in the Viburnum opulus fruit. The aim of the study was to determine the influence of V. opulus fresh juice (FJ) and juice purified by solid phase extraction (PJ) on osteogenesis processes with osteosarcoma Saos-2 cell lines. V. opulus purified juice revealed stronger potential as an inducer of Saos-2 osteogenic differentiation. Saos-2 cells matrix mineralization was evaluated with alkaline phosphatase (ALP) activity measurement and alizarin red S staining. Gene expression analysis showed the elevation of the mRNA levels of Runt-related transcription factor 2 (RUNX2), ALP, collagen type 1 and osteonectin, whereas the nuclear factor-κB ligand and osteoprotegerin ratio (RANKL/OPG) decreased. Furthermore, V. opulus was able to diminish the secretion of pro-inflammatory cytokines Il6 and TNFα, however had no effect on vascular endothelial growth factor (VEGF). It decreased intracellular oxidative stress and induced DNA repair, but had no effect on the growth inhibition of lactic acid beneficial microorganisms.

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Bone and Mineral Metabolism Phenotypes in MEN1-Related and Sporadic Primary Hyperparathyroidism, before and after Parathyroidectomy

Cells ◽

10.3390/cells10081895 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1895

Author(s):

Francesca Marini ◽

Francesca Giusti ◽

Federica Cioppi ◽

Davide Maraghelli ◽

Tiziana Cavalli ◽

...

Keyword(s):

Primary Hyperparathyroidism ◽

Bone Mass ◽

Bone Metabolism ◽

Mineral Metabolism ◽

Parathyroid Tissue ◽

Young Patients ◽

Surgical Removal ◽

Mineral Density ◽

Bone Mass Loss ◽

Before And After

Primary hyperparathyroidism (PHPT) is the most common endocrinopathy in multiple endocrine neoplasia type 1 (MEN1). Persistent levels of increased parathyroid hormone (PTH) result in a higher incidence of osteopenia and osteoporosis compared to the general population. Surgical removal of hyper-functioning parathyroid tissue is the therapy of choice. This retrospective study evaluated the effect of parathyroidectomy (PTX) on bone metabolism and bone mass in two series of patients with MEN1 PHPT and sporadic PHPT (sPHPT) by comparing bone metabolism-related biochemical markers and bone mineral density (BMD) before and after surgery. Our data confirmed, in a higher number of cases than in previously published studies, the efficacy of PTX, not only to rapidly restore normal levels of PTH and calcium, but also to normalize biochemical parameters of bone resorption and bone formation, and to improve spine and femur bone mass, in both MEN1 PHPT and sPHPT. Evaluation of single-patient BMD changes after surgery indicates an individual variable bone mass improvement in a great majority of MEN1 PHPT patients. In MEN1 patients, PTX is strongly suggested in the presence of increased PTH and hypercalcemia to prevent/reduce the early-onset bone mass loss and grant, in young patients, the achievement of the bone mass peak; routine monitoring of bone metabolism and bone mass should start from adolescence. Therapy with anti-fracture drugs is indicated in MEN1 patients with BMD lower than the age-matched normal values.

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Effect of dexamethasone as osteogenic supplementation in in vitro osteogenic differentiation of stem cells from human exfoliated deciduous teeth

Journal of Materials Science Materials in Medicine ◽

10.1007/s10856-020-06475-6 ◽

2021 ◽

Vol 32 (1) ◽

Author(s):

Mariane Beatriz Sordi ◽

Raissa Borges Curtarelli ◽

Izabella Thaís da Silva ◽

Gislaine Fongaro ◽

Cesar Augusto Magalhães Benfatti ◽

...

Keyword(s):

Stem Cells ◽

Extracellular Matrix ◽

Osteogenic Differentiation ◽

Culture Media ◽

Deciduous Teeth ◽

Free Calcium ◽

Alizarin Red ◽

Matrix Mineralization ◽

Media Supplementation

AbstractIn in vitro culture systems, dexamethasone (DEX) has been applied with ascorbic acid (ASC) and β-glycerophosphate (βGLY) as culture media supplementation to induce osteogenic differentiation of mesenchymal stem cells. However, there are some inconsistencies regarding the role of DEX as osteogenic media supplementation. Therefore, this study verified the influence of DEX culture media supplementation on the osteogenic differentiation, especially the capacity to mineralize the extracellular matrix of stem cells from human exfoliated deciduous teeth (SHED). Five groups were established: G1—SHED + Dulbecco’s Modified Eagles’ Medium (DMEM) + fetal bovine serum (FBS); G2—SHED + DMEM + FBS + DEX; G3—SHED + DMEM + FBS + ASC + βGLY; G4—SHED + DMEM + FBS + ASC + βGLY + DEX; G5—MC3T3-E1 + α Minimal Essential Medium (MEM) + FBS + ASC + βGLY. DNA content, alkaline phosphatase (ALP) activity, free calcium quantification in the extracellular medium, and extracellular matrix mineralization quantification through staining with von Kossa, alizarin red, and tetracycline were performed on days 7 and 21. Osteogenic media supplemented with ASC and β-GLY demonstrated similar effects on SHED in the presence or absence of DEX for DNA content (day 21) and capacity to mineralize the extracellular matrix according to alizarin red and tetracycline quantifications (day 21). In addition, the presence of DEX in the osteogenic medium promoted less ALP activity (day 7) and extracellular matrix mineralization according to the von Kossa assay (day 21), and more free calcium quantification at extracellular medium (day 21). In summary, the presence of DEX in the osteogenic media supplementation did not interfere with SHED commitment into mineral matrix depositor cells. We suggest that DEX may be omitted from culture media supplementation for SHED osteogenic differentiation in vitro studies.

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Concentrated Growth Factors (CGF) Induce Osteogenic Differentiation in Human Bone Marrow Stem Cells

Biology ◽

10.3390/biology9110370 ◽

2020 ◽

Vol 9 (11) ◽

pp. 370

Author(s):

Alessio Rochira ◽

Luisa Siculella ◽

Fabrizio Damiano ◽

Andrea Palermo ◽

Franco Ferrante ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Osteogenic Differentiation ◽

Bone Marrow Stem Cells ◽

Protein Quantification ◽

Human Bone ◽

Human Bone Marrow ◽

Osteogenic Medium ◽

Alizarin Red ◽

Matrix Mineralization

Bone regeneration is a complex process regulated by several factors that control overlapping biological processes, coordinating interactions among distinct cell populations. There is a great interest in identifying new strategies for inducing osteogenesis in a safe and efficient manner. Concentrated Growth Factor (CGF) is an autologous blood derived product obtained by centrifugation of venous blood following the procedure set on the Silfradent device. In this study the effects of CGF on osteogenic differentiation of human Bone Marrow Stem Cells (hBMSC) in vitro have been investigated; hBMSC were cultured with CGF or osteogenic medium, for 21 days. The osteogenic differentiation was evaluated measuring alkaline phosphatase (ALP) enzyme activity, matrix mineralization by alizarin red staining and through mRNA and protein quantification of osteogenic differentiation markers by Real-time PCR and Western blotting, respectively. The treatment with CGF stimulated ALP activity and promoted matrix mineralization compared to control and seems to be more effective than osteogenic medium. Also, hBMSC lost mesenchymal markers and showed other osteogenic features. Our study showed for the first time that CGF alone is able to induce osteogenic differentiation in hBMSC. The application of CGF on hBMSC osteoinduction might offer new clinical and biotechnological strategies in the tissue regeneration field.

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The Roles of Epigenetics Regulation in Bone Metabolism and Osteoporosis

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2020.619301 ◽

2021 ◽

Vol 8 ◽

Author(s):

Fei Xu ◽

Wenhui Li ◽

Xiao Yang ◽

Lixin Na ◽

Linjun Chen ◽

...

Keyword(s):

Dna Methylation ◽

Histone Modification ◽

Osteogenic Differentiation ◽

Bone Metabolism ◽

Bone Fragility ◽

Research Progress ◽

Epigenetic Mechanisms ◽

Mineral Density ◽

Transcriptional Regulatory ◽

Non Coding Rnas

Osteoporosis is a metabolic disease characterized by decreased bone mineral density and the destruction of bone microstructure, which can lead to increased bone fragility and risk of fracture. In recent years, with the deepening of the research on the pathological mechanism of osteoporosis, the research on epigenetics has made significant progress. Epigenetics refers to changes in gene expression levels that are not caused by changes in gene sequences, mainly including DNA methylation, histone modification, and non-coding RNAs (lncRNA, microRNA, and circRNA). Epigenetics play mainly a post-transcriptional regulatory role and have important functions in the biological signal regulatory network. Studies have shown that epigenetic mechanisms are closely related to osteogenic differentiation, osteogenesis, bone remodeling and other bone metabolism-related processes. Abnormal epigenetic regulation can lead to a series of bone metabolism-related diseases, such as osteoporosis. Considering the important role of epigenetic mechanisms in the regulation of bone metabolism, we mainly review the research progress on epigenetic mechanisms (DNA methylation, histone modification, and non-coding RNAs) in the osteogenic differentiation and the pathogenesis of osteoporosis to provide a new direction for the treatment of bone metabolism-related diseases.

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TIMP-1 inhibits proliferation and osteogenic differentiation of hBMSCs through Wnt/β-catenin signaling

Bioscience Reports ◽

10.1042/bsr20181290 ◽

2019 ◽

Vol 39 (1) ◽

Cited By ~ 5

Author(s):

Tangzhao Liang ◽

Wenling Gao ◽

Lei Zhu ◽

Jianhua Ren ◽

Hui Yao ◽

...

Keyword(s):

Cyclin D1 ◽

Osteogenic Differentiation ◽

Tissue Inhibitor Of Metalloproteinase ◽

Peroxisome Proliferator ◽

Mrna Levels ◽

Alizarin Red ◽

Differentiation Potential ◽

Peroxisome Proliferator Activated Receptor ◽

Alp Activity ◽

Related Transcription Factor

Abstract The present study aimed to evaluate the effect of tissue inhibitor of metalloproteinase-1 (TIMP-1) on the proliferation and osteogenic differentiation potential of human bone marrow-derived MSCs (hBMSCs). hBMSCs with stable TIMP-1 overexpression or TIMP-1 knockdown were generated. Osteogenic differentiation was assessed by Alizarin Red S staining, alkaline phosphatase (ALP) activity and expression of specific markers. Compared with the vehicle controls, TIMP-1 knockdown significantly promoted the growth of hBMSCs. TIMP-1 knockdown up-regulated β-catenin and cyclin D1 proteins. During osteogenic differentiation, TIMP-1 knockdown elevated the deposition of calcium nodules, ALP activity and the mRNA levels of the osteogenic markers sex determining region Y-box 9 (Sox9), CCAAT-enhancer-binding protein and peroxisome proliferator-activated receptor γ. During osteogenic differentiation, TIMP-1 knockdown significantly enhanced the up-regulation of osteocalcin proteins. Meanwhile, TIMP-1 overexpression attenuated the Wnt/activator Wnt3a-induced up-regulation cyclin D1 and Runt-related transcription factor 2 (RUNX-2) (during osteogenic differentiation) proteins, while TIMP-1 knockdown restored the inhibitor Dickkopf 1-induced inhibition effect on the expression of β-catenin, cyclin D1 and RUNX-2. TIMP-1 plays a negative regulatory role in the proliferation and osteogenesis of hBMSCs, at least partially, through Wnt/β-catenin signaling.

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Moringa Oleifera Leaf Extracts Protect BMSC Osteogenic Induction Following Peroxidative Damage by Activating The PI3K/Akt /Foxo1 Pathway

10.21203/rs.3.rs-57508/v1 ◽

2020 ◽

Author(s):

Meiling Liu ◽

Haifeng Ding ◽

Hongzhi Wang ◽

Manfeng Wang ◽

Xiaowei Wu ◽

...

Keyword(s):

Cell Viability ◽

Osteogenic Differentiation ◽

Moringa Oleifera ◽

Caspase 3 ◽

Therapeutic Effects ◽

Mrna Levels ◽

Leaf Extracts ◽

Alizarin Red ◽

Peroxidative Damage ◽

Osteogenic Induction

Abstract Objective: We aimed to investigate the therapeutic effects of Moringa oleifera leaf extracts on osteogenic induction of rat bone marrow mesenchymal stem cells (BMSCs) following peroxidative damage and to explore the underlying mechanisms. Methods: Conditioned medium was used to induce osteogenic differentiation of BMSCs, which were treated with H2O2, Moringa oleifera leaf extracts-containing serum, or the phosphatidyl inositol-3 kinase (PI3K) inhibitor Wortmannin, alone or in combination. Cell viability was measured using the MTT assay. Cell cycle was assayed using flow cytometry. Expression levels of Akt, phosphorylated (p)Akt, Foxo1, and cleaved caspase-3 were analyzed using Western blot analysis. The mRNA levels of osteogenesis-associated genes, including alkaline phosphatase (ALP), collagen І, osteopontin (OPN), and Runx2, were detected using qRT-PCR. Reactive oxygen species (ROS) and malondialdehyde (MDA) levels, as well as superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and ALP activity were detected using commercially available kits. Osteogenic differentiation capability was determined using alizarin red staining. Results: During osteogenic induction of rat BMSCs, H2O2 reduced cell viability and proliferation, inhibited osteogenesis, increased ROS and MDA levels, and decreased SOD and GSH-PX activity. H2O2 significantly reduced pAkt and Foxo1 expression, and increased cleaved caspase-3 levels in BMSCs. Additional treatments with Moringa oleifera leaf extracts partially reversed the H2O2-induced changes. Wortmannin partially attenuated the effects of Moringa oleifera leaf extracts on protein expression of Foxo1, pAkt, and cleaved caspase-3, as well as mRNA levels of osteogenesis-associated genes.Conclusion: Moringa oleifera leaf extracts ameliorate peroxidative damage and enhance osteogenic induction of rat BMSCs by activating the PI3K/Akt/Foxo1 pathway.

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Downregulation of Cardiotrophin-Like Cytokine 1(CLCF1) Inhibits Osteoblastic Differentiation by Regulating RANKL/ OPG Pathway

10.21203/rs.3.rs-111138/v1 ◽

2020 ◽

Author(s):

Juan Chen ◽

Huijuan Xu ◽

Yunjing Ye ◽

Lihua Xie ◽

Shengqiang Li ◽

...

Keyword(s):

Bone Metabolism ◽

Postmenopausal Osteoporosis ◽

Osteoblast Differentiation ◽

Gene Knockout ◽

Raji Cell ◽

Osteoblastic Differentiation ◽

Control Group ◽

Mineral Density ◽

Alizarin Red ◽

Il Interleukin

Abstract BackgroundCardiotrophin-like cytokine factor 1 (CLCF1) is a member of the IL (interleukin)-6-type cytokine. Although its immunomodulatory functions are well defined, data on the physiological and pathological functions of the CLCF1 in bone metabolism remains scant. Here, we interrogated the functions and mechanisms of CLCF1 in osteoblast differentiation.MethodsA total of 109 patients with postmenopausal osteoporosis (PMOP) and 94 control group participants were included in this study. Quantitative reverse transcription PCR (RT-qPCR) and Western blot techniques were used to profile the expression of the CLCF1, nuclear factor-kB ligand and its decoy receptor osteoprotegerin (RANKL/OPG), as well as the janus activated kinase (JAK2)/transcription3 (STAT3) pathway. To elucidate the effect of CLCF1 on bone metabolism, we established CLCF1 gene-knockout Raji cell lines and Raji-MG-63 co-culture system, and interrogated the osteoblast differentiation by measuring the activity of ALP and the level of Alizarin-Red S staining.ResultsOur data demonstrated that the expression of CLCF1 was highly downregulated in PMOP compared with the control group. Moreover, the level of CLCF1 was proportional to the bone mineral density (BMD) in the postmenopausal osteoporosis. Furthermore, the deletion of CLCF1 gene inhibits osteoblast differentiation by regulating the RANKL/OPG system.ConclusionOur findings unravel the previously unidentified roles of CLCF1 in the bone-immune crosstalk, as well as potential therapeutic strategies for postmenopausal osteoporosis.

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The miR-187 Induced Bone Reconstruction and Healing in a Mouse Model of Osteoporosis, and Accelerated Osteoblastic Differentiation of Human Multipotent Stromal Cells by Targeting BARX2

10.21203/rs.3.rs-47428/v1 ◽

2020 ◽

Author(s):

Jun Zhang ◽

Feng Pan ◽

Benseng Tang ◽

Jing Li ◽

Zhengang Zha

Keyword(s):

Mouse Model ◽

Osteogenic Differentiation ◽

Bone Repair ◽

Osteoporotic Fractures ◽

Fracture Repair ◽

Luciferase Reporter ◽

Blue Staining ◽

Bone Reconstruction ◽

Alizarin Red ◽

Matrix Mineralization

Abstract Background: Multiple microRNAs (miRNAs) have been proven to regulate osteogenic differentiation by affecting the Runx2 signaling pathway. The intervention of miRNA can delay the progress of osteoporosis (OP) and induce fracture repair by affecting bone regeneration. However, the function and mechanism of miR-187 in osteoporotic fractures are still unknown. Methods: We first established the OP mouse model. Next, the BMD value was certified by iDXA. The miR-187 level in the OP mice and serum of OP patients was identified through qRT-PCR. Bone repair and bone healing were assessed through toluidine blue staining and X-ray, and BARX2 expression was also confirmed. Osteogenesis-related proteins, ALP activity, and the matrix mineralization state were evaluated by western blot, ALP staining, and Alizarin Red staining in hMSCs after transfection with miR-187 mimics, miR-187 inhibitor, or human BarH-like homeobox 2 (BARX2) siRNA. Moreover, the interplay between miR-187 and BARX2 was identified through the dual-luciferase reporter.Results: The BMD value was notably reduced in the OP mice, and miR-187 was markedly downregulated in the OP mice and serum of OP patients. Meanwhile, we proved that miR-187 induced bone reconstruction and healing, and downregulated BARX2 in the OP mouse model. We also proved that BARX2 was a direct target of miR-187, and could be significantly downregulated by miR-187. Furthermore, miR-187 induced osteogenic differentiation of hMSCs by targeting BARX2.Conclusions: The miR-187 might have a significant therapeutic effect in osteoporotic fractures. miR-187 accelerated osteogenic differentiation of hMSCs by directly regulating BARX2.

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Quercetin promotes bone marrow mesenchymal stem cell proliferation and osteogenic differentiation through the H19/miR-625-5p axis to activate the Wnt/β-catenin pathway

BMC Complementary Medicine and Therapies ◽

10.1186/s12906-021-03418-8 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Wei Bian ◽

Shunqiang Xiao ◽

Lei Yang ◽

Jun Chen ◽

Shifang Deng

Keyword(s):

Bone Marrow ◽

Osteogenic Differentiation ◽

Protein Level ◽

Cell Counting ◽

Alizarin Red ◽

Matrix Mineralization ◽

Stem Cell Proliferation ◽

Alp Activity ◽

Marrow Mesenchymal Stem Cells ◽

Quercetin Treatment

Abstract Background Quercetin and H19 can promote osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). However, whether quercetin regulates H19 expression to promote osteogenic differentiation of BMSCs is unclear. Methods BMSC proliferation, matrix mineralization, and alkaline phosphatase (ALP) activity were assessed using the Cell Counting Kit-8, ALP assay kit, and alizarin red staining kit, respectively. Expression of H19, miR-625-5p, BMP-2, osteocalcin, and RUNX2 were measured by qRT-PCR; β-catenin protein level was measured by western blotting. Results Quercetin promoted BMSC proliferation, enhanced ALP activity, and upregulated the expression of BMP-2, osteocalcin, and RUNX2 mRNAs, suggesting that it promoted osteogenic differentiation of BMSCs. Moreover, quercetin increased H19 expression, while the effect of quercetin on BMSCs was reversed by silencing H19 expression. Additionally, miR-625-5p, interacted with H19, was downregulated during quercetin-induced BMSC osteogenic differentiation, which negatively correlated with H19 expression. Silencing miR-625-5p expression promoted BMSC proliferation and osteogenic differentiation, whereas miR-625-5p overexpression weakened the effect of quercetin on BMSCs. Finally, quercetin treatment or downregulation of miR-625-5p expression increased β-catenin protein level in BMSCs. Upregulation or downregulation of miR-625-5p or H19 expression, respectively, inhibited β-catenin protein level in quercetin treated-BMSCs. Conclusion H19 promotes, while miR-625-5p inhibits BMSC osteogenic differentiation. Quercetin activates the Wnt/β-catenin pathway and promotes BMSC osteogenic differentiation via the H19/miR-625-5p axis.

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Intermittent Administration of Parathyroid Hormone Enhances Odonto/Osteogenic Differentiation of Stem Cells from the Apical Papilla via JNK and P38 MAPK Pathways

Stem Cells International ◽

10.1155/2020/5128128 ◽

2020 ◽

Vol 2020 ◽

pp. 1-13 ◽

Cited By ~ 1

Author(s):

Xiyao Pang ◽

Ying Zhuang ◽

Zehan Li ◽

Shuanglin Jing ◽

Qin Cai ◽

...

Keyword(s):

Stem Cells ◽

Parathyroid Hormone ◽

Osteogenic Differentiation ◽

P38 Mapk ◽

Cell Counting ◽

Mrna Levels ◽

Alizarin Red ◽

Alp Activity ◽

Mapk Pathways ◽

Apical Papilla

Objective. Parathyroid hormone (PTH) is considered to be essential during the tooth development. Stem cells from the apical papilla (SCAPs) are responsible for dentine formation. However, the interaction between PTH and SCAPs remains unclear. This study was aimed at investigating the effects of PTH on odonto/osteogenic differentiation capacity of SCAPs and elucidating the underlying molecular mechanisms. Materials and Methods. Here, SCAPs were isolated and identified in vitro. Effects of PTH on the proliferation of SCAPs were determined by Cell Counting Kit-8 (CCK-8), flow cytometry (FCM), and EdU. Alkaline phosphatase (ALP) activity, alizarin red staining, Western blot, and RT-PCR were carried out to detect the odonto/osteogenic differentiation of PTH-treated SCAPs as well as the participation of the MAPK signaling pathway. Results. An ALP activity assay determined that 10-8 mol/L PTH was the optimal concentration for the induction of SCAPs with no significant influence on the proliferation of SCAPs as indicated by CCK-8, FCM, and EdU. The expression of odonto/osteogenic markers was significantly upregulated in mRNA levels and protein levels. Moreover, intermittent treatment of PTH also increased phosphorylation of JNK and P38, and the differentiation was suppressed following the inhibition of JNK and P38 MAPK pathways. Conclusion. PTH can regulate the odonto/osteogenic differentiation of SCAPs via JNK and P38 MAPK pathways.

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