scholarly journals Mechanical Sensing Element PDLIM5 Promotes Osteogenesis of Human Fibroblasts by Affecting the Activity of Microfilaments

Biomolecules ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 759
Author(s):  
Xiaolan Huang ◽  
Rongmei Qu ◽  
Yan Peng ◽  
Yuchao Yang ◽  
Tingyu Fan ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Morphological Changes ◽  
Human Fibroblasts ◽  
Inhibitory Effect ◽  
Induction Medium ◽  
Sensing Element ◽  
Differentiation Potential ◽  
Osteogenic Induction Medium ◽  
Osteogenic Induction ◽  
Related Proteins

Human skin fibroblasts (HSFs) approximate the multidirectional differentiation potential of mesenchymal stem cells, so they are often used in differentiation, cell cultures, and injury repair. They are an important seed source in the field of bone tissue engineering. However, there are a few studies describing the mechanism of osteogenic differentiation of HSFs. Here, osteogenic induction medium was used to induce fibroblasts to differentiate into osteoblasts, and the role of the mechanical sensitive element PDLIM5 in microfilament-mediated osteogenic differentiation of human fibroblasts was evaluated. The depolymerization of microfilaments inhibited the expression of osteogenesis-related proteins and alkaline phosphatase activity of HSFs, while the polymerization of microfilaments enhanced the osteogenic differentiation of HSFs. The evaluation of potential protein molecules affecting changes in microfilaments showed that during the osteogenic differentiation of HSFs, the expression of PDLIM5 increased with increasing induction time, and decreased under the state of microfilament depolymerization. Lentivirus-mediated PDLIM5 knockdown by shRNA weakened the osteogenic differentiation ability of HSFs and inhibited the expression and morphological changes of microfilament protein. The inhibitory effect of knocking down PDLIM5 on HSF osteogenic differentiation was reversed by a microfilament stabilizer. Taken together, these data suggest that PDLIM5 can mediate the osteogenic differentiation of fibroblasts by affecting the formation and polymerization of microfilaments.

scholarly journals Optimizing Osteogenic Differentiation of Ovine Adipose-Derived Stem Cells by Osteogenic Induction Medium and FGFb, BMP2, or NELL1 In Vitro

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Emil Østergaard Nielsen ◽  
Li Chen ◽  
Jonas Overgaard Hansen ◽  
Matilda Degn ◽  
Søren Overgaard ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Basic Medium ◽  
Induction Medium ◽  
Control Analysis ◽  
Alizarin Red ◽  
Osteogenic Induction Medium ◽  
Osteogenic Induction ◽  
Fibroblast Growth Factor Basic

Although adipose-derived stromal cells (ADSCs) have been a major focus as an alternative to autologous bone graft in orthopedic surgery, bone formation potential of ADSCs is not well known and cytokines as osteogenic inducers on ADSCs are being investigated. This study aimed at isolating ADSCs from ovine adipose tissue (AT) and optimizing osteogenic differentiation of ovine ADSCs (oADSC) by culture medium and growth factors. Four AT samples were harvested from two female ovine (Texel/Gotland breed), and oADSCs were isolated and analyzed by flow cytometry for surface markers CD29, CD44, CD31, and CD45. Osteogenic differentiation was made in vitro by seeding oADSCs in osteogenic induction medium (OIM) containing fibroblast growth factor basic (FGFb), bone morphogenetic protein 2 (BMP2), or NEL-like molecule 1 (NELL1) in 4 different dosages (1, 10, 50, and 100 ng/ml, respectively). Basic medium (DMEM) was used as control. Analysis was made after 14 days by Alizarin red staining (ARS) and quantification. This study successfully harvested AT from ovine and verified isolated cells for minimal criteria for adipose stromal cells which suggests a feasible method for isolation of oADSCs. OIM showed significantly higher ARS to basic medium, and FGFb 10 ng/ml revealed significantly higher ARS to OIM alone after 14 days.

scholarly journals TMT-Based Proteomic Explores the Influence of DHEA on the Osteogenic Differentiation of hBMSCs

2021 ◽  
Vol 9 ◽  
Author(s):  
Xiaonan Liang ◽  
Mingwei He ◽  
Bo Zhu ◽  
Yongjia Zhu ◽  
Xixi He ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Metabolic Pathways ◽  
Osteoblast Differentiation ◽  
Bioinformatic Analysis ◽  
Induction Medium ◽  
Complete Medium ◽  
Tandem Mass ◽  
Osteogenic Induction Medium ◽  
Osteogenic Induction ◽  
Underlying Mechanisms

Dehydroepiandrosterone (DHEA) has been revealed to implicate in facilitating osteoblast differentiation of human bone marrow mesenchymal stem cells (hBMSCs) and inhibiting osteoporosis (OP). However, the underlying molecular mechanism remains largely unknown. Here, we induced osteogenic differentiation of hBMSCs derived from elders using an osteogenic induction medium with or without DHEA. The results showed that osteogenic induction medium (OIM) with DHEA could significantly promote the proliferation and osteogenic differentiation of hBMSCs than OIM alone. By using a Tandem Mass Tag (TMT) labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) technology, we screened out 604 differentially expressed proteins (DEPs) with at least one unique peptide were identified [524: OIM vs. complete medium (CM), and 547: OIM+DHEA vs. CM], among these proteins, 467 DEPs were shared in these two different comparative groups. Bioinformatic analysis revealed these DEPs are mainly enriched in metabolic pathways. Interestingly, the expression levels of the DEPs in the metabolic pathways showed a more noticeable change in the OIM+DHEA vs. CM group than OIM vs. CM group. Moreover, the protein-protein interaction (PPI) network analysis revealed that three potential proteins, ATP5B, MT-CYB, and MT-ATP6, involved in energy metabolism, might play a key role in osteogenic differentiation induced by OIM+DHEA. These findings offer a valuable clue for us to better understand the underlying mechanisms involved in osteoblast differentiation of hBMSCs caused by DHEA and assist in applying DHEA in hBMSCs-based therapy for osteogenic regeneration.

scholarly journals Surface Modification of Biodegradable Mg-Based Scaffolds for Human Mesenchymal Stem Cell Proliferation and Osteogenic Differentiation

Materials ◽  
2021 ◽  
Vol 14 (2) ◽  
pp. 441
Author(s):  
Si-Han Wang ◽  
Shiao-Pieng Lee ◽  
Chung-Wei Yang ◽  
Chun-Min Lo
Keyword(s):  
Mechanical Properties ◽  
Surface Modification ◽  
Osteogenic Differentiation ◽  
Magnesium Alloys ◽  
Human Mesenchymal Stem Cells ◽  
Human Mesenchymal Stem Cell ◽  
Induction Medium ◽  
Culture Dish ◽  
Osteogenic Induction Medium ◽  
Osteogenic Induction

Magnesium alloys with coatings have the potential to be used for bone substitute alternatives since their mechanical properties are close to those of human bone. However, the surface modification of magnesium alloys to increase the surface biocompatibility and reduce the degradation rate remains a challenge. Here, FHA-Mg scaffolds were made of magnesium alloys and coated with fluorohydroxyapatite (FHA). Human mesenchymal stem cells (hMSCs) were cultured on FHA-Mg scaffolds and cell viability, proliferation, and osteogenic differentiation were investigated. The results showed that FHA-Mg scaffolds display a nano-scaled needle-like structure of aggregated crystallites on their surface. The average Mg2+ concentration in the conditioned media collected from FHA-Mg scaffolds (5.8–7.6 mM) is much lower than those collected from uncoated, Mg(OH)2-coated, and hydroxyapatite (HA)-coated samples (32.1, 17.7, and 21.1 mM, respectively). In addition, compared with hMSCs cultured on a culture dish, cells cultured on FHA-Mg scaffolds demonstrated better proliferation and comparable osteogenic differentiation. To eliminate the effect of osteogenic induction medium, hMSCs were cultured on FHA-Mg scaffolds in culture medium and an approximate 66% increase in osteogenic differentiation was observed three weeks later, indicating a significant effect of the nanostructured surface of FHA-Mg scaffolds on hMSC behaviors. With controllable Mg2+ release and favorable mechanical properties, porous FHA-Mg scaffolds have a great potential in cell-based bone regeneration.

scholarly journals MEK1/2 Inhibitor (GDC0623) Promotes Osteogenic Differentiation of Primary Osteoblasts Inhibited by IL-1β through the MEK-Erk1/2 and Jak/Stat3 Pathways

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Zeng-Qiao Zhang ◽  
Xiao-Shen Hu ◽  
Ye-Chen Lu ◽  
Jun-Peng Zhang ◽  
Wen-Yao Li ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Western Blotting ◽  
Cell Counting ◽  
Nuclear Antigen ◽  
Control Group ◽  
Induction Medium ◽  
Alizarin Red ◽  
Real Time Quantitative Pcr ◽  
Osteogenic Induction Medium ◽  
Osteogenic Induction

Objective. We evaluated the effects and mechanisms of GDC0623 on osteogenic differentiation of osteoblasts induced by IL-1β. Methodology. Osteoblasts were treated with 20 ng/ml IL-1β and 0.1 µM GDC0623. Cell proliferation levels were evaluated by the cell counting kit 8 (CCK8), EdU assay, and western blotting [proliferating cell nuclear antigen (PCNA) and Cyclin D1]. Osteoblasts were cultured in an osteogenic induction medium for 1–3 weeks after which their differentiations were assessed by alkaline phosphatase (ALP) staining, Alizarin Red staining, calcium concentration, immunocytochemistry staining, real-time quantitative PCR (RT-qPCR), and immunofluorescence staining. The osteogenesis-associated mechanisms were further evaluated by western blotting using appropriate antibodies. Results. Relative to the control group, IL-1β induced the rapid proliferation of osteoblasts and suppressed their osteogenic differentiations by upregulating the activities of MEK-Erk1/2 as well as Jak-Stat3 pathways and by elevating MMP13 and MMP9 levels. However, blocking of the MEK-Erk1/2 signaling pathway by GDC0623 treatment reversed these effects. Conclusion. Inhibition of Jak-Stat3 pathway by C188-9 downregulated the expression levels of MMP9 and MMP13, activated MEK-Erk1/2 pathway, and inhibited osteogenic differentiation.

scholarly journals In Vitro Fabrication of Hybrid Bone/Cartilage Complex Using Mouse Induced Pluripotent Stem Cells

2020 ◽  
Vol 21 (2) ◽  
pp. 581 ◽  
Author(s):  
Phoonsuk Limraksasin ◽  
Takeru Kondo ◽  
Maolin Zhang ◽  
Hiroko Okawa ◽  
Thanaphum Osathanon ◽  
...  
Keyword(s):  
Cartilage Tissue ◽  
Marker Genes ◽  
Induction Medium ◽  
Mechanical Stimuli ◽  
Tissue Formation ◽  
Osteogenic Induction Medium ◽  
Osteogenic Induction ◽  
Chondrogenic Marker ◽  
Induced Pluripotent

Cell condensation and mechanical stimuli play roles in osteogenesis and chondrogenesis; thus, they are promising for facilitating self-organizing bone/cartilage tissue formation in vitro from induced pluripotent stem cells (iPSCs). Here, single mouse iPSCs were first seeded in micro-space culture plates to form 3-dimensional spheres. At day 12, iPSC spheres were subjected to shaking culture and maintained in osteogenic induction medium for 31 days (Os induction). In another condition, the osteogenic induction medium was replaced by chondrogenic induction medium at day 22 and maintained for a further 21 days (Os-Chon induction). Os induction produced robust mineralization and some cartilage-like tissue, which promoted expression of osteogenic and chondrogenic marker genes. In contrast, Os-Chon induction resulted in partial mineralization and a large area of cartilage tissue, with greatly increased expression of chondrogenic marker genes along with osterix and collagen 1a1. Os-Chon induction enhanced mesodermal lineage commitment with brachyury expression followed by high expression of lateral plate and paraxial mesoderm marker genes. These results suggest that combined use of micro-space culture and mechanical stimuli facilitates hybrid bone/cartilage tissue formation from iPSCs, and that the bone/cartilage tissue ratio in iPSC constructs could be manipulated through the induction protocol.

scholarly journals microRNA-214-3p Suppresses Ankylosing Spondylitis Fibroblast Osteogenesis via BMP–TGFβ Axis and BMP2

2021 ◽  
Vol 11 ◽  
Author(s):  
Lixiang Ding ◽  
Yukun Yin ◽  
Yu Hou ◽  
Haoran Jiang ◽  
Ji Zhang ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Ankylosing Spondylitis ◽  
Osteogenic Differentiation ◽  
Target Gene ◽  
Loss Of Function ◽  
Alizarin Red ◽  
Neck Fracture ◽  
Osteogenic Induction ◽  
Related Proteins ◽  
Bone Morphogenic Protein 2

Recent investigations suggest microRNAs (miRs) exert functions in fibroblast osteogenesis in ankylosing spondylitis (AS), an inflammatory rheumatic disease. But the mechanism of miR-214-3p in osteogenic differentiation in AS is not clearly understood yet. In this study, fibroblasts were obtained from the capsular ligament of patients with AS and femoral neck fracture and cultured for osteogenic induction and identified. The roles of miR-214-3p and bone morphogenic protein 2 (BMP2) in AS fibroblast osteogenesis were assessed via gain- and loss-of-function, alizarin red S staining, and alkaline phosphatase (ALP) detection. Levels of miR-214-3p, BMP2, osteogenic differentiation-related proteins, and BMP–TGFβ axis-related proteins were further measured. Consequently, miR-214-3p was downregulated in AS fibroblasts, with enhanced ALP activity and calcium nodules, which were reversed by miR-214-3p overexpression. BMP2 was a target gene of miR-214-3p and promoted AS fibroblast osteogenesis by activating BMP–TGFβ axis, while miR-214-3p inhibited AS fibroblast osteogenesis by targeting BMP2. Together, miR-214-3p could prevent AS fibroblast osteogenic differentiation by targeting BMP2 and blocking BMP–TGFβ axis. This study may offer a novel insight for AS treatment.

scholarly journals Exosomal circLPAR1 Promoted Osteogenic Differentiation of Homotypic Dental Pulp Stem Cells by Competitively Binding to hsa-miR-31

2020 ◽  
Vol 2020 ◽  
pp. 1-13
Author(s):  
Liangkun Xie ◽  
Zheng Guan ◽  
Mingzhu Zhang ◽  
Sha Lyu ◽  
Nattawut Thuaksuban ◽  
...  
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Dental Pulp ◽  
Expression Profiles ◽  
Circular Rna ◽  
Inhibitory Effect ◽  
Dental Pulp Stem Cells ◽  
Great Promise ◽  
Human Dental Pulp ◽  
Osteogenic Induction

Human dental pulp stem cells (DPSCs) hold great promise in bone regeneration. However, the exact mechanism of osteogenic differentiation of DPSCs remains unknown, especially the role of exosomes played in. The DPSCs were cultured and received osteogenic induction; then, exosomes from osteogenic-induced DPSCs (OI-DPSC-Ex) at different time intervals were isolated and sequenced for circular RNA (circRNA) expression profiles. Gradually, increased circular lysophosphatidic acid receptor 1 (circLPAR1) expression was found in the OI-DPSC-Ex coincidentally with the degree of osteogenic differentiation. Meanwhile, results from osteogenic differentiation examinations showed that the OI-DPSC-Ex had osteogenic effect on the recipient homotypic DPSCs. To investigate the mechanism of exosomal circLPAR1 on osteogenic differentiation, we verified that circLPAR1 could competently bind to hsa-miR-31, by eliminating the inhibitory effect of hsa-miR-31 on osteogenesis, therefore promoting osteogenic differentiation of the recipient homotypic DPSCs. Our study showed that exosomal circRNA played an important role in osteogenic differentiation of DPSCs and provided a novel way of utilization of exosomes for the treatment of bone deficiencies.

scholarly journals LncRNA DANCR and miR-320a suppressed osteogenic differentiation in osteoporosis by directly inhibiting the Wnt/β-catenin signaling pathway

2020 ◽  
Vol 52 (8) ◽  
pp. 1310-1325
Author(s):  
Cheng-Gong Wang ◽  
Yi-He Hu ◽  
Shi-Long Su ◽  
Da Zhong
Keyword(s):  
Osteogenic Differentiation ◽  
Signaling Pathway ◽  
Morphological Changes ◽  
Osteoporosis Treatment ◽  
Inhibitory Effect ◽  
Luciferase Assay ◽  
Blue Staining ◽  
Alcian Blue Staining ◽  
Alizarin Red

Abstract Our study aimed to determine how lncRNA DANCR, miR-320a, and CTNNB1 interact with each other and regulate osteogenic differentiation in osteoporosis. qRT-PCR and western blotting were performed to determine the expression of DANCR, miR-320a, CTNNB1, and the osteoporosis- or Wnt/β-catenin pathway-related markers T-cell factor 1 (TCF-1), runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), osteocalcin (OCN), and osteopontin (OPN). Interactions between CTNNB1, DANCR, and miR-320a were predicted by bioinformatics approaches and validated using a luciferase assay. Osteoblastic phenotypes were evaluated by ALP staining, ALP activity assay and Alizarin Red staining. The bilateral ovariectomy method was used to establish an in vivo osteoporosis model. Bone morphological changes were examined using hematoxylin and eosin (H&E) and Alcian Blue staining. The expression levels of DANCR and miR-320a in BMSCs derived from osteoporosis patients were upregulated, whereas CTNNB1 expression was downregulated compared with that in healthy controls. Importantly, we demonstrated that miR-320a and DANCR acted independently from each other and both inhibited CTNNB1 expression, whereas the inhibitory effect was additive when miR-320a and DANCR were cooverexpressed. Moreover, we found that DANCR overexpression largely abrogated the effect of the miR-320a inhibitor on CTNNB1 expression and the Wnt/β-catenin signaling pathway in BMSCs during osteogenic differentiation. We further confirmed the results above in BMSCs derived from an osteoporosis animal model. Taken together, our findings revealed that DANCR and miR-320a regulated the Wnt/β-catenin signaling pathway during osteogenic differentiation in osteoporosis through CTNNB1 inhibition. Our results highlight the potential value of DANCR and miR-320a as promising therapeutic targets for osteoporosis treatment.

Sign in / Sign up

Export Citation Format

Share Document