scholarly journals Effects of Toll-Like Receptors 3 and 4 in the Osteogenesis of Stem Cells

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Chen Qi ◽  
Xu Xiaofeng ◽  
Wang Xiaoguang
Keyword(s):  
Stem Cells ◽  
Western Blot ◽  
Type I Collagen ◽  
Wnt Signaling Pathway ◽  
Inhibitory Effect ◽  
Type I ◽  
Toll Like Receptors ◽  
Blank Group ◽  
Alizarin Red ◽  
Marrow Mesenchymal Stem Cells

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.

scholarly journals Promotion of Hepatic Differentiation of Bone Marrow Mesenchymal Stem Cells on Decellularized Cell-Deposited Extracellular Matrix

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Hongliang He ◽  
Xiaozhen Liu ◽  
Liang Peng ◽  
Zhiliang Gao ◽  
Yun Ye ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Extracellular Matrix ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Type I Collagen ◽  
Type Iii Collagen ◽  
Type I ◽  
Hepatic Differentiation ◽  
Marrow Mesenchymal Stem Cells

Interactions between stem cells and extracellular matrix (ECM) are requisite for inducing lineage-specific differentiation and maintaining biological functions of mesenchymal stem cells by providing a composite set of chemical and structural signals. Here we investigated if cell-deposited ECM mimickedin vivoliver's stem cell microenvironment and facilitated hepatogenic maturation. Decellularization process preserved the fibrillar microstructure and a mix of matrix proteins in cell-deposited ECM, such as type I collagen, type III collagen, fibronectin, and laminin that were identical to those found in native liver. Compared with the cells on tissue culture polystyrene (TCPS), bone marrow mesenchymal stem cells (BM-MSCs) cultured on cell-deposited ECM showed a spindle-like shape, a robust proliferative capacity, and a suppressed level of intracellular reactive oxygen species, accompanied with upregulation of two superoxide dismutases. Hepatocyte-like cells differentiated from BM-MSCs on ECM were determined with a more intensive staining of glycogen storage, an elevated level of urea biosynthesis, and higher expressions of hepatocyte-specific genes in contrast to those on TCPS. These results demonstrate that cell-deposited ECM can be an effective method to facilitate hepatic maturation of BM-MSCs and promote stem-cell-based liver regenerative medicine.

scholarly journals Three-Dimensional Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Promotes Matrix Metallopeptidase 13 (MMP13) Expression in Type I Collagen Hydrogels

2021 ◽  
Vol 22 (24) ◽  
pp. 13594
Author(s):  
Luis Oliveros Anerillas ◽  
Paul J. Kingham ◽  
Mikko J. Lammi ◽  
Mikael Wiberg ◽  
Peyman Kelk
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Type I Collagen ◽  
Bone Defects ◽  
Type I ◽  
Collagen Gels ◽  
Alizarin Red ◽  
Matrix Metallopeptidase

Autologous bone transplantation is the principal method for reconstruction of large bone defects. This technique has limitations, such as donor site availability, amount of bone needed and morbidity. An alternative to this technique is tissue engineering with bone marrow-derived mesenchymal stem cells (BMSCs). In this study, our aim was to elucidate the benefits of culturing BMSCs in 3D compared with the traditional 2D culture. In an initial screening, we combined BMSCs with four different biogels: unmodified type I collagen (Col I), type I collagen methacrylate (ColMa), an alginate and cellulose-based bioink (CELLINK) and a gelatin-based bioink containing xanthan gum (GelXA-bone). Col I was the best for structural integrity and maintenance of cell morphology. Osteogenic, adipogenic, and chondrogenic differentiations of the BMSCs in 2D versus 3D type I collagen gels were investigated. While the traditional pellet culture for chondrogenesis was superior to our tested 3D culture, Col I hydrogels (i.e., 3D) favored adipogenic and osteogenic differentiation. Further focus of this study on osteogenesis were conducted by comparing 2D and 3D differentiated BMSCs with Osteoimage® (stains hydroxyapatite), von Kossa (stains anionic portion of phosphates, carbonates, and other salts) and Alizarin Red (stains Ca2+ deposits). Multivariate gene analysis with various covariates showed low variability among donors, successful osteogenic differentiation, and the identification of one gene (matrix metallopeptidase 13, MMP13) significantly differentially expressed in 2D vs. 3D cultures. MMP13 protein expression was confirmed with immunohistochemistry. In conclusion, this study shows evidence for the suitability of type I collagen gels for 3D osteogenic differentiation of BMSCs, which might improve the production of tissue-engineered constructs for treatment of bone defects.

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

2021 ◽  
Vol 11 (8) ◽  
pp. 1630-1635
Author(s):  
Bin Wu ◽  
Fenghua Bai ◽  
Jianping Lin ◽  
Guangji Wang ◽  
Wentao Cai ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Osteogenic Differentiation ◽  
Type I Collagen ◽  
Type I ◽  
Smad Signaling ◽  
Alp Activity ◽  
Smad Signaling Pathway ◽  
Marrow Mesenchymal Stem Cells

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.

Exosomes from bone marrow mesenchymal stem cells promoted osteogenic differentiation by delivering miR-196a that targeted Dkk1 to activate Wnt/β-catenin pathway

2020 ◽  
Author(s):  
Zhi Peng ◽  
Zhenkai Lou ◽  
Zhongjie Li ◽  
Shaobo Li ◽  
Kaishun Yang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Western Blot ◽  
Osteogenic Differentiation ◽  
Osteoblast Differentiation ◽  
Negative Regulator ◽  
Luciferase Assay ◽  
Alizarin Red ◽  
Marrow Mesenchymal Stem Cells

Abstract Background: Osteoporosis is the most common bone metabolic disease. Emerging evidence suggests that exosomes are secreted by diverse cells such as bone marrow mesenchymal stem cells (BMSCs), and play important role in cell-to-cell communication and tissue homeostasis. Recently, the discovery of exosomes has attracted attention in the field of bone remodeling. Methods: The exosomes were extracted from BMSCs and labeled by PKH-67, and then incubated with hFOB1.19 cells to investigate the miR-196a function on the osteoblast differentiation of hFOB1.19. The osteoblast differentiation was detected via alizarin red staining and the expression of osteoblast genes were detected by western blot. The cell apoptosis was detected by flow cytometer. The target relationship of miR-196a and Dickkopf-1 (Dkk1) were verified by luciferase assay and western blot. Results: Here we demonstrated that exosomes extracted from BMSCs (BMSC-exo) significantly promoted hFOB1.19 differentiation to osteoblasts. We found that BMSC-exo were enriched with miR-196a and delivered miR-196a to hFOB1.19 cells to inhibit its target Dkk1, which is a negative regulator of Wnt/β-catenin pathway. Conclusion: BMSC-exo activated Wnt/β-catenin pathway to promote osteogenic differentiation, while BMSC-exo failed to exert the effects when miR-196a was deprived. In conclusion, miR-196a delivered by exosomes from BMSCs plays an essential role in enhancing osteoblastic differentiation by targeting Dkk1 to activate Wnt/β-catenin pathway.

Effect of Hepatic Kinase B1 (LKB1) on Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells During Senescence

2020 ◽  
Vol 10 (2) ◽  
pp. 246-251
Author(s):  
Wenxiao Jiang ◽  
Yijun Zhang ◽  
Ye Huang ◽  
Yunfeng Cheng ◽  
Zhigang Liu
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Type I Collagen ◽  
Mtor Pathway ◽  
Nodule Formation ◽  
Type I ◽  
Alp Activity ◽  
Marrow Mesenchymal Stem Cells

Hepatic kinase B1 (LKB1) is a tumor suppressor and regulates cell proliferation and apoptosis. However, whether LKB1 affects bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation of during aging remains unclear. Two BMSCs derived from Zempster24−/− (aging) and Zempster24+/+ (normal) mice were cultured in vitro followed by measurement of LKB1 expression by real-time quantitative PCR and Western blot. LKB1 siRNA was transfected into Zempster24−/−BMSCs and LKB1 expression was measured. 14 days after osteogenic induction, mineralized nodule formation was evaluated by alizarin red staining, expression of Calcin, type I collagen, RUNX2 and OPN mRNA expression was measured, together with alkaline phosphatase (ALP) activity and the PI3K/mTOR pathway activity. Compared with normal BMSCs, LKB1 expression was significantly increased, calcified nodules were decreased, with reduced expression of osteocalcin, type I collagen, RUNX2 and OPN mRNA as well as decreased ALP activity and PI3K/mTOR signaling protein expression (P < 0.05). LKB1 siRNA transfection into senescent BMSCs down-regulated LKB1 expression, increased calcification nodule formation, expression of osteocalcin, type I collagen, RUNX2 and OPN mRNA, as well as increased ALP activity and PI3K/mTOR pathway protein expression (P < 0.05). Aging can promote the increase of LKB1 expression and inhibit the osteogenic differentiation of BMSCs. Down-regulation of LKB1 expression in BMSCs during senescence can promote osteogenic differentiation through regulating PI3K/mTOR pathway.

scholarly journals Icariin Regulates the Bidirectional Differentiation of Bone Marrow Mesenchymal Stem Cells through Canonical Wnt Signaling Pathway

2017 ◽  
Vol 2017 ◽  
pp. 1-12 ◽  
Author(s):  
Jun-ming Huang ◽  
Yuan Bao ◽  
Wei Xiang ◽  
Xing-zhi Jing ◽  
Jia-chao Guo ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Western Blot ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Wnt Signaling Pathway ◽  
Canonical Wnt Signaling ◽  
Marrow Mesenchymal Stem Cells ◽  
Canonical Wnt

Fat infiltration within the bone marrow is easily observed in some postmenopausal women. Those fats are mainly derived from bone marrow mesenchymal stem cells (BMMSCs). The increment of adipocytes derived from BMMSCs leads to decreased osteoblasts derived from BMMSCs, so the bidirectional differentiation of BMMSCs significantly contributes to osteoporosis. Icariin is the main extractive of Herba Epimedii which is widely used in traditional Chinese medicine. In this experiment, we investigated the effect of icariin on the bidirectional differentiation of BMMSCs through quantitative real-time PCR, immunofluorescence, western blot, and tissue sections in vitro and in vivo. We found that icariin obviously promotes osteogenesis and inhibits adipogenesis through detecting staining and gene expression. Micro-CT analysis showed that icariin treatment alleviated the loss of cancellous bone of the distal femur in ovariectomized (OVX) mice. H&E staining analysis showed that icariin-treated OVX mice obtained higher bone mass and fewer bone marrow lipid droplets than OVX mice. Western blot and immunofluorescence showed that icariin regulates the bidirectional differentiation of BMMSCs via canonical Wnt signaling. This study demonstrates that icariin exerts its antiosteoporotic effect by regulating the bidirectional differentiation of BMMSCs through the canonical Wnt signaling pathway.

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