scholarly journals Osteogenic and chondrogenic differentiation potential of mesenchymal stem cells obtained from the bone marrow and placenta

2021 ◽  
Vol 18 (1) ◽  
pp. 36-45
Author(s):  
H. A. Zhernasechanka ◽  
Ya. I. Isaikina ◽  
T. V. Filipovich ◽  
E. G. Liakh
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Alkaline Phosphatase ◽  
Mesenchymal Stem Cells ◽  
Chondrogenic Differentiation ◽  
Cartilage Tissue ◽  
Osteogenic Potential ◽  
Fibrin Gel ◽  
Alizarin Red ◽  
Differentiation Potential

Mesenchymal stem cells (MSC) represent a perspective resource for cell biotechnology. However the question of chondrogenic and osteogenic capacity of MSC of different origin remains under study.The aim of this study was to analyze the osteo-chondrogenic differentiation potential of MSC obtained from the bone marrow and placenta. The results of our studies have indicated that bone marrow-derived and placenta-derived MSC showed a chondrogenic potential in vitro after a chondrogenic induction with specific differentiation media. But for bone marrowderived MSC, the chondrogenic program was realized by expression of collagens (Coll2, Coll10), while in placenta-derived MSC cultures we found a progressive increase in COMP and Ver expression, so bone marrow-derived MSC is more preferable for use in cartilage tissue engineering. Regarding the results on alkaline phosphatase and alizarin red staining, bone marrowderived MSC showed a more significant osteogenic potential compared to placenta-derived MSC. Bone marrow-derived MSC in the composition of fibrin gel after osteogenic induction on the 14th day exhibited the activity of alkaline phosphatase, calcium depositions inside the cells and extracellular matrix, the increase in Sp7 and DMP expression.

scholarly journals Osteoporosis is accompanied by reduced CD274 expression in human bone marrow-derived mesenchymal stem cells

2021 ◽  
Vol 41 ◽  
pp. 603-615
Author(s):  
A-N Zeller ◽  
◽  
M Selle ◽  
Z Gong ◽  
M Winkelmann ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Cell Surface ◽  
Chondrogenic Differentiation ◽  
Surface Antigens ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Differentiation Potential ◽  
Proliferation Potential

Underlying pathomechanisms of osteoporosis are still not fully elucidated. Cell-based therapy approaches pose new possibilities to treat osteoporosis and its complications. The aim of this study was to quantify differences in human bone marrow-derived mesenchymal stem cells (hBMSCs) between healthy donors and those suffering from clinically manifest osteoporosis. Cell samples of seven donors for each group were selected retrospectively from the hBMSC cell bank of the Trauma Department of Hannover Medical School. Cells were evaluated for their adipogenic, osteogenic and chondrogenic differentiation potential, for their proliferation potential and expression of surface antigens. Furthermore, a RT2 Osteoporosis Profiler PCR array, as well as quantitative real-time PCR were carried out to evaluate changes in gene expression. Cultivated hBMSCs from osteoporotic donors showed significantly lower cell surface expression of CD274 (4.98 % ± 2.38 %) than those from the control group (26.03 % ± 13.39 %; p = 0.007), as assessed by flow cytometry. In osteoporotic patients, genes involved in inhibition of the anabolic WNT signalling pathway and those associated with stimulation of bone resorption were significantly upregulated. Apart from these changes, no significant differences were found for the other cell surface antigens, adipogenic, osteogenic and chondrogenic differentiation ability as well as proliferation potential. These findings supported the theory of an influence of CD274 on the regulation of bone metabolism. CD274 might be a promising target for further investigations of the pathogenesis of osteoporosis and of cell-based therapies involving MSCs.

scholarly journals Osteogenic differentiation potential and marker gene expression of different porcine bone marrow mesenchymal stem cell subpopulations selected in different basal media

2020 ◽  
Author(s):  
Sangeetha Kannan ◽  
Jyotirmoy Ghosh ◽  
Sujoy K. Dhara
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Marker Gene ◽  
Microscopical Examination ◽  
Alizarin Red ◽  
Differentiation Potential ◽  
Basal Media ◽  
Selection Of

AbstractMultipotent porcine mesenchymal stem cells (pMSC) are indispensable for research and therapeutic use. Derivation and culture media might affect the selection of MSC subpopulation and thus the differentiation potential of cells. In this study we evaluated the effects of αMEM, aDMEM, M199, αMEM/M199, aDMEM/M199 and αMEM/aDMEM media on porcine bone marrow MSC derivation; pre-differentiation expression of ALP, COL1A1, SPP1 and BGLAP osteogenic marker genes at passage 5 and 10 pMSC; and differentiation potential of passage 5 pMSC. Morphological changes and matrix formation in osteogenic cells were evaluated by microscopical examination and calcium deposit in osteocytes was confirmed by Alizarin Red S staining. Results indicated media independent selection of different bone marrow MSC subpopulations with different surface marker gene expressions. Many pMSC subpopulations in different media had CD14+ expressing cells. We also observed basal media dependent changes in osteogenic markers expression and differentiation potential of pMSC. The αMEM/aDMEM media grown pMSC showed best osteogenic differentiation potential. We thus recommended the testing of αMEM/aDMEM mixed media in other species for pre-differentiation MSC culture that are intended for better osteogenic differentiation.SummaryPre-differentiation basal media influence osteogenic differentiation potential of mesenchymal stem cells (MSC). Among the tested media, αMEM/aDMEM was the best for pre-differentiation porcine MSC culture intending to use in osteogenesis.

Differential Expression of MicroRNA-3148 in Patients with Osteoporosis and Its Impacts on the Osteogenic Differentiation of Rat Bone Marrow Mesenchymal Stem Cells

2021 ◽  
Vol 11 (5) ◽  
pp. 957-962
Author(s):  
Ainiwaerjiang Damaola ◽  
Maerdan Aierken ◽  
Mieralimu Muertizha ◽  
Abudouaini Abudoureheman ◽  
Haishan Lin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Serum Samples ◽  
Alizarin Red ◽  
Differentiation Potential ◽  
Qrt Pcr ◽  
Marrow Mesenchymal Stem Cells ◽  
Rat Bone

We aimed to explore the effects of rat bone marrow mesenchymal stem cells (BMSCs) on osteogenic differentiation via analyzing miR-3148 expression in patients with osteoporosis. Realtime quantitative PCR was conducted for assessing microRNA-3148 expression. BMSCs from SD rats were transfected with microRNA-3148 mimics and microRNA-3148 inhibitor via liposomal trans-fection method utilizing Lipo2000, followed by analysis of microRNA-3148 level. After 10-days of osteogenic differentiation induction, alkaline phosphatase (ALP) staining and alizarin red (ARS) staining were done to investigate the osteogenic differentiation potential. Simultaneously, qRT-PCR measured the expression of osteogenesis marker genes (BMP and Runx2) in each group. qRT-PCR analysis revealed a high expression of miR-3148 in the bone tissue and the serum samples from patients with osteoporosis in comparison with healthy individuals. In addition, miRNA-3148 mimics could retard the osteogenic differentiation of BMSCs, while microRNA-3148 inhibitor could prompt the procedure. MicroRNA-3148 was highly expressed in the skeletal tissues and the serum samples from patients with osteoporosis and it could restrain the differentiation of BMSCs into osteoblasts, suggesting that it might be a novel therapeutic target for treating osteoporosis.

scholarly journals Osteogenic differentiation potential of porcine bone marrow mesenchymal stem cell subpopulations selected in different basal media

Biology Open ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. bio053280
Author(s):  
Sangeetha Kannan ◽  
Jyotirmoy Ghosh ◽  
Sujoy K. Dhara
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Alizarin Red ◽  
Differentiation Potential ◽  
Osteogenic Lineage ◽  
Osteogenic Markers ◽  
Basal Media ◽  
Selection Of

ABSTRACTMultipotent porcine mesenchymal stem cells (pMSC) are invaluable for research and therapeutic use in regenerative medicine. Media used for derivation and expansion of pMSC may play an important role for the selection of MSC subpopulation at an early stage and thereby, the specific basal medium may also affect differentiation potential of these cells. The present study was undertaken to evaluate the effects of αMEM, aDMEM, M199, αMEM/M199, aDMEM/M199 and αMEM/aDMEM media on (1) porcine bone marrow MSC derivation; (2) expression of number of osteogenic markers (ALP, COL1A1, SPP1 and BGLAP) at 5th and 10th passage in pMSC before differentiation; and (3) differentiation of pMSC (at 5th passage) to osteogenic lineage. Morphological changes and matrix formation in osteogenic cells were evaluated by microscopic examination. Calcium deposits in osteocytes were confirmed by Alizarin Red S staining. Based on expression of different markers, it was evident that selection of bone marrow pMSC subpopulations was independent of basal media used. However, the differentiation of those pMSCs, specifically to osteogenic lineage, was dependent on the medium used for expansion of pMSC at the pre-differentiation stage. We demonstrated here that the pMSC grown in combined αMEM/aDMEM (1:1) medium expressed number of osteogenic markers and these pMSC underwent osteogenic differentiation most efficiently, in comparison to porcine mesenchymal stem cells grown in other media. In conclusion, osteogenic differentiation potential of pMSC maintained in αMEM/aDMEM medium was observed significantly higher compared to cells cultivated in other media and therefore, the combined medium αMEM/aDMEM (1:1) may preferentially be used for expansion of pMSC, if needed for osteogenic differentiation.

296 IN VITRO DIFFERENTIATION OF PORCINE BONE MARROW-DERIVED MESENCHYMAL STEM CELLS INTO HEPATOCYTE-LIKE CELLS

2013 ◽  
Vol 25 (1) ◽  
pp. 295
Author(s):  
B. Mohana Kumar ◽  
W. J. Lee ◽  
Y. M. Lee ◽  
R. Patil ◽  
S. L. Lee ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Class Ii ◽  
In Vitro Differentiation ◽  
Rt Pcr ◽  
Hepatic Differentiation ◽  
Alizarin Red ◽  
Differentiation Potential

Mesenchymal stem cells (MSC) are isolated from bone marrow or other tissues, and have properties of self renewal and multilineage differentiation ability. The current study investigated the in vitro differentiation potential of porcine bone marrow derived MSCs into hepatocyte-like cells. The MSC were isolated from the bone marrow of adult miniature pigs (7 months old, T-type, PWG Micro-pig®, PWG Genetics, Seoul, Korea) and adherent cells with fibroblast-like morphology were cultured on plastic. Isolated MSCs were positive for CD29, CD44, CD73, CD90, and vimentin, and negative for CD34, CD45, major histocompatibility complex-class II (MHC-class II), and swine leukocyte antigen-DR (SLA-DR) by flow cytometry analysis. Further, trilineage differentiation of MSC into osteocytes (alkaline phosphatase, von Kossa and Alizarin red), adipocytes (Oil Red O), and chondrocytes (Alcian blue) was confirmed. Differentiation of MSC into hepatocyte-like cells was induced with sequential supplementation of growth factors, cytokines, and hormones for 21 days as described previously (Taléns-Visconti et al. 2006 World J. Gastroenterol. 12, 5834–5845). Morphological analysis, expression of liver-specific markers, and functional assays were performed to evaluate the hepatic differentiation of MSC. Under hepatogenic conditions, MSC acquired cuboidal morphology with cytoplasmic granules. These hepatocyte-like cells expressed α-fetoprotein (AFP), albumin (ALB), cytokeratin 18 (CK18), cytochrome P450 7A1 (CYP7A1), and hepatocyte nuclear factor 1 (HNF-1) markers by immunofluorescence assay. In addition, the expression of selected markers was demonstrated by Western blotting analysis. In accordance with these features, RT-PCR revealed transcripts of AFP, ALB, CK18, CYP7A1, and HNF-1α. Further, the relative expression levels of these transcripts were analysed by quantitative RT-PCR after normalizing to the expression of the endogenous control, glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Data were analysed statistically by one-way ANOVA using PASW statistics 18 (SPSS Inc., Chicago, IL, USA), and significance was considered at P < 0.05. The results showed that the relative expressions of selected marker genes in hepatocyte-like cells were significantly increased compared with that in untreated MSC. The generated hepatocyte-like cells showed glycogen storage as analysed by periodic acid-Schiff (PAS) staining. Moreover, the induced cells produced urea at Day 21 of culture compared with control MSC. In conclusion, our results indicate the potential of porcine MSC to differentiate in vitro into hepatocyte-like cells. Further studies on the functional properties of hepatocyte-like cells are needed to use porcine MSC as an ideal source for liver cell therapy and preclinical drug evaluation. This work was supported by Basic Science Research Program through the National Research Foundation (NRF), funded by the Ministry of Education, Science and Technology (2010-0010528) and the Next-Generation BioGreen 21 Program (No. PJ009021), Rural Development Administration, Republic of Korea.

178 Differential Behavior of Porcine Mesenchymal Stem Cells from Bone Marrow and Adipose During Osteogenic Differentiation on a Glycosaminoglycan Hydrogel Scaffold for Bone and Cartilage Tissue Engineering

2018 ◽  
Vol 30 (1) ◽  
pp. 229 ◽  
Author(s):  
S. A. Womack ◽  
D. J. Milner ◽  
D. W. Weisgerber ◽  
B. A. Harley ◽  
M. B. Wheeler
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Cartilage Tissue Engineering ◽  
Cartilage Tissue ◽  
Scaffold Material ◽  
Alizarin Red ◽  
And Cartilage

The pig is an ideal species for use in tissue engineering studies targeted towards repair of bone and cartilage defects. Novel collagen-glycosaminoglycan hydrogel (CG) scaffolds have shown promise for supporting bone and cartilage growth from mesenchymal stem cells. In order to determine the suitability of these scaffolds for use in porcine model systems for bone and cartilage tissue engineering, we have begun to investigate the behaviour of porcine mesenchymal stem cells on this material. The purpose of this study was to determine whether mesenchymal stem cells from adipose (ASC) and bone marrow (BMSC) form bone on a CG scaffold material. Primary BMSC and ASC from 6-month-old Yorkshire pigs were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) with 10% fetal bovine serum. The ASC and BMSC were then trypsinized at passage 4 or 5 and used to seed ~4-mm-diameter CG scaffolds with 2 million cells/scaffold. Scaffolds were seeded by suspending the cells in medium that had been equilibrated for 30 min, and then placing the CG scaffold into the medium. This method of seeding was determined to be most effective in previous experiments. Scaffolds were then cultured for 7 days in DMEM followed by 21 days in osteogenic media. At the conclusion of the incubation period, the diameter of the scaffolds was measured, and they were fixed with 4% paraformaldehyde and cryosectioned. Then, 10-µm sections were stained with Alizarin Red to assay for mineralization, a hallmark of osteogenic differentiation. Both ASC- and BMSC-loaded scaffolds showed Alizarin Red staining throughout the section after incubation, demonstrating that both undergo osteogenesis on the scaffold material (n = 4). During osteogenic differentiation, scaffolds seeded with both ASC and BMSC showed a decrease in diameter. Unseeded scaffolds showed no decrease in size when in media. The BMSC scaffolds demonstrated a more extensive decrease in size than ASC. The average diameter of ASC loaded scaffolds after differentiation was 2.49 ± 0.39 mm, and that of BMSC-loaded scaffolds was 1.47 ± 0 0.18 mm (n = 3, P < 0.05, Student’s t-test). This suggests a differential ability of ASC and BMSC to break down and metabolize the scaffold matrix, and may indicate that one cell type may be preferable to the other for repairing osteogenic defects using these scaffolds. Current experiments underway will analyse expression of matrix-degrading enzymes to determine the source of the difference between cell types in scaffold shrinkage during differentiation. We will also quantify mineralization in ASC- v. BMSC-loaded scaffolds and assay gene expression of osteogenic markers to determine if there is a difference in osteogenic potential between sources of mesenchymal stem cells on these scaffolds.

scholarly journals Effect of miR-192-5p on Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells in Idiopathic Scoliosis by Regulating Wnt/β-catenin Signaling Pathway via RSPO1

2021 ◽  
Author(s):  
Yifan Yang ◽  
Jing Xu ◽  
Qingxin Su ◽  
Yiran Wu ◽  
Qizheng Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Alkaline Phosphatase ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Signaling Pathway ◽  
Alizarin Red ◽  
Alkaline Phosphatase Staining ◽  
Marrow Mesenchymal Stem Cells ◽  
Related Proteins

Abstract BackgroundIdiopathic scoliosis (IS) is the most common structural scoliosis, which seriously affects not only patient’s physical and mental health but also quality of patient’s life. Abnormal osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) is one of the causes of IS. However, the regulation mechanism of osteogenic differentiation of BMSCs in patients with IS remains to be further studied.MethodsSerum samples of 135 patients with IS were collected, and the expression of miRNA were detected by RT-qPCR. BMSCs from patients with IS were collected and the expression of miR-192-5p in BMSCs from IS patients and normal BMSCs was detected by RT-qPCR. Double luciferase reporter genes assay was used to verify the targeting relationship between miR-192-5p and RSPO1. The levels of RSPO1, osteogenic related proteins (OC, OPN and RUNX2) and Wnt/β-catenin signaling pathway related proteins (WNT3A and β-catenin) were detected by Western blotting. Alkaline phosphatase staining and alizarin red staining were used to evaluate the osteogenesis of BMSCs.ResultsmiR-192-5p was significantly up-regulated in serum and BMSCs of patients with IS. Alkaline phosphatase staining and alizarin red staining showed that miR-192-5p inhibitor promoted the osteogenic differentiation of BMSCs from IS patients. miR-192-5p targeted down-regulated the expression of RSPO1 in BMSCs from IS patients. In addition, overexpression of RSPO1 activated Wnt/β-catenin signaling pathway in BMSCs from IS patients. Furthermore, miR-192-5p/RSPO1 axis regulated levels of osteogenic related proteins (OC, OPN and RUNX2) in BMSCs from IS patients through Wnt/β-catenin signaling pathway, and affected the osteogenic differentiation of BMSCs.ConclusionmiR-192-5p, which was highly expressed in patients with IS, inhibited Wnt/β-catenin signaling pathway by down-regulating RSPO1 protein and then reduced the osteogenic differentiation ability of BMSCs.

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