Comparative study of osteogenic differentiation potential of mesenchymal stem cells derived from bone marrow and adipose tissue of osteoporotic female rats

AbstractMesenchymal stem cells (MSCs) are important in regenerative medicine because of their potential for multi-differentiation. Bone marrow, chorion and placenta have all been suggested as potential sources for clinical application. However, the osteogenic differentiation potential of MSCs derived from chorion or placenta is not very efficient. Bone morphogenetic protein-2 (BMP-2) plays an important role in bone development. Its effect on osteogenic augmentation has been addressed in several studies. Recent studies have also shown a relationship between miRNAs and osteogenesis. We hypothesized that miRNAs targeted to Runt-related transcription factor 2 (Runx-2), a major transcription factor of osteogenesis, are responsible for regulating the differentiation of MSCs into osteoblasts. This study examines the effect of BMP-2 on the osteogenic differentiation of MSCs isolated from chorion and placenta in comparison to bone marrow-derived MSCs and investigates the role of miRNAs in the osteogenic differentiation of MSCs from these sources. MSCs were isolated from human bone marrow, chorion and placenta. The osteogenic differentiation potential after BMP-2 treatment was examined using ALP staining, ALP activity assay, and osteogenic gene expression. Candidate miRNAs were selected and their expression levels during osteoblastic differentiation were examined using real-time RT-PCR. The role of these miRNAs in osteogenesis was investigated by transfection with specific miRNA inhibitors. The level of osteogenic differentiation was monitored after anti-miRNA treatment. MSCs isolated from chorion and placenta exhibited self-renewal capacity and multi-lineage differentiation potential similar to MSCs isolated from bone marrow. BMP-2 treated MSCs showed higher ALP levels and osteogenic gene expression compared to untreated MSCs. All investigated miRNAs (miR-31, miR-106a and miR148) were consistently downregulated during the process of osteogenic differentiation. After treatment with miRNA inhibitors, ALP activity and osteogenic gene expression increased over the time of osteogenic differentiation. BMP-2 has a positive effect on osteogenic differentiation of chorion- and placenta-derived MSCs. The inhibition of specific miRNAs enhanced the osteogenic differentiation capacity of various MSCs in culture and this strategy might be used to promote bone regeneration. However, further in vivo experiments are required to assess the validity of this approach.

Download Full-text

Osteogenic differentiation of adipose tissue-derived mesenchymal stem cells cultured with different concentrations of prolactin

Arquivo Brasileiro de Medicina Veterinária e Zootecnia ◽

10.1590/1678-4162-9364 ◽

2017 ◽

Vol 69 (6) ◽

pp. 1573-1580

Author(s):

K.P. Oliveira ◽

A.M.S. Reis ◽

A.P. Silva ◽

C.L.R. Silva ◽

A.M. Goes ◽

...

Keyword(s):

Stem Cells ◽

Alkaline Phosphatase ◽

Mesenchymal Stem Cells ◽

Adipose Tissue ◽

Osteogenic Differentiation ◽

Bone Sialoprotein ◽

Collagen I ◽

Female Rats ◽

Osteogenic Potential ◽

Vitro Effect

ABSTRACT The objective was to evaluate the in vitro effect of prolactin in osteogenic potential of adipose tissue-derived mesenchymal stem cells (ADSCs) in female rats. ADSCs were cultured in osteogenic medium with and without the addition of prolactin and distributed into three groups: 1) ADSCs (control), 2) ADSCs with addition of 100ng/mL of prolactin and 3) ADSCs with addition of 300ng/mL of prolactin. At 21 days of differentiation, the tests of MTT conversion into formazan crystals, percentage of mineralized nodules and cells per field and quantification of genic transcript for alkaline phosphatase, osteopontin, osteocalcin, bone sialoprotein, BMP-2 and collagen I by real-time RT-PCR were made. The addition of prolactin reduced the conversion of MTT in group 3 and increased the percentage of cells per field in the groups 2 and 3, however without significantly increasing the percentage of mineralized nodules and the expression of alkaline phosphatase, osteopontin, osteocalcin, bone sialoprotein, BMP-2 and collagen I. In conclusion, the addition of prolactin in concentrations of 100ng/mL and 300ng/mL does not change the osteogenic differentiation to the ADSCs of female rats despite increase in the cellularity of the culture.

Download Full-text

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

10.1101/2020.04.27.063230 ◽

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.

Download Full-text

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

Journal of Biomaterials and Tissue Engineering ◽

10.1166/jbt.2021.2635 ◽

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.

Download Full-text

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

Biology Open ◽

10.1242/bio.053280 ◽

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.

Download Full-text

Biological Characteristics and Osteogenic Differentiation of Ovine Bone Marrow Derived Mesenchymal Stem Cells Stimulated with FGF-2 and BMP-2

International Journal of Molecular Sciences ◽

10.3390/ijms21249726 ◽

2020 ◽

Vol 21 (24) ◽

pp. 9726

Author(s):

Sandra Gromolak ◽

Agnieszka Krawczenko ◽

Agnieszka Antończyk ◽

Krzysztof Buczak ◽

Zdzisław Kiełbowicz ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Collagen Type ◽

Collagen Type I ◽

Biological Characteristics ◽

Immunofluorescent Staining ◽

Type I ◽

Differentiation Potential

Cell-based therapies using mesenchymal stem cells (MSCs) are a promising tool in bone tissue engineering. Bone regeneration with MSCs involves a series of molecular processes leading to the activation of the osteoinductive cascade supported by bioactive factors, including fibroblast growth factor-2 (FGF-2) and bone morphogenetic protein-2 (BMP-2). In this study, we examined the biological characteristics and osteogenic differentiation potential of sheep bone marrow MSCs (BM-MSCs) treated with 20 ng/mL of FGF-2 and 100 ng/mL BMP-2 in vitro. The biological properties of osteogenic-induced BM-MSCs were investigated by assessing their morphology, proliferation, phenotype, and cytokine secretory profile. The osteogenic differentiation was characterized by Alizarin Red S staining, immunofluorescent staining of osteocalcin and collagen type I, and expression levels of genetic markers of osteogenesis. The results demonstrated that BM-MSCs treated with FGF-2 and BMP-2 maintained their primary MSC properties and improved their osteogenic differentiation capacity, as confirmed by increased expression of osteocalcin and collagen type I and upregulation of osteogenic-related gene markers BMP-2, Runx2, osterix, collagen type I, osteocalcin, and osteopontin. Furthermore, sheep BM-MSCs produced a variety of bioactive factors involved in osteogenesis, and supplementation of the culture medium with FGF-2 and BMP-2 affected the secretome profile of the cells. The results suggest that sheep osteogenic-induced BM-MSCs may be used as a cellular therapy to study bone repair in the preclinical large animal model.

Download Full-text