• Journal of Life Science
• /
• v.24 no.11
• /
• pp.1238-1243
• /
• 2014

Mesenchymal stem cells (MSCs) have been widely used as cellular therapeutic agents. They have their own characteristic stemness, and thus, they can be used in the treatment of many chronic diseases and in anticancer therapy. MSC therapy has many advantages over chemical therapy. MSC therapy is based on self or homogeneous origin; as such, it is expected to be effective in the treatment of various diseases. In addition, microRNAs in particular have been studied for their structure and function, and they are also expected to prove effective for use as therapeutic agents in cancer or chronic diseases. MicroRNAs are largely associated with metabolism and homeostasis. Therefore, over- or under-expression of microRNAs leads to chronic diseases. Conversely, effective control of the expression of specific microRNAs reduces the risk of many chronic diseases. However, there have been no reports thus far on the synergistic effects of MSCs and microRNAs. Therefore, in this study, we examined the relationship between MSCs and microRNAs using placenta-derived MSCs (PDSCs), bone marrow-derived MSCs (BM-MSCs), and fibroblast (WI-38) cells. We studied the expression of some microRNAs in MSCs and compared the expression in each cell line and cell passage. As a result, we found that the expression of microRNA-34a was higher in PDSCs than in BM-MSCs and that the expression of microRNA-27a, 33a, 33b, and 211 was higher in BM-MSCs than in PDSCs. Therefore, we expect that each MSC line will be used as cell therapy, considering its expressed functional microRNA.

• Journal of Microbiology and Biotechnology
• /
• v.18 no.5
• /
• pp.975-982
• /
• 2008

We investigated whether transplantation of osteogenically differentiated bone marrow-derived mesenchymal stem cells (BMMSCs) and the use of an hydroxyapatite (HAp) scaffold can enhance the in vivo bone formation efficacy of human BMMSCs. Three months after implantation to the subcutaneous dorsum of athymic mice, transplantation of osteogenically differentiated human BMMSCs increased the bone formation area and calcium deposition to 7.1- and 6.2-folds, respectively, of those of transplantation of undifferentiated BMMSCs. The use of the HAp scaffold increased the bone formation area and calcium deposition to 3.7- and 3.5-folds, respectively, of those of a polymer scaffold. Moreover, a combination of transplantation of osteogenically differentiated BMMSCs and HAp scaffold further increased the bone formation area and calcium deposition to 10.6- and 9.3-folds, respectively, of those of transplantation of undifferentiated BMMSCs seeded onto polymer scaffolds. The factorial experimental analysis showed that osteogenic differentiation of BMMSCs prior to transplantation has a stronger positive effect than the HAp scaffold on in vivo bone formation.

• Reproductive and Developmental Biology
• /
• v.32 no.1
• /
• pp.21-25
• /
• 2008

Bone marrow mesenchymal stem cells (BMMSCs) have the capacity for self-renewal and differentiation into a variety of cell types. They represent an attractive source of cells for gene and cell therapy. The purpose of this study is to direct the specific expression of the DsRed reporter gene in $Sca-1^+$ BMMSCs differentiated into a cardiomyogenic lineage. We constructed the prMLC-2v-DsRed vector expressing DsRed under the control of the 309 tp fragment of the rat MLC-2v 5'-flanking region. The specific expression of the DsRed reporter gene under the transcriptional control of the 309 bp fragment of the rat MLC-2v promoter was tested in 5-azacytidine healed-$Sca-1^+$ BMMSCs over 2 weeks after the prMLC-2v-DsRed transfection. The prMLC-2v-DsRed was specifically expressed in the $Sca-1^+$ BMMSCs with cardiomyogenic lineage differentiation and it demonstrates that the 309 bp sequences of the rat MLC-2v 5'-flanking region is sufficient to confer cardiac specific expression on a DsRed reporter gene. The cardiac-specific promoter-driven reporter vector provides an important tool for the study of stem cell differentiation and cell replacement therapy in ischemic cardiomyopathy.

• Journal of Animal Science and Technology
• /
• v.57 no.5
• /
• pp.18.1-18.8
• /
• 2015

Background: Quantitative real time reverse transcription PCR (qRT-PCR) is one of the most important techniques for gene-expression analysis in molecular based studies. Selecting a proper internal control gene for normalizing data is a crucial step in gene expression analysis via this method. The expression levels of reference genes should be remained constant among cells in different tissues. However, it seems that the location of cells in different tissues might influence their expression. The purpose of this study was to determine whether the source of mesenchymal stem cells (MSCs) has any effect on expression level of three common reference genes (GAPDH, ${\beta}$-actin and ${\beta}2$-microglobulin) in equine marrow- and adipose-derived undifferentiated MSCs and consequently their reliability for comparative qRT-PCR. Materials and methods: Adipose tissue (AT) and bone marrow (BM) samples were harvested from 3 mares. MSCs were isolated and cultured until passage 3 (P3). Total RNA of P3 cells was extracted for cDNA synthesis. The generated cDNAs were analyzed by quantitative real-time PCR. The PCR reactions were ended with a melting curve analysis to verify the specificity of amplicon. Results: The expression levels of GAPDH were significantly different between AT- and BM-derived MSCs (p < 0.05). Differences in expression level of ${\beta}$-actin (P < 0.001) and B2M (P < 0.006.) between MSCs derived from AT and BM were substantially higher than GAPDH. In addition, the fold change in expression levels of GAPDH, ${\beta}$-actin and B2M in AT-derived MSCs compared to BM-derived MSCs were 2.38, 6.76 and 7.76, respectively. Conclusion: This study demonstrated that GAPDH and especially ${\beta}$-actin and B2M express in different levels in equine AT- and BM-derived MSCs. Thus they cannot be considered as reliable reference genes for comparative quantitative gene expression analysis in MSCs derived from equine bone marrow and adipose tissue.

• IMMUNE NETWORK
• /
• v.22 no.6
• /
• pp.49.1-49.15
• /
• 2022

Exosomes derived from mesenchymal stem cells (MSCs) could protect against myocardial infarction (MI). TLR4 is reported to play an important role in MI, while microRNA-182-5p (miR-182-5p) negatively regulates TLR4 expression. Therefore, we hypothesize that MSCs-derived exosomes overexpressing miR-182-5p may have beneficial effects on MI. We generated bone marrow mesenchymal stem cells (BM-MSCs) and overexpressed miR-182-5p in these cells for exosome isolation. H₂O₂-stimulated neonatal mouse ventricle myocytes (NMVMs) and MI mouse model were employed, which were subjected to exosome treatment. The expression of inflammatory factors, heart function, and TLR4 signaling pathway activation were monitored. It was found that miR-182-5p decreased TLR4 expression in BM-MSCs and NMVMs. Administration of exosomes overexpressing miR-182-5p to H₂O₂-stimulated NMVMs enhanced cell viability and suppressed the expression of inflammatory cytokines. In addition, they promoted heart function, suppressed inflammatory responses, and de-activated TLR4/NF-κB signaling pathway in MI mice. In conclusion, miR-182-5p transferred by the exosomes derived from BM-MSCs protected against MI-induced impairments by targeting TLR4.

• Journal of the Korea Academia-Industrial cooperation Society
• /
• v.16 no.3
• /
• pp.1964-1971
• /
• 2015

Human bone marrow or human umbilical cord vein derived-mesenchymal stem cells (hBM-MSCs or hUC-MSCs) have known as a potentially useful cell type for clinical therapeutic applications. We investigated two-pore domain potassium (K2P) channels in these cells. K2P channels play a major role in setting the resting membrane potential in many cell types. Among them, TREK1 is targets of hydrogen, hypoxia, polyunsaturated fatty acids, antidepressant, and neurotransmitters. We investigated whether hBM-MSCs and hUC-MSCs express functional TREK1 channel using RT-PCR analysis and patch clamp technique. Potassium channel with a single channel conductance of 100 pS was found in hUC-MSCs and BM-MSCs and the channel was activated by membrane stretch (-5 mmHg ~ -15 mmHg), arachidonic acid ($10{\mu}M$) and intracellular acidosis (pH 6.0). These electrophysiological properties were similar to those of TREK1. Our results suggest that TREK1 is functionally present in hBM-MSCs and hUC-MSCs, where they contribute to its resting membrane potential.

• Journal of Korean Neurosurgical Society
• /
• v.46 no.4
• /
• pp.397-402
• /
• 2009

Objective : In this study, the authors assessed the ability of rat bone marrow derived mesenchymal stem cells (BMDMSCs), in the presence of a growth factor, fibroblast growth factor-4 (FGF-4) and hydroxyapatite, to act as a scaffold for posterolateral spinal fusion in a rat model. Methods : Using a rat posterolateral spine fusion model. the experimental study comprised 3 groups. Group 1 was composed of 6 animals that were implanted with 0.08 gram hydroxyapatite only. Group 2 was composed of 6 animals that were implanted with 0.08 gram hydroxyapatite containing $1{\times}10^6/60{\mu}L$ rat of BMDMSCs. Group 3 was composed of 6 animals that were implanted with 0.08 gram hydroxyapatite containing $1{\times}10^6/60{\mu}L$ of rat BMDMSCs and FGF-4 $1{\mu}G$ to induce the bony differentiation of the BMDMSCs. Rats were assessed using radiographs obtained at 4, 6, and 8 weeks postoperatively. After sacrifice, spines were explanted and assessed by manual palpation, high-resolution microcomputerized tomography, and histological analysis. Results : Radiographic, high-resolution microcomputerized tomographic, and manual palpation revealed spinal fusion in five rats (83%) in Group 2 at 8 weeks. However, in Group 1, three (60%) rats developed fusion at L4-L5 by radiography and two (40%) by manual palpation in radiographic examination. In addition, in Group 3, bone fusion was observed in only 50% of rats by manual palpation and radiographic examination at this time. Conclusion : The present study demonstrates that 0.08 gram of hydroxyapatite with $1{\times}10^6/60{\mu}L$ rat of BMDMSCs induced bone fusion. FGF4, added to differentiate primitive $1{\times}10^6/60{\mu}L$ rat of BMDMSCs did not induce fusion. Based on histologic data, FGF-4 appears to induce fibrotic change rather than differentiation to bone by $1{\times}10^6/60{\mu}L$ rat of BMDMSCs.

• Molecules and Cells
• /
• v.24 no.2
• /
• pp.255-260
• /
• 2007

Bone marrow (BM) derived mesenchymal stem cells (MSC) are pluripotent cells which can differentiate into osteogenic, adipogenic and other lineages. In spite of the broad interest, the information about the changes in BM cell composition, in particularly about the variation of MSC number and their properties in relation to the age of the donor is still controversial. The aim of this study was to investigate the age associated changes in variations of BM cell composition, phenotype and differentiation capacities of MSC using a rat model. Cell populations were characterized by flow cytometry using light scattering parameters, DNA content and a set of monoclonal antibodies. Single cell analysis was performed by conventional fluorescent microscopy. In vitro culture of MSC was established and their phenotype and capability for in vitro differentiation into osteogenic and adipogenic cells was shown. Age related changes in tibiae and femurs, amount of BM tissue, BM cell composition, proportions of separated MSC and yield of MSC in 2 weeks of in vitro culture were found. At the same time, neither change in phenotype no in differentiation capacities of MSC was registered. Age-related changes of the number of MSC should be taken into account whenever MSC are intended to be used for investigations.

• International Journal of Stem Cells
• /
• v.15 no.2
• /
• pp.217-226
• /
• 2022

Background and Objectives: Stroke is the most common cause of human death and functional disability, resulting in more than 5 million deaths worldwide each year. Bone marrow mesenchymal stem cells (BMSCs) are a kind of stem cell that are able to self-renew and differentiate into many types of tissues. Therefore, BMSCs have the potential to replace damaged neurons and promote the reconstruction of nerve conduction pathways and connective tissue. However, it remains unknown whether transplanted BMSCs promote angiogenesis or improve the tissue microenvironment directly or indirectly through paracrine interactions. This study aimed to determine the therapeutic effect of BMSCs on ischemic stroke with hypertension in a rodent model and to explore the possible mechanisms underlying any benefits. Methods and Results: Middle cerebral artery occlusion was used to establish the experimental stroke model. The area of cerebral infarction, expression of vascular endothelial growth factor (VEGF) and glial cell line-derived neurotrophic factor (GDNF), and increment of astrocyte were measured by TTC staining, western blot, real-time quantitative polymerase chain reaction (RT-qPCR) and immunocytochemistry. The results showed a smaller area of cerebral infarction and improved neurological function scores in animals treated with BMSCs compared to controls. The results of RT-qPCR and western blot assays showed higher expression of VEGF and GDNF in BMSC-treated animals compared with controls. Our study also showed that one round of BMSCs transplantation significantly promoted the proliferation of subventricular zone and cortical cells, especially astrocytes, on the ischemic side following cerebral ischemia. Conclusions: Above findings support that BMSCs have therapeutic effects for ischemic stroke complicated with hypertension, which may occur via up-regulated expression of VEGF and GDNF and reduction of neuronal apoptosis, thereby promoting the recovery of nerve function.

• IMMUNE NETWORK
• /
• v.14 no.1
• /
• pp.54-65
• /
• 2014

Bone marrow-derived mesenchymal stem cells (MSCs) are multipotent, with the ability to differentiate into different cell types. Additionally, the immunomodulatory activity of MSCs can downregulate inflammatory responses. The use of MSCs to repair injured tissues and treat inflammation, including in neuroimmune diseases, has been extensively explored. Although MSCs have emerged as a promising resource for the treatment of neuroimmune diseases, attempts to define the molecular properties of MSCs have been limited by the heterogeneity of MSC populations. We recently developed a new method, the subfractionation culturing method, to isolate homogeneous human clonal MSCs (hcMSCs). The hcMSCs were able to differentiate into fat, cartilage, bone, neuroglia, and liver cell types. In this study, to better understand the properties of neurally differentiated MSCs, gene expression in highly homogeneous hcMSCs was analyzed. Neural differentiation of hcMSCs was induced for 14 days. Thereafter, RNA and genomic DNA was isolated and subjected to microarray analysis and DNA methylation array analysis, respectively. We correlated the transcriptome of hcMSCs during neural differentiation with the DNA methylation status. Here, we describe and discuss the gene expression profile of neurally differentiated hcMSCs. These findings will expand our understanding of the molecular properties of MSCs and contribute to the development of cell therapy for neuroimmune diseases.