• The Korean Journal of Physiology and Pharmacology
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• v.16 no.6
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• pp.405-411
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• 2012

The spontaneous axon regeneration of damaged neurons is limited after spinal cord injury (SCI). Recently, mesenchymal stem cell (MSC) transplantation was proposed as a potential approach for enhancing nerve regeneration that avoids the ethical issues associated with embryonic stem cell transplantation. As SCI is a complex pathological entity, the treatment of SCI requires a multipronged approach. The purpose of the present study was to investigate the functional recovery and therapeutic potential of human MSCs (hMSCs) and polymer in a spinal cord hemisection injury model. Rats were subjected to hemisection injuries and then divided into three groups. Two groups of rats underwent partial thoracic hemisection injury followed by implantation of either polymer only or polymer with hMSCs. Another hemisection-only group was used as a control. Behavioral, electrophysiological and immunohistochemical studies were performed on all rats. The functional recovery was significantly improved in the polymer with hMSC-transplanted group as compared with control at five weeks after transplantation. The results of electrophysiologic study demonstrated that the latency of somatosensory-evoked potentials (SSEPs) in the polymer with hMSC-transplanted group was significantly shorter than in the hemisection-only control group. In the results of immunohistochemical study, ${\beta}$-gal-positive cells were observed in the injured and adjacent sites after hMSC transplantation. Surviving hMSCs differentiated into various cell types such as neurons, astrocytes and oligodendrocytes. These data suggest that hMSC transplantation with polymer may play an important role in functional recovery and axonal regeneration after SCI, and may be a potential therapeutic strategy for SCI.

• The Korean Journal of Physiology and Pharmacology
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• v.20 no.6
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• pp.657-667
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• 2016

Critical limb ischemia (CLI) is one of the most severe forms of peripheral artery diseases, but current treatment strategies do not guarantee complete recovery of vascular blood flow or reduce the risk of mortality. Recently, human bone marrow derived mesenchymal stem cells (MSCs) have been reported to have a paracrine influence on angiogenesis in several ischemic diseases. However, little evidence is available regarding optimal cell doses and injection frequencies. Thus, the authors undertook this study to investigate the effects of cell dose and injection frequency on cell survival and paracrine effects. MSCs were injected at $10^6$ or $10^5$ per injection (high and low doses) either once (single injection) or once in two consecutive weeks (double injection) into ischemic legs. Mice were sacrificed 4 weeks after first injection. Angiogenic effects were confirmed in vitro and in vivo, and M2 macrophage infiltration into ischemic tissues and rates of limb salvage were documented. MSCs were found to induce angiogenesis through a paracrine effect in vitro, and were found to survive in ischemic muscle for up to 4 weeks dependent on cell dose and injection frequency. In addition, double high dose and low dose of MSC injections increased vessel formation, and decreased fibrosis volumes and apoptotic cell numbers, whereas a single high dose did not. Our results showed MSCs protect against ischemic injury in a paracrine manner, and suggest that increasing injection frequency is more important than MSC dosage for the treatment CLI.

• Journal of Korean Neurosurgical Society
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• v.62 no.2
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• pp.153-165
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• 2019

Objective : Spinal cord injury (SCI) is a very serious health problem, usually caused by a trauma and accompanied by elevated levels of inflammation indicators. Stem cell-based therapy is promising some valuable strategies for its functional recovery. Nestin-positive progenitor and/or stem cells (SC) isolated from pancreatic islets (PI) show mesenchymal stem cell (MSC) characteristics. For this reason, we aimed to analyze the effects of rat pancreatic islet derived stem cell (rPI-SC) delivery on functional recovery, as well as the levels of inflammation factors following SCI. Methods : rPI-SCs were isolated, cultured and their MSC characteristics were determined through flow cytometry and immunofluorescence analysis. The experimental rat population was divided into three groups : 1) laminectomy & trauma, 2) laminectomy & trauma & phosphate-buffered saline (PBS), and 3) laminectomy+trauma+SCs. Green fluorescent protein (GFP) labelled rPI-SCs were transplanted into the injured rat spinal cord. Their motilities were evaluated with Basso, Beattie and Bresnahan (BBB) Score. After 4-weeks, spinal cord sections were analyzed for GFP labeled SCs and stained for vimentin, $S100{\beta}$, brain derived neurotrophic factor (BDNF), 2',3'-cyclic-nucleotide 3'-phosphodiesterase (CNPase), vascular endothelial growth factor (VEGF) and proinflammatory (interleukin [IL]-6, transforming growth factor $[TGF]-{\beta}$, macrophage inflammatory protein [MIP]-2, myeloperoxidase [MPO]) and anti-inflammatory (IL-1 receptor antagonis) factors. Results : rPI-SCs were revealed to display MSC characteristics and express neural and glial cell markers including BDNF, glial fibrillary acidic protein (GFAP), fibronectin, microtubule associated protein-2a,b (MAP2a,b), ${\beta}3$-tubulin and nestin as well as anti-inflammatory prostaglandin E2 receptor, EP3. The BBB scores showed significant motor recovery in group 3. GFP-labelled cells were localized on the injury site. In addition, decreased proinflammatory factor levels and increased intensity of anti-inflammatory factors were determined. Conclusion : Transplantation of PI-SCs might be an effective strategy to improve functional recovery following spinal cord trauma.

• International Journal of Molecular Medicine
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• v.45 no.2
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• pp.678-686
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• 2020

Use of bone marrow aspirate (BMA) is a clinically advantageous cell therapeutic that bypasses the need for elaborate ex vivo cell culturing. However, a low level of bone marrow-mesenchymal stem cells (BM-MSCs) in the BMA and weak survival rate of these cells post-transplantation entails an insufficient efficacy in vivo. Moreover, stem cell activity in BMA is impaired by age or background diseases. Thus, in order to enrich the BM-MSC pool and improve cell survival, novel cell preconditioning technologies are required. In this study, it has been revealed that the pretreatment of repetitive electromagnetic stimulation (rEMS) is capable of enhancing fibroblastic colony-forming units and cell proliferation in the BM-MSCs, possibly via transient nitric oxide production and extracellular signal regulated kinase 1/2 activation. Notably, this effect was more apparent in stem cells isolated from older patients than from young patients. Furthermore, the rEMS-pretreated cells showed ~53% higher cell survival, compared with the untreated cells, after cell transplantation in mice with no signs of tumorigenesis. Collectively, transient rEMS preconditioning could be utilized to enhance the activity of stem cells and thus, application of rEMS preconditioning to stem cells isolated from older patients is expected to improve the therapeutic effect of stem cells.

• International Journal of Oral Biology
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• v.43 no.2
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• pp.77-82
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• 2018

The mesenchymal stem cells (MSCs) that reside in dental tissues hold a great potential for future applications in regenerative dentistry. In this study, we used human dental pulp cells, isolated from the molars (DPCs), in order to establish the organoid culture. DPCs were established after growing pulp cells in an MSC expansion media (MSC-EM). DPCs were subjected to organoid growth media (OGM) in comparison with human dental pulp stem cells (DPSCs). Inside the extracellular matrix in the OGM, the DPCs and DPSCs readily formed vessel-like structures, which were not observed in the MSC-EM. Immunocytochemistry analysis and flow cytometry analysis showed the elevated expression of CD31 in the DPCs and DPSCs cultured in the OGM. These results suggest endothelial cell-prone differentiation of the DPCs and DPSCs in organoid culture condition.

• Journal of Embryo Transfer
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• v.33 no.2
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• pp.85-97
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• 2018

The trans-differentiation potential of mesenchymal stem cells (MSCs) is employed, but there is little understanding of the cell source-dependent trans-differentiation potential of MSCs into corneal epithelial cells. In the present study, we induced trans-differentiation of MSCs derived from umbilical cord matrix (UCM-MSCs) and from dental tissue (D-MSCs), and we comparatively evaluated the in vitro trans-differentiation properties of both MSCs into corneal epithelial-like cells. Specific cell surface markers of MSC (CD44, CD73, CD90, and CD105) were detected in both UCM-MSCs and D-MSCs, but MHCII and CD119 were significantly lower (P < 0.05) in UCM-MSCs than in D-MSCs. In UCM-MSCs, not only expression levels of Oct3/4 and Nanog but also proliferation ability were significantly higher (P < 0.05) than in D-MSCs. In vitro differentiation abilities into adipocytes and osteocytes were confirmed for both MSCs. UCM-MSCs and D-MSCs were successfully trans-differentiated into corneal epithelial cells, and expression of lineage-specific markers (Cytokeratin-3, -8, and -12) were confirmed in both MSCs using immunofluorescence staining and qRT-PCR analysis. In particular, the differentiation capacity of UCM-MSCs into corneal epithelial cells was significantly higher (P < 0.05) than that of D-MSCs. In conclusion, UCM-MSCs have higher differentiation potential into corneal epithelial-like cells and have lower expression of CD119 and MHC class II than D-MSCs, which makes them a better source for the treatment of corneal opacity.

• Biomolecules & Therapeutics
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• v.30 no.1
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• pp.28-37
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• 2022

Treatment options for patients with chronic kidney disease (CKD) are currently limited; therefore, there has been significant interest in applying mesenchymal stem/stromal cell (MSC)-based therapy to treat CKD. However, MSCs harvested from CKD patients tend to show diminished viability and proliferation due to sustained exposure to uremic toxins in the CKD environment, which limits their utility for cell therapy. The application of melatonin has been demonstrated to improve the therapeutic efficacy of MSCs derived from and engrafted to tissues in patients suffering from CKD, although the underlying biological mechanism has not been elucidated. In this study, we observed overexpression of hexokinase-2 (HK2) in serum samples of CKD patients and MSCs harvested from an adenine-fed CKD mouse model (CKD-mMSCs). HK2 upregulation led to increased production levels of methylglyoxal (MG), a toxic metabolic intermediate of abnormal glycolytic processes. The overabundance of HK2 and MG was associated with impaired mitochondrial function and low cell proliferation in CKD-mMSCs. Melatonin treatment inhibited the increases in HK2 and MG levels, and further improved mitochondrial function, glycolytic metabolism, and cell proliferation. Our findings suggest that identifying and characterizing metabolic regulators such as HK2 in CKD may improve the efficacy of MSCs for treating CKD and other kidney disorders.

• Nuclear Medicine and Molecular Imaging
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• v.42 no.5
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• pp.394-400
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• 2008

Purpose: Quantitative comparison of transgene expression within stem cells between lentivirus and adenovirus-mediated delivery systems has not been reported. Here, we evaluated the human sodium iodide symporter (hNIS) gene expression in rat mesenchymal stem cell (rMSC) transduced by lentivirus or adenovirus, and compared the hNIS expression quantitatively between the two delivery systems. Materials and Methods: Lentiviral-mediated hNIS expressing rMSC (lenti-hNIS-rMSC) was constructed by cloning hNIS gene into pLenti6/UbC/V5-DEST (Invitrogen) to obtain pLenti-hNIS, transducing rMSC with the pLenti-hNIS, and selecting with blasticidin for 3 weeks. Recombinant adenovirus expressing hNIS gene (Rad-hNIS) was produced by homologous recombination and transduction efficiency of Rad-hNIS into rMSC evaluated by Rad-GFP was $19.1{\pm}4.7%$, $54.0{\pm}6.4%$, $85.7{\pm}8.7%$, and $98.4{\pm}1.3%$ at MOI 1, 5, 20, and 100, respectively. The hNIS expressions in lenti-hNIS-rMSC or adeno-hNIS-rMSC were assessed by immunocytochemistry, western blot, and 1-125 uptake. Results: Immunocytochemistry and western blot analyses revealed that hNIS expressions in lenti-hNIS-rMSC were greater than those in adeno-hNIS-rMSC at MOI 20 but lower than at MOI 50. However in vitro 1-125 uptake test demonstrated that iodide uptake in lenti-hNIS-rMSC ($29,704{\pm}6,659\; picomole/10^6\;cells$) was greater than that in adeno-hNIS-rMSC at MOI 100 ($6,168{\pm}2,134\;picomole/10^6\;cells$). Conclusion: Despite lower amount of expressed protein, hNIS function in rMSC was greater by lentivirus than by adenovirus mediated expression. Stem cell tracking using hNIS as a reporter gene should be conducted in consideration of relative vector efficiency for transgene expression.

• Biomolecules & Therapeutics
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• v.23 no.6
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• pp.517-524
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• 2015

Human mesenchymal stem cells (MSCs) have been used in cell-based therapy to promote revascularization after peripheral or myocardial ischemia. High levels of reactive oxygen species (ROS) are involved in the senescence and apoptosis of MSCs, causing defective neovascularization. Here, we examined the effect of the natural antioxidant lycopene on oxidative stress-induced apoptosis in MSCs. Although $H_2O_2$ ($200{\mu}M$) increased intracellular ROS levels in human MSCs, lycopene ($10{\mu}M$) pretreatment suppressed $H_2O_2$-induced ROS generation and increased survival. $H_2O_2$-induced ROS increased the levels of phosphorylated p38 mitogen activated protein kinase (MAPK), Jun-N-terminal kinase (JNK), ataxia telangiectasia mutated (ATM), and p53, which were inhibited by lycopene pretreatment. Furthermore, lycopene pretreatment decreased the expression of cleaved poly (ADP ribose) polymerase-1 (PARP-1) and caspase-3 and increased the expression of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax), which were induced by $H_2O_2$ treatment. Moreover, lycopene significantly increased manganese superoxide dismutase (MnSOD) expression and decreased cellular ROS levels via the PI3K-Akt pathway. Our findings show that lycopene pretreatment prevents ischemic injury by suppressing apoptosis-associated signal pathway and enhancing anti-oxidant protein, suggesting that lycopene could be developed as a beneficial broad-spectrum agent for the successful MSC transplantation in ischemic diseases.

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