• Proceedings of the KSAR Conference
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• 2002.06a
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• pp.4-4
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• 2002

Male germ cell apoptosis has been extensively explored in rodent. In contrast, very little is known about their susceptibility to apoptosis stimuli of developing germ cell stages at the time when germ cell depletion after busulfan treatment occurs. Furthermore, it is still unanswered how spermatogonial stem cells are resistant to busulfan treatment. Spontaneous apoptosis of germ cells was observed in the testis of adult mice and experimentally induced busulfan treated mice increased this apoptosis to such an extent that there was a decrease in the weight of the testis. (omitted)

• International Journal of Oral Biology
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• v.34 no.4
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• pp.177-183
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• 2009

Human mesenchymal stem cell (hMSCs) isolated from human adult bone marrow have self-renewal capacity and can differentiate into multiple cell types in vitro and in vivo. A number of studies have now demonstrated that MSCs can differentiate into various neuronal populations. Due to their autologous characteristics, replacement therapy using MSCs is considered to be safe and does not involve immunological complications. The basic helix-loop-helix (bHLH) transcription factor Olig2 is necessary for the specification of both oligodendrocytes and motor neurons during vertebrate embryogenesis. To develop an efficient method for inducing neuronal differentiation from MSCs, we attempted to optimize the culture conditions and combination with Olig2 gene overexpression. We observed neuron-like morphological changes in the hMSCs under these induction conditions and examined neuronal marker expression in these cells by RTPCR and immunocytochemistry. Our data demonstrate that the combination of Olig2 overexpression and neuron-specific conditioned medium facilitates the neuronal differentiation of hMSCs in vitro. These results will advance the development of an efficient stem cell-mediated cell therapy for human neurodegenerative diseases.

• Archives of Plastic Surgery
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• v.44 no.5
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• pp.370-377
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• 2017

Background Composite grafts are frequently used for facial reconstruction. However, the unpredictability of the results and difficulties with large defects are disadvantages. Adipose-derived stem cells (ADSCs) express several cytokines, and increase the survival of random flaps and fat grafts owing to their angiogenic potential. Methods This study investigated composite graft survival after ADSC injection. Circular chondrocutaneous composite tissues, 2 cm in diameter, from 15 New Zealand white rabbits were used. Thirty ears were randomly divided into 3 groups. In the experimental groups (1 and 2), ADSCs were subcutaneously injected 7 days and immediately before the operation, respectively. Similarly, phosphate-buffered saline was injected in the control group just before surgery in the same manner as in group 2. In all groups, chondrocutaneous composite tissue was elevated, rotated 90 degrees, and repaired in its original position. Skin flow was assessed using laser Doppler 1, 3, 6, 9, and 12 days after surgery. At 1 and 12 days after surgery, the viable area was assessed using digital photography; the rabbits were euthanized, and immunohistochemical staining for CD31 was performed to assess neovascularization. Results The survival of composite grafts increased significantly with the injection of ADSCs (P<0.05). ADSC injection significantly improved neovascularization based on anti-CD31 immunohistochemical analysis and vascular endothelial growth factor expression (P<0.05) in both group 1 and group 2 compared to the control group. No statistically significant differences in graft survival, anti-CD31 neovascularization, or microcirculation were found between groups 1 and 2. Conclusions Treatment with ADSCs improved the composite graft survival, as confirmed by the survival area and histological evaluation. The differences according to the injection timing were not significant.

• Journal of Life Science
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• v.26 no.12
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• pp.1355-1359
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• 2016

Human bone marrow mesenchymal stem cells (hBM-MSCs) can differentiate into various cell types including osteoblasts, adipocytes, chondrocytes, and myocytes. Previous studies, including our own, have shown that MSCs can also differentiate into neuron-like cells. However, their rate of neuronal differentiation is not sufficient for application to stem cell therapy, which requires well-defined cell types. For this purpose, we first examined the expression of neuronal lineage markers (GFAP, MAP-2, KCNH1, Nestin, NF-M, and Tuj-1) by real-time PCR, western blot, and immunocytochemical staining. The expressions of the astrocyte marker GFAP and neuronal markers NF-M and Tuj-1 increased in neuronal differentiated MSCs (dMSCs). To improve the neuronal differentiation efficiency, PDE4, an important signaling intermediator in the progression of neuronal differentiation, was modulated using well-known inhibitors such as rolipram or resveratrol and then differentiated into neuronal cells (Roli- or RSV-dMSCs). The expressions of NF-M, Tuj-1 were increased while that of GFAP decreased in Roli- and RSV-dMSCs, which were examined by real-time PCR, western blot, and immunocytochemical staining. From these experiments, we have found that the neuronal differentiation efficiency can be ameliorated by the modulation of PDE4 activity.

• Journal of Life Science
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• v.26 no.1
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• pp.8-16
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• 2016

Reactive oxygen species (ROS), at appropriate concentrations, mediate various normal cellular functions, including defense against pathogens, signal transduction, cellular growth, and gene expression. A recent study demonstrated that ROS and ROS-generating NADPH oxidase (Nox) are important in self-renewal and neuronal differentiation of subventricular zone (SVZ) neural stem cells in adult mouse brains. In this study, we found that endogenous ROS were detected in SVZ neural stem cells cultured from postnatal mouse brains. Nox4 was predominantly expressed in cultured cells, while the levels of the Nox1 and Nox2 transcripts were very low. In addition, the Nox4 gene was highly upregulated (by up to 10-fold) during neuronal differentiation. Immunocytochemical analysis detected the Nox4 protein mainly in neurons positive for the neuronal specific tubulin Tuj1. After differentiation, endogenous ROS were detected exclusively in neuron-like cells with processes. In addition, perturbation of the cellular redox state with N-acetyl cysteine, a ROS scavenger, during neuronal differentiation greatly inhibited neurogenesis. Lastly, knockdown of Nox4 using short hairpin RNA decreased neurogenesis. These findings suggest that Nox4 may be a major ROS-generating enzyme in postnatal SVZ neural stem cells, and Nox4-mediated ROS generation may be important in their neuronal differentiation.

• Molecules and Cells
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• v.42 no.2
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• pp.104-112
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• 2019

Tracking the fate of individual cells and their progeny through lineage tracing has been widely used to investigate various biological processes including embryonic development, homeostatic tissue turnover, and stem cell function in regeneration and disease. Conventional lineage tracing involves the marking of cells either with dyes or nucleoside analogues or genetic marking with fluorescent and/or colorimetric protein reporters. Both are imaging-based approaches that have played a crucial role in the field of developmental biology as well as adult stem cell biology. However, imaging-based lineage tracing approaches are limited by their scalability and the lack of molecular information underlying fate transitions. Recently, computational biology approaches have been combined with diverse tracing methods to overcome these limitations and so provide high-order scalability and a wealth of molecular information. In this review, we will introduce such novel computational methods, starting from single-cell RNA sequencing-based lineage analysis to DNA barcoding or genetic scar analysis. These novel approaches are complementary to conventional imaging-based approaches and enable us to study the lineage relationships of numerous cell types during vertebrate, and in particular human, development and disease.

• BMB Reports
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• v.53 no.6
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• pp.329-334
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• 2020

Inflammasomes are cytosolic, multiprotein complexes that act at the frontline of the immune responses by recognizing pathogen- or danger-associated molecular patterns or abnormal host molecules. Mesenchymal stem cells (MSCs) have been reported to possess multipotency to differentiate into various cell types and immunoregulatory effects. In this study, we investigated the expression and functional regulation of NLR Family Pyrin Domain Containing 3 (NLRP3) inflammasome in human umbilical cord blood-derived MSCs (hUCB-MSCs). hUCB-MSCs expressed inflammasome components that are necessary for its complex assembly. Interestingly, NLRP3 inflammasome activation suppressed the differentiation of hUCB-MSCs into osteoblasts, which was restored when the expression of adaptor proteins for inflammasome assembly was inhibited. Moreover, the suppressive effects of MSCs on T cell responses and the macrophage activation were augmented in response to NLRP3 activation. In vivo studies using colitic mice revealed that the protective abilities of hUCB-MSCs increased after NLRP3 stimulation. In conclusion, our findings suggest that the NLRP3 inflammasome components are expressed in hUCB-MSCs and its activation can regulate the differentiation capability and the immunomodulatory effects of hUCB-MSCs.

• Journal of Animal Reproduction and Biotechnology
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• v.35 no.1
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• pp.65-72
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• 2020

Fish ovarian germline stem cells (OGSCs) that have the abilities to self-renew and differentiate into functional gametes can be used in various researches and applications. A main issue to be solved for effective utilization of fish OGSCs is the development of their stable in vitro culture condition, but only few researches about fish OGSC culture have been reported so far. In this study, in order to find the clues to develop the culture condition for OGSCs from Japanese medaka (Oryzias latipes), we tried to establish somatic cell lines as a candidate for the feeder cells and evaluated its supporting effects on the culture of ovarian cell populations from O. latipes. As the results, the somatic cell lines could be established only from the embryonic tissues among three tissues derived from embryos, fins and ovaries. Three embryonic cell lines were tested as a feeder cell for the culture of ovarian cell population and all three cell lines induced cell aggregation formation of the cultured ovarian cells whereas the feeder-free condition did not. Furthermore, a significant cellular proliferation was observed in the ovarian cells cultured on two of three cell lines. As a trial to increase the capacity of the cell lines as a feeder cell that supports the proliferation of the cultured ovarian cells, we subsequently established a stable line that expresses the foreign O. latipes fibroblast growth factor 2 (FGF2) from an embryonic cell line and evaluated its effectiveness as a feeder cell. The ovarian cells cultured on FGF2 expressing feeder cells still formed cell aggregates but did not show a significant increase in cellular proliferation compared to those cultured on non-transformed feeder cells. The results from this study will provide the fundamental information for in vitro culture of medaka OGSCs.

• Anatomy and Cell Biology
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• v.56 no.2
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• pp.219-227
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• 2023

Adult neurogenesis has been reported in the hypothalamus, subventricular zone and subgranular zone in the hippocamp. Recent studies indicated that new cells in the hypothalamus are affected by diet. We previously showed beneficial effects of safflower seed oil (SSO), a rich source of linoleic acid (LA; 74%), on proliferation and differentiation of neural stem cells (NSCs) in vitro. In this study, the effect of SSO on hypothalamic neurogenesis was investigated in vivo, in comparison to synthetic LA. Adult mice were treated with SSO (400 mg/kg) and pure synthetic LA (300 mg/kg), at similar concentrations of LA, for 8 weeks and then hypothalamic NSCs were cultured and subsequently used for Neurosphere-forming assay. In addition, serum levels of brain-derived neurotrophic factor (BNDF) were measured using enzyme-linked immunosorbent assay. Administration of SSO for 8 weeks in adult mice promoted the proliferation of NSCs isolated from SSO-treated mice. Immunofluorescence staining of the hypothalamus showed that the frequency of astrocytes (glial fibrillary acidic protein⁺ cells) are not affected by LA or SSO. However, the frequency of immature (doublecortin⁺ cells) and mature (neuronal nuclei⁺ cells) neurons significantly increased in LA- and SSO-treated mice, compared to vehicle. Furthermore, both LA and SSO caused a significant increase in the serum levels of BDNF. Importantly, SSO acted more potently than LA in all experiments. The presence of other fatty acids in SSO, such as oleic acid and palmitic acid, suggests that they could be responsible for SSO positive effect on hypothalamic proliferation and neurogenesis, compared to synthetic LA at similar concentrations.

• Journal of Life Science
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• v.28 no.12
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• pp.1455-1460
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• 2018

Due to a rapidly expanding aging population, the incidence of degenerative bone disease has increased, and efforts to handle the issue using regenerative medicine have become more important. In order to control various bone diseases such as osteoarthritis and osteoporosis, regenerative medicine utilizing adult stem cells has been extensively studied. And it is now clear that the mitochondrial energy metabolism, oxidative phosphorylation, is important for the process of stem cell differentiation. Interestingly, a recent study reported that salicylate promotes mitochondrial biogenesis by regulating the expression of $PGC-1{\alpha}$ in murine cells. However, the possible effects of salicylate on osteogenic differentiation through increased mitochondrial biogenesis in stem cells remain unknown. Thus, here we investigated whether salicylate could influence osteogenic differentiation and mitochondrial biogenesis of periosteum-derived mesenchymal stem cells (POMSCs). We found that salicylate treatments of POMSCs undergoing osteogenic differentiation increased the activity of alkaline phosphatase, a well-known early marker of bone cell differentiation. In addition, we observed that mitochondrial mass was increased by salicylate treatments in POMSCs. Together, these results indicate that salicylate can enhance osteogenic differentiation and mitochondrial biogenesis in POMSCs. Therefore, the findings in this study suggest that small molecules augmenting mitochondrial function such as salicylate can be a novel modulator for osteogenic differentiation and regenerative medicine.