• Development and Reproduction
• /
• v.13 no.3
• /
• pp.173-181
• /
• 2009

One of the most extensively studied populations of multipotent adult stem cells are mesenchymal stem cells (MSCs). MSCs derived from the human umbilical cord vein (HUC-MSCs) are morphologically and immunophenotypically similar to MSCs isolated from bone marrow. HUC-MSCs are multipotent stem cells, differ from hematopoietic stem cells and can be differentiated into neural cells. Since neural tissue has limited intrinsic capacity of repair after injury, the identification of alternate sources of neural stem cells has broad clinical potential. We isolated mesenchymal-like stem cells from the human umbilical cord vein, and studied transdifferentiation-promoting conditions in neural cells. Dopaminergic neuronal differentiation of HUC-MSCs was also studied. Neural differentiation was induced by adding bFGF, EGF, dimethyl sulfoxide (DMSO) and butylated hydroxyanisole (BHA) in N2 medium and N2 supplement. The immunoreactive cells for $\beta$-tubulin III, a neuron-specific marker, GFAP, an astrocyte marker, or Gal-C, an oligodendrocyte marker, were found. HUC-MSCs treated with bFGF, SHH and FGF8 were differentiated into dopaminergic neurons that were immunopositive for tyrosine hydroxylase (TH) antibody. HUC-MSCs treated with DMSO and BHA rapidly showed the morphology of multipolar neurons. Both immunocytochemistry and RT-PCR analysis indicated that the expression of a number of neural markers including NeuroD1, $\beta$-tubulin III, GFAP and nestin was markedly elevated during this acute differentiation. While the stem cell markers such as SCF, C-kit, and Stat-3 were not expressed after neural differentiation, we confirmed the differentiation of dopaminergic neurons by TH/$\beta$-tubulin III positive cells. In conclusion, HUC-MSCs can be differentiated into dopaminergic neurons and these findings suggest that HUC-MSCs are alternative cell source of therapeutic treatment for neurodegenerative diseases.

• The korean journal of orthodontics
• /
• v.31 no.6 s.89
• /
• pp.579-587
• /
• 2001

Cleft palate has been studied with epidemiologic and molecular methods, and many etiologic factors have been examined closely Among the research methods, biologic molecule research has been the most important method for cleft palate formation study The $TGF-\beta$ played an important role in cell migration, epithelial-mesenchymal transdifferentiation, extracellular matrix synthesis and deposition. But there was not much research on the correlation cleft palate induced by beta-aminonitroproprionitrile(BAPN) and $TGF-\beta$ expression. The purpose of the present study was to examine how $TGF-\beta$ is expressed in cleft palate rats. 4 Timed-pregnant Sprague-Dawley rats were obtained on the 10th gestation day. On the 13th day of gestation, BAPN-monofumarate salts (${(C_3H_6N_2)}_2{\cdot}C_4H_4O_4$) were individually, ovally administered to 3 pregnant rats at a ratio of 1g/kg body weight. And 4 pregnant rats were sacrificed on day 20 post coitus (p.c.). The $TGF-\beta$ expression in the cleft formed rats fetuses showed the following patterns : 1. Osteoblast and mesenchymal cells of the cleft pa)ate rats were of low expression compared with those of the control rats. 2. The cleft palate rats didn't show uy difference in the $TGF-\beta$ expression of osteocyte item the control rats. 3. In western blot analysis, the thickness of band of $TGF-\beta$ in the cleft palate rats was thinner and more diluted than that of the control rats.

• Journal of Animal Science and Technology
• /
• v.63 no.4
• /
• pp.934-953
• /
• 2021

Ciglitazone is a member of the thiazolidinedione family, and specifically binds to peroxisome proliferator-activated receptor-γ (PPARγ), thereby promoting adipocyte differentiation. We hypothesized that ciglitazone as a PPARγ ligand in the absence of an adipocyte differentiation cocktail would increase adiponectin and adipogenic gene expression in bovine satellite cells (BSC). Muscle-derived BSCs were isolated from six, 18-month-old Yanbian Yellow Cattle. The BSC were cultured for 96 h in differentiation medium containing 5 µM ciglitazone (CL), 10 µM ciglitazone (CM), or 20 µM ciglitazone (CH). Control (CON) BSC were cultured only in a differentiation medium (containing 2% horse serum). The presence of myogenin, desmin, and paired box 7 (Pax7) proteins was confirmed in the BSC by immunofluorescence staining. The CL, CM, and CH treatments produced higher concentrations of triacylglycerol and lipid droplet accumulation in myotubes than those of the CON treatment. Ciglitazone treatments significantly increased the relative expression of PPARγ, CCAAT/enhancer-binding protein alpha (C/EBPα), C/EBPβ, fatty acid synthase, stearoyl-CoA desaturase, and perilipin 2. Ciglitazone treatments increased gene expression of Pax3 and Pax7 and decreased expression of myogenic differentiation-1, myogenin, myogenic regulatory factor-5, and myogenin-4 (p < 0.01). Adiponectin concentration caused by ciglitazone treatments was significantly greater than CON (p < 0.01). RNA sequencing showed that 281 differentially expressed genes (DEGs) were found in the treatments of ciglitazone. DEGs gene ontology (GO) analysis showed that the top 10 GO enrichment significantly changed the biological processes such as protein trimerization, negative regulation of cell proliferation, adipocytes differentiation, and cellular response to external stimulus. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that DEGs were involved in the p53 signaling pathway, PPAR signaling pathway, biosynthesis of amino acids, tumor necrosis factor signaling pathway, non-alcoholic fatty liver disease, PI3K-Akt signaling pathway, and Wnt signaling pathway. These results indicate that ciglitazone acts as PPARγ agonist, effectively increases the adiponectin concentration and adipogenic gene expression, and stimulates the conversion of BSC to adipocyte-like cells in the absence of adipocyte differentiation cocktail.

• Journal of the korean academy of Pediatric Dentistry
• /
• v.28 no.2
• /
• pp.238-246
• /
• 2001

Bone morphogenetic proteins(BMPs), members of the transforming growth factor $\beta$(TGF-$\beta$) superfamily were first identified as the factors that induce ectopic bone formation in vivo, when implanted into muscular tissue. Especially BMP-2 inhibits terminal differentiation of C2C12 myoblasts and converts them into osteoblast lineage cells. In the molecular mechanism of the signal transduction of TGF-$\beta$ and related factors, intracellular signaling proteins were identified as Smad. In previous study, it has been reported that Smad 1 and Smad 5, which belong to the R-Smad family mediate BMP signaling, were involved in the induction of osteoblast differentiation in C2C12 cells. To understnad the role of Smads involved in osteogenic transdifferentiation in C2C12 cell, in present study, after we stably transfected C2C12 cells with each. Smad(Smad 1,Smad 5) expression vector, cultured for 3 days and stained for alkaline phophatase activity. ALP activity positive cells appeared in the Smad 1, Smad 5 stably transfected cell even in the abscence of BMP. After transiently co-transfected C2C12 cells with each Smad expression vector and ALP promoter, it was examined that Smad 1 and Smad 5 expression vector had increased about 2 fold ALP promoter activity in the abscence of BMP. These result suggested that both Smad 1 and Smad 5 were involved in the intracellular BMP signals which induce osteoblast differentiation in C2C12 cells. The effect of BMP on C2C12 cells with Smad 1, Smad 5 transfected were studied by using northern blot analysis. the treatment of BMP upregulated ALP mRNA level in three groups, especially upregulation of ALP was larger in Smad 1, Smad 5 transfected cell than control group. Pretreatment with cycloheximide($10{\mu}g/ml$), a protein synthesis inhibitor resulted in blocking the ALP gene expression even in BMP(100ng/ml) treated cell. These results suggested that Smad increased the level of ALP mRNA via the synthesis of a certain transcriptional regulatory protein.

• Development and Reproduction
• /
• v.11 no.3
• /
• pp.235-243
• /
• 2007

Human umbilical cord blood(HUCB) contains a rich source of hematopoietic stem cells, mesenchymal stem cells and endothelial cell precursors. Mesenchymal stem cells(MSCs) in HUCB are multipotent stem cells, differ from hematopoietic stem cells and can be differentiated into neural cells. We studied on transdifferentiation-promoting conditions in neural cells and cholinergic neuron induction of HUCB-derived MSCs. Neural differentiation was induced by addingdimethyl sulphoxide(DMSO) and butylated hydroxyanisole(BHA) in Dulbeco's Modified Essential Medium(DMEM) and fetal bovine serum(FBS). Differentiation of MSCs to cholinergic neurons was induced by combined treatment with basic fibroblast growth factor(bFGF), retinoic acid(RA) and sonic hedgehog(Shh). MSCs treated with DMSO and BHA rapidly assumed the morphology of multipolar neurons. Both immunocytochemistry and RT-PCR analysis indicated that the expression of a number of neural markers including $\beta$-tubulin III, GFAP and MBP, was markedly elevated during this acute differentiation. The differentiation rate was about $32.3{\pm}2.9%$ for $\beta$-tubulin III-positive cells, $11.0{\pm}0.9%$ for GFAP, and $9.4{\pm}1.0%$ for Gal-C. HUCB-MSCs treated combinatorially with bFGF, RA and Shh were differentiated into cholinergic neurons. After cholinergic neuronal differentiation, the $\beta$-tubulin III-positive cell population of total cells was $31.3{\pm}3.2%$ and of differentiated neuronal population, $70.0{\pm}7.8%$ was ChAT-positive showing 3 folds higher in cholinergic population than neural induction. Conclusively, HUCB-derived MSCs can be differentiated into neural and cholinergic neurons and these findings suggest that HUCB are alternative cell source of treatment for neurodegenerative diseases such as Alzheimer's disease.

• Development and Reproduction
• /
• v.12 no.1
• /
• pp.31-39
• /
• 2008

Neural tissue has limited intrinsic capacity of repair after injury, and the identification of alternate sources of neural stem cells has broad clinical potential. We isolated mesechymal-like stem cells from human adipose tissues (AT-MSCs), and studied on transdifferentiation-promoting conditions in neural cells. Dopaminergic and cholinergic neuron induction of AT-MSCs was also studied. Neural differentiation was induced by adding bFGF, EGF, dimethyl sulphoxide (DMSO) and butylated hydroxyanisole(BHA) in N2 Medium and N2 supplement. The immunoreactive cells for $\beta$-tubulin III, a neuron-specific marker, GFAP, an astrocyte marker, or Gal-C, an oligodendrocyte marker, were found. AT-MSCs treated with bFGF, SHH and FGF8 were differentiatied into dopaminergic neurons that were immunopositive for TH antibody. Differentiation of MSCs to cholinergic neurons was induced by combined treatment with basic fibroblast growth factor (bFGF), retinoic acid (RA) and sonic hedgehog (Shh). AT-MSCs treated with DMSO and BHA rapidly assumed the morphology of multipolar neurons. Both immunocytochemistry and RT-PCR analysis indicated that the expression of a number of neural markers including neuro D1, $\beta$-tubulin III, GFAP and nestinwas markedly elevated during this acute differentiation. While the stem cell markers such as SCF, C-kit, and Stat-3 were not expressed after preinduction medium culture, we confirmed the differentiation of dopaminergic and cholinergic neurons by TH/$\beta$-tubulin III or ChAT/ $\beta$-tubulin III positive cells. Conclusively, AT-MSCs can be differentiated into dopaminergic and cholinergic neuronsand these findings suggest that AT-MSCs are alternative cell source of treatment for neurodegenerative diseases.

• Clinical and Experimental Pediatrics
• /
• v.52 no.8
• /
• pp.944-952
• /
• 2009

Purpose : To investigate the effects of angiotensin II inhibition on the epithelial to mesenchymal transition (EMT) in the developing kidney, we tested the expression of EMT markers and nestin in angiotensin converting enzyme (ACE) inhibitor-treated kidneys. Methods : Newborn rat pups were treated with enalapril (30 mg/kg/d) or a vehicle for 7 days. Immunohistochemistry for the expression of ${\alpha}$-smooth muscle actin (SMA), E-cadherin, vimentin, and nestin were performed. The number of positively-stained cells was determined under 100 magnification in 10 random fields. Results : In the enalapril-treated group, ${\alpha}SMA-positive$ cells were strongly expressed in the dilated tubular epithelial cells. The number of ${\alpha}SMA-positive$ cells in the enalapril-treated group increased in both the renal cortex and medulla, compared to the control group (P<0.05). The expression of E-cadherin-positive cells was dramatically reduced in the cortical and medullary tubular epithelial cells in the enalapril-treated group (P<0.05). The number of vimentin- and nestin-positive cells in the cortex was not different in comparisons between the two groups; however, their expression increased in the medullary tubulointerstitial cells in the enalapril-treated group (P<0.05). Conclusion : Our results show that ACE inhibition in the developing kidney increases the renal EMT by up-regulating ${\alpha}SMA$ and down-regulating E-cadherin. Enalapril treatment was associated with increased expression of vimentin and nestin in the renal medulla, suggesting that renal medullary changes during the EMT might be more prominent, and ACE inhibition might differentially modulate the expression of EMT markers in the developing rat kidney.