• Molecules and Cells
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• 제24권1호
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• pp.139-147
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• 2007

Although transforming growth factors (TGFs) are implicated in the process of endochondral ossification, which is initiated by the differentiation of mesenchymal cells into chondrocytes, it is not clear how $TGF-{\beta}3$ regulates the chondrogenic differentiation of limb bud mesenchymal cells. Here, differential display polymerase chain reaction (DD-PCR) screening and RT-PCR analysis revealed that transcripts of A Disintegrin And Metalloprotease 10 (ADAM 10) decreased during the chondro-inhibitory action of $TGF-{\beta}3$ on cultured chick leg bud mesenchymal cells. Electroporation of ADAM 10 morpholino antisense oligonucleotides inhibited the ectodomain shedding of delta-1, and cell proliferation and subsequent precartilage condensation, in a manner similar to that caused by $TGF-{\beta}3$. The suppression of mesenchymal cell proliferation induced by $TGF-{\beta}3$ and ADAM 10 morpholino antisense oligonucleotides was reversed by activation of ADAM 10 with phorbol 12-myristate 13-acetate (PMA) or knockdown of Notch-1 with siRNA. Collectively, these data indicate that, in cultured chick leg bud mesenchyme cells, $TGF-{\beta}3$ downregulates ADAM 10 and inhibits cell proliferation and subsequent precartilage condensation by inhibiting the ectodomain shedding of delta-1, and that this results in the activation of Notch signaling.

• Asian-Australasian Journal of Animal Sciences
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• 제26권4호
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• pp.588-595
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• 2013

Mesenchymal stem cells (MSCs) are often known to have a therapeutic potential in the cell-mediated repair for fatal or incurable diseases. In this study, canine umbilical cord MSCs (cUC-MSCs) were isolated from umbilical cord matrix (n = 3) and subjected to proliferative culture for 5 consecutive passages. The cells at each passage were characterized for multipotent MSC properties such as proliferation kinetics, expression patterns of MSC surface markers and self-renewal associated markers, and chondrogenic differentiation. In results, the proliferation of the cells as determined by the cumulative population doubling level was observed at its peak on passage 3 and stopped after passage 5, whereas cell doubling time dramatically increased after passage 4. Expression of MSC surface markers (CD44, CD54, CD61, CD80, CD90 and Flk-1), molecule (HMGA2) and pluripotent markers (sox2, nanog) associated with self-renewal was negatively correlated with the number of passages. However, MSC surface marker (CD105) and pluripotent marker (Oct3/4) decreased with increasing the number of subpassage. cUC-MSCs at passage 1 to 5 underwent chondrogenesis under specific culture conditions, but percentage of chondrogenic differentiation decreased with increasing the number of subpassage. Collectively, the present study suggested that sequential subpassage could affect multipotent properties of cUC-MSCs and needs to be addressed before clinical applications.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제32권6호
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• pp.495-505
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• 2006

In this study we evaluate the effects of bFGF-BB and PDGF on in vitro proliferation, differentiation and mineralization of mesenchymal stem cells (MSCs) from rat. MSCs were prepared from the bone marrow of 6 or 7-week-old male rats with a technique previously described by Maniatopoulos et al. in 1988. Lineage differentiation to osteogenesis, chondrogenesis and adipogenesis were performed. At first, we characterized the cultured cell on passage 1, 3, 5, 7 with immunocytochemical staining using CD29, 44, 34, 45, ${\alpha}$-SMA and type I collagen. And to study the effects of bFGF and PDGF-BB on proliferation, differentiation and mineralization, we seeded the expanded cell at a density of 6 $6{\times}10^3\;cells/cm^2$ to 100-mm dish for evaluation of cell proliferation and MTT assay was carried out on day 2, 4, 7, 9. We also resuspended the cells with same density $(6{\times}10^3\;cells/cm^2)$ to 24 well plates for subculture. On the following day, the attached cells were exposed to 2.5ng/ml bFGF and/or 25ng/ml PDGF-BB daily during 5 days. The osteocalcin (OC) level was assessed and mineral contents were evaluated with alizarin red S staining on subculture day 2, 7, 14, 21. We identified the mesenchymal stem cell from the bone marrow derived cells of rat through their successful multi-differentiation and stable display of its phenotype. And bFGF and PDGF-BB showed the effect that inhibited osteoblastic differentiation and mineralization mildly in above concentration at in vitro culture. This study was supported by grant 04-2004-0120 from the Seoul National University Hospital Research Fund.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제33권3호
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• pp.204-210
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• 2007

This study was intended to compare the cranial base morphology between the mandibular prognathism and maxillary retrognathism in skeletal class III patients. The subject of the present study was composed of 88 patients divided into two groups; Group 1 (Skeletal Class III with mandibular prognathism. SNA within normal range, SNB over normal range, n=54) and Group 2(Skeletal Class III with maxillary retrognathism. SNA below normal range, SNB within normal range, n=34). Lateral cephalogram were taken immediate before surgery and 18 landmarks were used to analyze the characteristics of cranial base and maxillomandibular skeleton. The result revealed that cranial base angle is significantly smaller in Group 1 than Group 2, which implies the influence of the cranial base angulation on the mandibular position. However the posterior cranial base length did not influence the mandibular horizontal position and anterior cranial base length did not influence the maxillary horizontal position. As the anterior cranial base length was closely related with ramal height, it is recommendable to investigate the regulatory mechanism of chondrogenesis of cranial base and condyle cartilage in the future research.

• 대한음성언어의학회:학술대회논문집
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• 대한음성언어의학회 2003년도 제19회 학술대회
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• pp.183-183
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• 2003

Background and Objectives : Cartilage reconstruction is one of medical issue in otolaryngology. Tissue engineering is presently being utilized in part of cartilage repair. Sources of cells for tissue engineering are chondrocyte from mature cartilage and bone marrow mesenchymal stem cells that are able to differentiate into chondrocyte. Recent studies have shown that adipose tissue have mesenchymal stem cells which can differentiate into adipogenic, chondrogenic myogenic osteogenic cells and neural cell in vitro. In this study, we have examined chondrogenic potential of the canine adipose tissue-derived mesenchymal stem cell(ATSC). Materials and Methods : We harvested canine adipose tissue from inguinal area. ATSCs were enzymatically released from canine adipose tissue. Under appropriate culture conditions, ATSCs were induced to differentiate into the chondrocyte lineages using micromass culture technique. We used immunostain to type II collagen and toluidine blue stain to confirm chondrogenic differentiation of ATSCs. Results : We could isolate ATSCs from canine adipose tissue. ATSCs expressed CD29 and CD44 which are specific surface markers of mesenchymal stem cell. ATSCs differentiated into micromass that has positive response to immunostain of type II collagen and toluidine blue stain. Conclusion : In vitro, ATSCs differentiated into cells that have characteristic cartilage matrix molecules in the presence of lineage-specific induction factors. Adipose tissue may represent an alternative source to bone marrow-derived MSCs.

• 한방재활의학과학회지
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• 제30권4호
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• pp.1-16
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• 2020

Objectives The main purpose of this study was to evaluate the bone healing effect of Pahyeolsandong-tang (PHT)(Poxiesanteng-tang) extract in tibia fracture-induced mice. Methods PHT was extracted using a solution of 35% ethanol in 60℃ for 8 hours. Mice were randomly divided into 4 groups (normal, control, PHT 50 and PHT 100). Mice of experimental groups were medicated with PHT 50 or 100 mg/kg for 7 to 21 days. To clarify the effect of bone fracture healing, relative messenger RNA (mRNA) expressions of osteocalcin (OCN), runt-related transcription factor 2 (Runx2), osterix (OSX), Sox9, collagen type II alpha 1 chain (Col2a1), receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegerin (OPG) were examined. Results In in vitro experiment, relative mRNA expression of OCN, Runx2, Col2a1 was significantly increased in PHT treated group to compare with control differentiation group. In in vivo experiment, relative mRNA expression of OCN, Runx2, OSX, Sox9, Col2a1, RANKL, OPG was significantly increased in PHT treated group. Conclusions This study showed that PHT accelerates bone fracture healing through the activation of osteoclasts and osteoblasts. It was showed that PHT significantly promotes osteoblasts differentiation by osteoblast differentiation markers such as OCN, Runx2, Col1a2. Also it was investigated that PHT had stimulatory effect on osteoblasts function through enhancing OCN, Runx2, OSX, Sox9, Col2a1 and, osteoclasts function through enhancing RANKL and OPG markers. PHT effectively promotes bone fracture healing process through activation of osteoblasts and osteoclasts.

• 대한수의학회지
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• 제58권4호
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• pp.211-217
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• 2018

Mesenchymal stem cells (MSCs) are useful candidates for tissue engineering and cell therapy. Physiological cell environment not only connects cells to each other, but also connects cells to the extracellular matrix that provide mechanical support, thus exposing the entire cell surface and activating signaling pathways. Hydrogel is a polymeric material that swells in water and maintains a distinct 3-dimensional (3D) network structure by cross linking. In this study, we investigated the optimized cellular function for canine adipose tissue-derived MSCs (cAD-MSCs) using hydrogel. We observed that the expression levels of Ki67 and proliferating cell nuclear antigen, which are involved in cell proliferation and stemness, were increased in transwell-hydrogel (3D-TN) compared to the transwell-normal (TN). Also, transforming growth factor-${\beta}1$ and SOX9, which are typical bone morphogenesis-inducing factors, were increased in 3D-TN compared to the TN. Collagen type II alpha 1, which is a chondrocyte-specific marker, was increased in 3D-TN compared to the TN. Osteocalcin, which is a osteocyte-specific marker, was increased in 3D-TN compared to the TN. Collectively, preconditioning cAD-MSCs via 3D culture systems can enhance inherent secretory properties that may improve the potency and efficacy of MSCs-based therapies for bone regeneration process.

• 생체재료학회지
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• 제22권4호
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• pp.337-345
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• 2018

Background: Human mesenchymal stem cells (hMSCs) are, due to their pluripotency, useful sources of cells for stem cell therapy and tissue regeneration. The phenotypes of hMSCs are strongly influenced by their microenvironment, in particular the extracellular matrix (ECM), the composition and structure of which are important in regulating stem cell fate. In reciprocal manner, the properties of ECM are remodeled by the hMSCs, but the mechanism involved in ECM remodeling by hMSCs under topographical stimulus is unclear. In this study, we therefore examined the effect of nanotopography on the expression of ECM proteins by hMSCs by analyzing the quantity and structure of the ECM on a nanogrooved surface. Methods: To develop the nanoengineered, hMSC-derived ECM, we fabricated the nanogrooves on a coverglass using a UV-curable polyurethane acrylate (PUA). Then, hMSCs were cultivated on the nanogrooves, and the cells at the full confluency were decellularized. To analyze the effect of nanotopography on the hMSCs, the hMSCs were re-seeded on the nanoengineered, hMSC-derived ECM. Results: hMSCs cultured within the nano-engineered hMSC-derived ECM sheet showed a different pattern of expression of ECM proteins from those cultured on ECM-free, nanogrooved surface. Moreover, hMSCs on the nano-engineered ECM sheet had a shorter vinculin length and were less well-aligned than those on the other surface. In addition, the expression pattern of ECM-related genes by hMSCs on the nanoengineered ECM sheet was altered. Interestingly, the expression of genes for osteogenesis-related ECM proteins was downregulated, while that of genes for chondrogenesis-related ECM proteins was upregulated, on the nanoengineered ECM sheet. Conclusions: The nanoengineered ECM influenced the phenotypic features of hMSCs, and that hMSCs can remodel their ECM microenvironment in the presence of a nanostructured ECM to guide differentiation into a specific lineage.

• 대한본초학회지
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• 제37권1호
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• pp.51-59
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• 2022

Objectives : The process through which mesenchymal cells condense and differentiate into chondrocytes to form new bone is known as endochondral bone formation. Chondrogenic differentiation and hypertrophy are essential steps in bone formation and are influenced by various factors. The stem bark and root bark of Eleutherococcus sessiliflorus (ES) have been widely used to treat growth retardation and arthritis in traditional Korean Medicine. In this study, we aimed to investigate the possible role of the stem bark of ES in the stimulation of chondrogenic differentiation in clonal murine chondrogenic ATDC5 cells. Methods : In ATDC5 cells treated with ES extract, cell viability and extracellular matrix production were determined using CCK-8 assay and Alcian blue staining, respectively, and alkaline phosphatase activity was measured. We also examined mRNA and protein expression levels of genes related to chondrogenic expression in ATDC5 cells using reverse transcription-polymerase chain reaction and western blot analyses. Results : ES extract increased the accumulation of Alcian blue-stained cartilage nodules and alkaline phosphatase activity in ATDC5 cells. It increased the mRNA expressions of chondrogenic markers including bone sialoprotein (BSP), cartilage collagens, Runt-related transcription factor-2 (RUNX-2), osteocalcin (OCN), β-catenin, and bone morphogenetic protein-2 (BMP-2), as well as the protein expressions of β-catenin, RUNX-2, BMP-2, and alkaline phosphatase (ALP). Conclusion : Taken together, these results suggest that ES extract exhibits a chondromodulating activity and therefore may be a possible agent for the treatment of bone growth disorders.

• BMB Reports
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• 제55권5호
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• pp.226-231
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• 2022

It is well-known that some species of lizard have an exceptional ability known as caudal autotomy (voluntary self-amputation of the tail) as an anti-predation mechanism. After amputation occurs, they can regenerate their new tails in a few days. The new tail section is generally shorter than the original one and is composed of cartilage rather than vertebrae bone. In addition, the skin of the regenerated tail distinctly differs from its original appearance. We performed a proteomics analysis for extracts derived from regenerating lizard tail tissues after amputation and found that endoplasmin (ENPL) was the main factor among proteins up-regulated in expression during regeneration. Thus, we performed further experiments to determine whether ENPL could induce chondrogenesis of tonsil-derived mesenchymal stem cells (T-MSCs). In this study, we found that chondrogenic differentiation was associated with an increase of ENPL expression by ER stress. We also found that ENPL was involved in chondrogenic differentiation of T-MSCs by suppressing extracellular signal-regulated kinase (ERK) phosphorylation.