• Archives of Plastic Surgery
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• v.35 no.6
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• pp.637-644
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• 2008

Purpose: The survival of bone marrow derived stem cell was reported several times. But the survival of adipose tissue derived stem cells(hASCs) was not mentioned on. We studied the adipose tissue derived stem cell's survival and effect on articular cartilage in rabbits. Methods: Osteoarthritis was induced in twenty New Zealand white rabbits by intraarticular injection of monosodium iodoacetate(MIA). After four weeks, hASCs were also injected into the knee joints space without any vehicle, but the control group received phosphate buffered saline only. The histologic grade of articular cartilage was measured in 4 and 8 weeks after the transplantation of hASC and the viability of injected stem cells measured by Fluorescent in situ Hybridization (FISH) examination. Results: After 4 and 8 weeks from hASCs transplantation, histologic grade was not significantly difference between two groups(p>0.05), and the Y chromosome of the transplanted hASCs was not detected in articular cartilage. Conclusion: We found that direct injection of hASC in joint space didn't work on damaged articular cartilage repair.

• BMB Reports
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• v.47 no.8
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• pp.469-474
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• 2014

Cell therapies utilizing mesenchymal stem cells (MSCs) have a great potential in many research and clinical settings. The mechanisms underlying the therapeutic effects of MSCs have been studied previously and the paracrine effects elicited by their production of various growth factors and cytokines were recognized as being crucial. However, the molecular controls that govern these paracrine effects remain poorly understood. To elucidate the molecular regulators of this process, we performed a global knockdown of microRNAs (miRNAs) in human adipose-derived mesenchymal stem cells (hADSCs) by inhibiting DGCR8, a key protein in miRNA biogenesis. Global disruption of miRNA biogenesis in hADSCs caused dramatic changes in the expression of subsets of growth factors and cytokines. By performing an extensive bioinformatic analysis, we were able to associate numerous putative miRNAs with these genes. Taken together, our results strongly suggest that miRNAs are essential for the production of growth factors and cytokines in hADSCs.

• Korean Chemical Engineering Research
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• v.51 no.6
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• pp.727-732
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• 2013

In this report, we present an inverse opal scaffold that can enhance the chondrogenic differentiation of human adipose-derived stem cells (hADSCs) without drug, gene, or cytokine supplement. Inverse opal scaffolds based on poly(D,L-lactide-co-glycolide) were formed with uniform $200{\mu}m$ pores. Due to uniform pore sizes and well-controlled interconnectivity of inverse opal scaffold, hADSCs were allowed to distribute homogeneously throughout the scaffolds. As a result, high cell density culture with scaffold was possible. Since the hADSCs cultured in inverse opal scaffolds were subjected to limited supplies of oxygen and nutrients, these cells were naturally preconditioned to a hypoxic environment that stimulated the up-regulation of hypoxia-inducible factor-$1{\alpha}$ (HIF-$1{\alpha}$). As a result, apoptotic activity of hADSCs until 3 weeks after initial cell seeding was significantly reduced and chondrogenic differentiation related molecular signal cascades were up regulated (transforming growth factor-beta, phosphorylated AKT, and phosphorylated p38 expression). In contrast, hADSCs cultured with small and non-uniform porous scaffolds showed significantly increased apoptotic activity with decreased chondrogenic differentiation. Taken together, inverse opal scaffold could potentially be used as an effective tool for improving chondrogenesis using stem cells.

• Molecules and Cells
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• v.38 no.4
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• pp.336-342
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• 2015

Propyl gallate (PG) used as an additive in various foods has antioxidant and anti-inflammatory effects. Although the functional roles of PG in various cell types are well characterized, it is unknown whether PG has effect on stem cell differentiation. In this study, we demonstrated that PG could inhibit adipogenic differentiation in human adipose tissue-derived mesenchymal stem cells (hAMSCs) by decreasing the accumulation of intracellular lipid droplets. In addition, PG significantly reduced the expression of adipocyte-specific markers including peroxisome proliferator-activated receptor-${\gamma}$ (PPAR-${\gamma}$), CCAAT enhancer binding protein-${\alpha}$ (C/EBP-${\alpha}$), lipoprotein lipase (LPL), and adipocyte fatty acid-binding protein 2 (aP2). PG inhibited adipogenesis in hAMSCs through extracellular regulated kinase (ERK) pathway. Decreased adipogenesis following PG treatment was recovered in response to ERK blocking. Taken together, these results suggest a novel effect of PG on adipocyte differentiation in hAMSCs, supporting a negative role of ERK1/2 pathway in adipogenic differentiation.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.32 no.2
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• pp.97-106
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• 2010

Aim of the study: An alternative source of adult stem cells that could be obtained in large quantities, under local anesthesia, with minimal discomfort would be advantageous. Adipose tissue could be processed to obtain a fibroblast-like population of cells or adipose tissue-derived stromal cells (ATSCs). This study was performed to confirm the availability of ATSCs in bone tissue engineering. Materials amp; Methods: In this study, adipose tissue-derived mesenchymal stem cell was extracted from the liposuctioned abdominal fat of 24-old human and cultivated, and the stem cell surface markers of CD 105 and SCF-R were confirmed by immunofluorescent staining. The proliferation of bone marrow mesenchymal stem cell and ATSCs were compared, and evaluated the osteogenic differentiation of ATSCs in a specific osteogenic induction medium. Osteogenic differentiation was assessed by von Kossa and alkaline phosphatase staining. Expression of osteocyte specific BMP-2, ALP, Cbfa-1, Osteopontin and osteocalcin were confirmed by RT-PCR. With differentiation of ATSCs, calcium concentration was assayed, and osteocalcin was evaluated by ELISA (Enzyme-linked immunosorbant assay). The bone formation by 5-week implantation of HA/TCP block loaded with bone marrow mesenchymal stem cells and ATSCs in the subcutaneous pocket of nude mouse was evaluated by histologic analysis. Results: ATSCs incubated in the osteogenic medium were stained positively for von Kossa and alkaline phosphatase staining. Expression of osteocyte specific genes was also detected. ATSCs could be easily identified through fluorescence microscopy, and bone formation in vivo was confirmed by using ATSC-loaded HA/TCP scaffold. Conclusions: The present results show that ATSCs have an ability to differentiate into osteoblasts and formed bone in vitro and in vivo. So ATSCs may be an ideal source for further experiments on stem cell biology and bone tissue engineering.

• Archives of Plastic Surgery
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• v.38 no.4
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• pp.345-350
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• 2011

Purpose: Although platelet-rich plasma (PRP) potentiate the wound healing activity of adipose-derived stem cells (ADSCs), its effect cannot be sustained for a prolonged period of time due to short duration of action. This led us to design and produce platelet-rich fibrin (PRF), in an effort to develop a tool which lasts longer, and apply it on wound healing. Methods: Two symmetrical skin defects were made on the back of seven nude mice. ADSCs were applied to each wound, combined with either PRP or PRF. The wound area was measured over 14 days. By day 16, the wound was harvested and histologic analysis was performed including counting of the blood vessel. Results: The healing rate was more accelerated in PRP group in the first 5 days (p<0.05). However, PRF group surpassed PRP group after 6 days (p<0.05). The average number of blood vessels observed in the PRF group was $6.53{\pm}0.51$, compared with $5.68{\pm}0.71$ for the PRP group. Conclusion: PRF exerts a slow yet pervasive influence over the two-week course of the wound healing process. Thus, PRF is probably more beneficial for promoting the activity of ADSCs for a sustained period of time.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.32 no.4
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• pp.299-305
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• 2010

Purpose: Adipose tissue is located beneath the skin, around internal organs, and in the bone marrow in humans. Its main role is to store energy in the form of fat, although it also cushions and insulates the body. Adipose tissue also has the ability to dynamically expand and shrink throughout the life of an adult. Recently, it has been shown that adipose tissue contains a population of adult multipotent mesenchymal stem cells and endothelial progenitor cells that, in cell culture conditions, have extensive proliferative capacity and are able to differentiate into several lineages, including, osteogenic, chondrogenic, endothelial cells, and myogenic lineages. Materials and Methods: This study focused on endothelial cell culture from the adipose tissue. Adipose tissues were harvested from buccal fat pad during bilateral sagittal split ramus osteotomy for surgical correction of mandibular prognathism. The tissues were treated with 0.075% type I collagenase. The samples were neutralized with DMEM/and centrifuged for 10 min at 2,400 rpm. The pellet was treated with 3 volume of RBC lysis buffer and filtered through a 100 ${\mu}m$ nylon cell strainer. The filtered cells were centrifuged for 10 min at 2,400 rpm. The cells were further cultured in the endothelial cell culture medium (EGM-2, Cambrex, Walkersville, Md., USA) supplemented with 10% fetal bovine serum, human EGF, human VEGF, human insulin-like growth factor-1, human FGF-$\beta$, heparin, ascorbic acid and hydrocortisone at a density of $1{\times}10^5$ cells/well in a 24-well plate. Low positivity of endothelial cell markers, such as CD31 and CD146, was observed during early passage of cells. Results: Increase of CD146 positivity was observed in passage 5 to 7 adipose tissue-derived cells. However, CD44, representative mesenchymal stem cell marker, was also strongly expressed. CD146 sorted adipose tissue-derived cells was cultured using immuno-magnetic beads. Magnetic labeling with 100 ${\mu}l$ microbeads per 108 cells was performed for 30 minutes at $4^{\circ}C$ a using CD146 direct cell isolation kit. Magnetic separation was carried out and a separator under a biological hood. Aliquous of CD146+ sorted cells were evaluated for purity by flow cytometry. Sorted cells were 96.04% positivity for CD146. And then tube formation was examined. These CD146 sorted adipose tissue-derived cells formed tube-like structures on Matrigel. Conclusion: These results suggest that adipose tissue-derived cells are endothelial cells. With the fabrication of the vascularized scaffold construct, novel approaches could be developed to enhance the engineered scaffold by the addition of adipose tissue-derived endothelial cells and periosteal-derived osteoblastic cells to promote bone growth.

• BMB Reports
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• v.50 no.11
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• pp.572-577
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• 2017

In most clinical applications, human mesenchymal stem cells (hMSCs) are expanded in large scale before their administration. Prolonged culture in vitro results in cellular senescence-associated phenotypes, including accumulation of reactive oxygen species (ROS) and decreased cell viabilities. Profiling of stem cell-related genes during in vitro expansion revealed that numerous canonical pathways were significantly changed. To determine the effect of selenocysteine (Sec), a rare amino acid found in several antioxidant enzymes, on the replicative senescence in hMSCs, we treated senescent hMSCs with Sec. Supplementation of Sec in the culture medium in late-passage hMSCs reduced ROS levels and improved the survival of hMSCs. In addition, a subset of key antioxidant genes and Sec-containing selenoproteins showed increased mRNA levels after Sec treatment. Furthermore, ROS metabolism and inflammation pathways were predicted to be downregulated. Taken together, our results suggest that Sec has antioxidant effects on the replicative senescence of hMSCs.

• Biomolecules & Therapeutics
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• v.22 no.4
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• pp.328-333
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• 2014

Vitiligo is a pigmentary disorder induced by a loss of melanocytes. In addition to replacement of pure melanocytes, cocultures of melanocytes with keratinocytes have been used to improve the repigmentation outcome in vitiligo treatment. We previously identified by in vitro studies, that adipose-derived stem cells (ADSCs) could be a potential substitute for keratinocytes in cocultures with melanocytes. In this study, the efficacy of pigmentation including durability of grafted melanocytes and short-term safety was examined in the nude mouse and Sprague-Dawley rat after grafting of primary cultured human melanocytes, with or without different ratios of primary cultured human ADSCs. Simultaneous grafting of melanocytes and ADSCs, which were separately cultured and mixed on grafting at the ratios of 1:1, 1:2, or 1:3, showed better efficacy than that of pure melanocytes. Grafting of melanocytes cocultured with ADSCs resulted in a similar outcome as the grafting of cell mixtures. Skin pigmentation by melanocytes : ADSCs at the ratios of 1:1 and 1:2 was better than at 1:3. No significant difference was observed between the 1-week and 2-week durations in coculturing. Time-course microscopic examination showed that the grafted melanocytes remained a little longer than 6-week post-grafting. No inflammatory cell infiltration was observed in the grafted skin and no melanocytes were detectable in other organs. Collectively, grafting of melanocytes and ADSCs was equally safe and more effective than grafting of melanocytes alone. Despite the absence of significant differences in efficacy between the group of 1:1 and that of 1:2 ratio, 1:2 ratio for 1-week coculturing may be better for clinical use from the cost-benefit viewpoint.

• Development and Reproduction
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• v.17 no.4
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• pp.409-420
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• 2013

Human serum (HS) has been reported to induce aggregation of human eyelid adipose-derived stem cells (HEACs) during high-density culture in vitro. The present study focused on the role of cell adhesion molecules and gelatinases during HS-induced aggregation of HEACs. HS-induced aggregation occurred between 9-15 days of culture. Cells aggregated by HS medium (HS-agg) showed stronger expression of ${\alpha}2$, ${\alpha}2B$, ${\alpha}X$, and CEACAM1 genes compared to non-aggregated cells in HS medium (HS-ex) or in control FBS-cultured cells. HS-agg were distinctly labeled with antibodies against ${\alpha}2$, ${\alpha}2B$, and ${\alpha}X$ proteins. Western blot results demonstrated that the two integrin proteins were greatly expressed in HS-agg compared to HS-ex and control FBS-cultured cells. Treatment of HEACs with anti-integrin ${\alpha}2$ antibody during culture in HS medium delayed aggregation formation. HS-agg exhibited strong expression of MMP1 and MMP9 compared to HS-ex or FBS-cultured cells. Conditioned media from HS-culture showed remarkable increase of MMP9 gelatinolytic activity in comparison to those from FBS-culture. However, there was no change of TIMP mRNA expression in relation to the HS-induced aggregation. Based on these results, it is suggested that integrin ${\alpha}2$, ${\alpha}2B$, and ${\alpha}X$, and MMP9 might play an important role in the HS-induced aggregation of HEACs.