• Advances in pediatric surgery
• /
• v.15 no.1
• /
• pp.1-10
• /
• 2009

Recently, amniotic fluid has gained attention as one of the potential sources for cell therapy and tissue engineering because it has characteristics of multipotent stem cells. However, current knowledge about what types of cells are naturally found in amniotic fluid is still limited. In this study, we aimed to investigate whether human amniotic fluid contains cells that have characteristics of respiratory cells. Samples of human amniotic fluid (5 mL per sample) obtained from amniocenteses were cultured with small airway growth medium (SAGM). Cells were grown until the third passage and the presence of type II alveolar cells were characterized by inverted microscopy, immunofluorescence, and reverse transcription polymerase chain reaction (RT-PCR). On inverted microscopy, cultured cells showed typical polygonal and cobblestone-like epithelial morphology. The morphology of cells was not changed after selection and passing. Immunofluorescence analysis demonstrated that the isolated cells stained positive for surfactant protein C (SPC), specific marker for type II alveolar cells. Cells also stained positive for TTF-1 protein but negative for CD 31 and vimentin. RT-PCR analysis of cells showed expression of SPC mRNA. This study has demonstrated that respiratory cells can be isolated and identified from human amniotic fluid cultured in SAGM medium. Our results may provide the basis for further investigations of amniotic fluid.

• Molecules and Cells
• /
• v.38 no.7
• /
• pp.663-668
• /
• 2015

hBMSCs are multipotent cells that are useful for tissue regeneration to treat degenerative diseases and others for their differentiation ability into chondrocytes, osteoblasts, adipocytes, hepatocytes and neuronal cells. In this study, biodegradable elastic hydrogels consisting of hydrophilic poly(ethylene glycol) (PEG) and hydrophobic poly(${\varepsilon}$-caprolactone) (PCL) scaffolds were evaluated for tissue engineering because of its biocompatibility and the ability to control the release of bioactive peptides. The primary cultured cells from human bone marrow are confirmed as hBMSC by immunohistochemical analysis. Mesenchymal stem cell markers (collagen type I, fibronectin, CD54, $integrin1{\beta}$, and Hu protein) were shown to be positive, while hematopoietic stem cell markers (CD14 and CD45) were shown to be negative. Three different hydrogel scaffolds with different block compositions (PEG:PCL=6:14 and 14:6 by weight) were fabricated using the salt leaching method. The hBMSCs were expanded, seeded on the scaffolds, and cultured up to 8 days under static conditions in Iscove's Modified Dulbecco's Media (IMDM). The growth of MSCs cultured on the hydrogel with PEG/PCL= 6/14 was faster than that of the others. In addition, the morphology of MSCs seemed to be normal and no cytotoxicity was found. The coating of the vascular endothelial growth factor (VEGF) containing scaffold with Matrigel slowed down the release of VEGF in vitro and promoted the angiogenesis when transplanted into BALB/c nude mice. These results suggest that hBMSCs can be supported by a biode gradable hydrogel scaffold for effective cell growth, and enhance the angiogenesis by Matrigel coating.

• Molecules and Cells
• /
• v.40 no.6
• /
• pp.386-392
• /
• 2017

Periodontal ligament stem cells (PDLSCs) are multipotent stem cells derived from periodontium and have mesenchymal stem cell (MSC)-like characteristics. Recently, the perivascular region was recognized as the developmental origin of MSCs, which suggests the in vivo angiogenic potential of PDLSCs. In this study, we investigated whether PDLSCs could be a potential source of perivascular cells, which could contribute to in vivo angiogenesis. PDLSCs exhibited typical MSC-like characteristics such as the expression pattern of surface markers (CD29, CD44, CD73, and CD105) and differentiation potentials (osteogenic and adipogenic differentiation). Moreover, PDLSCs expressed perivascular cell markers such as NG2, ${\alpha}-smooth$ muscle actin, platelet-derived growth factor receptor ${\beta}$, and CD146. We conducted an in vivo Matrigel plug assay to confirm the in vivo angiogenic potential of PDLSCs. We could not observe significant vessel-like structures with PDLSCs alone or human umbilical vein endothelial cells (HUVECs) alone at day 7 after injection. However, when PDLSCs and HUVECs were co-injected, there were vessel-like structures containing red blood cells in the lumens, which suggested that anastomosis occurred between newly formed vessels and host circulatory system. To block the $SDF-1{\alpha}$ and CXCR4 axis between PDLSCs and HUVECs, AMD3100, a CXCR4 antagonist, was added into the Matrigel plug. After day 3 and day 7 after injection, there were no significant vessel-like structures. In conclusion, we demonstrated the perivascular characteristics of PDLSCs and their contribution to in vivo angiogenesis, which might imply potential application of PDLSCs into the neovascularization of tissue engineering and vascular diseases.

• Molecules and Cells
• /
• v.41 no.11
• /
• pp.953-963
• /
• 2018

The stepwise development of T cells from a multipotent precursor is guided by diverse mechanisms, including interactions among lineage-specific transcription factors (TFs) and epigenetic changes, such as DNA methylation and hydroxymethylation, which play crucial roles in mammalian development and lineage commitment. To elucidate the transcriptional networks and epigenetic mechanisms underlying T-cell lineage commitment, we investigated genome-wide changes in gene expression, DNA methylation and hydroxymethylation among populations representing five successive stages of T-cell development (DN3, DN4, DP, $CD4^+$, and $CD8^+$) by performing RNA-seq, MBD-seq and hMeDIP-seq, respectively. The most significant changes in the transcriptomes and epigenomes occurred during the DN4 to DP transition. During the DP stage, many genes involved in chromatin modification were up-regulated and exhibited dramatic changes in DNA hydroxymethylation. We also observed 436 alternative splicing events, and approximately 57% (252) of these events occurred during the DP stage. Many stage-specific, differentially methylated regions were observed near the stage-specific, differentially expressed genes. The dynamic changes in DNA methylation and hydroxymethylation were associated with the recruitment of stage-specific TFs. We elucidated interactive networks comprising TFs, chromatin modifiers, and DNA methylation and hope that this study provides a framework for the understanding of the molecular networks underlying T-cell lineage commitment.

• Journal of the Korean Society of Radiology
• /
• v.5 no.1
• /
• pp.11-18
• /
• 2011

The periosteum contains multipotent cells that can differentiate into osteoblasts and chondrocytes. Cultured periosteum-derived cells (PDCs) have an osteogenic capacity. The purpose of this study was to evaluate the interaction of PDCs with bone graft biomaterial. After cell isolation from the calvarial periosteum of Sprague-Dawley rats, cultured PDCs were placed in critical-sized calvarial defects with beta-tricalcium phosphate (${\beta}$-TCP). All rats were sacrificed 8 weeks after bone graft surgery, and the bone regenerative ability of bone grafting sides was evaluated by plain radiography, micro-computed tomography (CT), and histological examination. PDCs grafted with ${\beta}$-TCP displayed enhanced calcification in the defect site, density of regenerated bone and new bone formation within the defect and its boundaries. Furthermore, these PDCs more efficiently regenerated new bone as compared to grafted ${\beta}$-TCP only. The results suggest that cultured PDCs have the potential to promote osteogenesis in bone defects.

• Journal of Acupuncture Research
• /
• v.23 no.5
• /
• pp.69-77
• /
• 2006

Background : Mesenchymal stem cells (MSCs) are present in most of the tissue matrix, taking part in their regeneration when injury or damage occurs. The aim of this study was to investigate the presence of cells with pluripotential characteristics in human subchondral bone and the capacity of these cells to differentiate to osteoblast. Methods : Human subchondral bone were digested with collagenase. Isolated cells were cultured with a-MEM, 15% FBS, 10-8M dexamethasone and 50 ng/mL ascoric acid. Cells from 0 day(isolated cells), 7 day (first subculture) and 14 days (third subculture) were used to carry out phenotypic characterization experiments flowcytometry analysis with 11 monoclonal antibodies) and osteogenic differentiation experiments. Osteogenic differentiation of cells was assessment by quantification of bone extracellular matrix components by following analysis: alkaline phosphatase(ALP) stains to detect ALP activity, RT-PCR and western blot to detect osteocalcin (OCN), osteopontin (OPN) and type I collagen(Col I), and Alizarin red stains to detect calcium deposition. Results : Flowcytometry analyses showed that in our population more than 98% of cells were positive for MSC markers: SH-2(CD105, 99%), CD29 (95%), CD73 (95%). Cells were negative for hematopoietic markers (CD11b, CD34, and CD45). Furthermore, cells showed positive stain to multipotent markers such as CDl17 (c-kit) (15.1%), and CD166 (74.9%), and cell adhesion molecules such as CD54 (78.1%) and CD106 (63.5%). The osteogenic specific marker analyses showed that the culture of these cells for 7 and 14 days stimulates ALP, OCN, OPN and Col I synthesis by RT-PCR and Western blot analysis. Also, after 14 days in the culture of MSCs induces mineralization by Arizarin red stain. Conclusion : In this work, we demonstrated a new and efficient method for osteoblastic differentiation of human subchondral bone stem cells. As MSCs takes part in reparative processes of adult tissues, these cells could play an important role in osteogenesis.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
• /
• v.33 no.5
• /
• pp.405-418
• /
• 2007

Mesenchymal stem cells(MSCs) have been though to be multipotent cells that can replicate that have the potential to differentiate into lineages of mesenchymal tissue including the bone, cartilage, fat, tendon, muscle, and marrow stroma. Especially, scaffolds to support cell-based tissue engineering are critical determinants of clinical efforts to regenerate and repair the body. Selection of a matrix carrier imvolves consideration of the matrix's role as a scaffold for physical support and host tissue integration as well as its ability to support of synergize the osteoinductive program of the implanted mesenchymal stem cell. The aim of this study is to evaluate the effect of autobone and Bio-$Oss^{(R)}$ to adherent mesenchymal stem cells as scaffolds on sinus augmentation with fibrin glue mixture in a rabbit model. 16 New Zealand White rabbits were divided randomly into 4 groups based on their time of sacrifice(1, 2, 4 and 8 weeks). First, mesenchymal stem cells were isolated from iliac crest marrow of rabbits and expanded in vitro. Cell culture was performed in accordance with the technique described by Tsutsumi et al. In the present study, the animals were sacrificed at 1, 2, 4 and 8 weeks after transplantation, and the bone formation ability of each sides was evaluated clinically, radiologically, histologically and histomorphologically. According to the histological observations, autobone scaffolds group showed integrated graft bone with host bone from sinus wall. At 2 and 4 weeks, it showed active newly formed bone and neovascularization. At 8 weeks, lamellae bone was observed in sinus graft material area. Radiologically, autobone with stem cell showed more radiopaque than Bio-$Oss^{(R)}$ scaffolds group. there were significant differences in bone volume between 4 and 8 weeks(p<0.05).

• The Korean Journal of Pain
• /
• v.33 no.1
• /
• pp.23-29
• /
• 2020

Background: Neuropathic pain (NP) is considered a clinically incurable condition despite various treatment options due to its diverse causes and complicated disease mechanisms. Since the early 2000s, multipotent human mesenchymal stem cells (hMSCs) have been used in the treatment of NP in animal models. However, the effects of hMSC injections have not been studied in chronic post-ischemia pain (CPIP) mice models. Here, we investigated whether intrathecal (IT) and intrapaw (IP) injections of hMSCs can reduce mechanical allodynia in CPIP model mice. Methods: Seventeen CPIP C57/BL6 mice were selected and randomized into four groups: IT sham (n = 4), IT stem (n = 5), IP sham (n = 4), and IP stem (n = 4). Mice in the IT sham and IT stem groups received an injection of 5 μL saline and 2 × 10⁴ hMSCs, respectively, while mice in the IP sham and IP stem groups received an injection of 5 μL saline and 2 × 10⁵ hMSCs, respectively. Mechanical allodynia was assessed using von Frey filaments from pre-injection to 30 days post-injection. Glial fibrillary acidic protein (GFAP) expression in the spinal cord and dorsal root ganglia were also evaluated. Results: IT and IP injections of hMSCs improved mechanical allodynia. GFAP expression was decreased on day 25 post-injection compared with the sham group. Injections of hMSCs improved allodynia and GFAP expression was decreased compared with the sham group. Conclusions: These results suggested that hMSCs may be also another treatment modality in NP model by ischemia-reperfusion.

• Applied Microscopy
• /
• v.39 no.3
• /
• pp.253-260
• /
• 2009

GM-CSF is a multipotent growth factor, which also plays an important role during the process of wound healing. rrhGM-CSF was specifically produced from rice cell culture in our laboratory (Hanson Biotech Co., Ltd, Daejeon). The rrhGMCSF contains more oligosaccharide side chains than any other types of GM-CSF. This work was taken to evaluate the influence on wound healing of rrhGM-CSF in male golden hamsters. Full thickness skin defects of 9 mm in diameter were made in the back of hamsters, and 100 ${\mu}L$ ointment containing rrhGM-CSF 50 ${\mu}g/mL$ was applied. Control groups were given ointment without rrhGM-CSF. The wound sizes were relatively reduced and skin was well regenerated in the experimental group compared with the control group. Structurally, reepithelialization and architecture of the skin following injury were well accomplished in the experimental group. And also, positive reaction of PCNA of the skin following injury was more prominent in rrhGM-CSF containing ointment treatment group. Since this type of GM-CSF has highly glycosylated side chains, the effectiveness might be retain longer and stable, regarding acceleration of wound healing in the animal model. The present study has important implications for further development of the therapeutic manipulation of wound healing using rrhGM-CSF.

• Archives of Plastic Surgery
• /
• v.37 no.4
• /
• pp.323-328
• /
• 2010

Purpose: Adipose-derived stromal cells (ADSCs) are multipotent cells that have been found to promote wound healing through the process of angiogenesis and reepithelialization. Generally, it is well known that the antigenicity of ADSCs doesn't affect stem cell therapy. In this study, we investigated the effect of allogeneic ADSCs in the wound healing process by applying allogeneic ADSCs on the wound healing splint model of mice. Methods: Adipose tissue was harvested from the epididymal fat pads of BALB/c and C57BL/6 mice. Twenty four mice BALB/c were divided into three groups; control, isogeneic, and allogeneic groups. Two full thickness defects with 6 mm diameters were created on the back of BALB/c mice. $1{\times}10^6$ ADSCs from BALB/c mice were applied on the isogeneic group. In the allogeneic group, ADSCs from the C57BL/6 mice were applied. No cells were applied to the control group. The sizes of the wounds were evaluated in 3, 5, 7, 10, and 14 days after the wounds were applied, and tissues were harvested in 7 and 14 days for histological analysis. Results: Wound healing rates had showed significant increase in 10, and 14 days when the isogeneic group was compared to the control group, but the allogeneic group showed significantly decrease compared to the isogeneic group (p<0.05). Histological scores in the isogeneic group were significantly high, but significantly lower in the allogeneic group when compared to the isogeneic group in 2 weeks (p<0.05). In the isogeneic group, thick inflammatory cell infiltration with abundant capillaries were observed in 1 week, and thick epithelium with many large capillaries were observed in 2 weeks. Conclusion: When isogeneic ADSCs were applied to wounds, they presented a faster wound healing rate compared to controls and the allogeneic group. Unlike general stem cell therapy, these findings suggest that cell therapy targeted at enhancing wound healing may benefit from the use of ADSCs with identical antigenicity, as opposed to allogeneic or xenogenic ADSCs.