• Journal of Microbiology and Biotechnology
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• v.27 no.12
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• pp.2156-2164
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• 2017

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is considered as an antitumor agent owing to its ability to induce apoptosis of cancer cells without imparting toxicity toward most normal cells. TRAIL is produced in poor yield because of its insoluble expression in the cytoplasm of E. coli. In this study, we achieved soluble expression of TRAIL by fusing maltose-binding protein (MBP), b'a' domain of protein disulfide isomerase (PDIb'a'), or protein disulfide isomerase at the N-terminus of TRAIL. The TRAIL was purified using subsequent immobilized metal affinity chromatography and amylose-binding chromatography, with the tag removal using tobacco etch virus protease. Approximately 4.5 mg of pure TRAIL was produced from 125 ml flask culture with a purification yield of 71.6%. The endotoxin level of the final product was $0.4EU/{\mu}g$, as measured by the Limulus amebocyte lysate endotoxin assay. The purified TRAIL was validated and shown to cause apoptosis of HeLa cells with an $EC_{50}$ and Hill coefficient of $0.6{{\pm}}0.03nM$ and $2.41{\pm}0.15$, respectively. The high level of apoptosis in HeLa cells following administration of purified TRAIL indicates the significance and novelty of this method for producing high-grade and high-yield TRAIL.

• Clinical and Experimental Pediatrics
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• v.46 no.5
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• pp.418-422
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• 2003

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2009.02a
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• pp.34-34
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• 2009

• Journal of Korean Neurosurgical Society
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• v.42 no.2
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• pp.118-124
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• 2007

Objective : Adipose tissue is derived from the embryonic mesoderm and contains a heterogenous stromal cell population. Authors have tried to verify the characteristics of stem cell of adipose derived stromal cells (ADSCs) and to investigate immunohistochemical findings after transplantation of ADSC into rat brain to evaluate survival, migration and differentiation of transplanted stromal cells. Methods : First ADSCs were isolated from human adipose tissue and induced adipose, osseous and neuronal differentiation under appropriate culture condition in vitro and examined phenotypes profile of human ADSCs in undifferentiated states using flow cytometry and immunohistochemical study. Human ADSCs were transplanted into the healthy rat brain to investigate survival, migration and differentiation after 4 weeks. Results : From human adipose tissue, adipose stem cells were harvested and subcultured for several times. The cultured ADSCs were differentiated into adipocytes, osteoctye and neuron-like cell under conditioned media. Flow cytometric analysis of undifferentiated ADSCs revealed that ADSCs were positive for CD29, CD44 and negative for CD34, CD45, CD117 and HLA-DR. Transplanted human ADSCs were found mainly in cortex adjacent to injection site and migrated from injection site at a distance of at least 1 mm along the cortex and corpus callosum. A few transplanted cells have differentiated into neuron and astrocyte. Conclusion : ADSCs were differentiated into multilineage cell lines through transdifferentiation. ADSCs were survived and migrated in xenograft without immunosuppression. Based on this data, ADSCs may be potential source of stem cells for many human disease including neurologic disorder.

• Journal of Korean Neurosurgical Society
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• v.62 no.2
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• pp.153-165
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• 2019

Objective : Spinal cord injury (SCI) is a very serious health problem, usually caused by a trauma and accompanied by elevated levels of inflammation indicators. Stem cell-based therapy is promising some valuable strategies for its functional recovery. Nestin-positive progenitor and/or stem cells (SC) isolated from pancreatic islets (PI) show mesenchymal stem cell (MSC) characteristics. For this reason, we aimed to analyze the effects of rat pancreatic islet derived stem cell (rPI-SC) delivery on functional recovery, as well as the levels of inflammation factors following SCI. Methods : rPI-SCs were isolated, cultured and their MSC characteristics were determined through flow cytometry and immunofluorescence analysis. The experimental rat population was divided into three groups : 1) laminectomy & trauma, 2) laminectomy & trauma & phosphate-buffered saline (PBS), and 3) laminectomy+trauma+SCs. Green fluorescent protein (GFP) labelled rPI-SCs were transplanted into the injured rat spinal cord. Their motilities were evaluated with Basso, Beattie and Bresnahan (BBB) Score. After 4-weeks, spinal cord sections were analyzed for GFP labeled SCs and stained for vimentin, $S100{\beta}$, brain derived neurotrophic factor (BDNF), 2',3'-cyclic-nucleotide 3'-phosphodiesterase (CNPase), vascular endothelial growth factor (VEGF) and proinflammatory (interleukin [IL]-6, transforming growth factor $[TGF]-{\beta}$, macrophage inflammatory protein [MIP]-2, myeloperoxidase [MPO]) and anti-inflammatory (IL-1 receptor antagonis) factors. Results : rPI-SCs were revealed to display MSC characteristics and express neural and glial cell markers including BDNF, glial fibrillary acidic protein (GFAP), fibronectin, microtubule associated protein-2a,b (MAP2a,b), ${\beta}3$-tubulin and nestin as well as anti-inflammatory prostaglandin E2 receptor, EP3. The BBB scores showed significant motor recovery in group 3. GFP-labelled cells were localized on the injury site. In addition, decreased proinflammatory factor levels and increased intensity of anti-inflammatory factors were determined. Conclusion : Transplantation of PI-SCs might be an effective strategy to improve functional recovery following spinal cord trauma.

• IMMUNE NETWORK
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• v.23 no.6
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• pp.44.1-44.16
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• 2023

Mesenchymal stem cells (MSCs) are effective in treating autoimmune diseases and managing various conditions, such as engraftment of allogeneic islets. Additionally, autologous and HLA-matched allogeneic MSCs can aid in the engraftment of human allogeneic kidneys with or without low doses of tacrolimus, respectively. However, HLA alloantigens are problematic because cell therapy uses more HLA-mismatched allogeneic cells than autologous for convenience and standardization. In particular, HLA-mismatched MSCs showed increased Ag-specific T/B cells and reduced viability faster than HLA-matched MSCs. In CRISPR/Cas9-based cell therapy, Cas9 induce T cell activation in the recipient's immune system. Interestingly, despite their immunogenicity being limited to the cells with foreign Ags, the accumulation of HLA alloantigen-sensitized T/B cells may lead to allograft rejection, suggesting that alloantigens may have a greater scope of adverse effects than foreign Ags. To avoid alloantigen recognition, the β2-microglobulin knockout (B2MKO) system, eliminating class-I MHC, was able to avoid rejection by alloreactive CD8 T cells compared to controls. Moreover, universal donor cells in which both B2M and Class II MHC transactivator (CIITA) were knocked out was more effective in avoiding immune rejection than single KO. However, B2MKO and CIITA KO system remain to be controlled and validated for adverse effects such as the development of tumorigenicity due to deficient Ag recognition by CD8 T and CD4 T cells, respectively. Overall, better HLA-matching or depletion of HLA alloantigens prior to cell therapy can reduce repetitive transplantation through the long-term survival of allogeneic cell therapy, which may be especially important for patients seeking allogeneic transplantation.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.2
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• pp.153-160
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• 2017

In this study, we aim to determine the in vivo effect of human umbilical cord blood-derived multipotent stem cells (hUCB-MSCs) on neuropathic pain, using three, principal peripheral neuropathic pain models. Four weeks after hUCB-MSC transplantation, we observed significant antinociceptive effect in hUCB-MSC-transplanted rats compared to that in the vehicle-treated control. Spinal cord cells positive for c-fos, CGRP, p-ERK, p-p 38, MMP-9 and MMP 2 were significantly decreased in only CCI model of hUCB-MSCs-grafted rats, while spinal cord cells positive for CGRP, p-ERK and MMP-2 significantly decreased in SNL model of hUCB-MSCs-grafted rats and spinal cord cells positive for CGRP and MMP-2 significantly decreased in SNI model of hUCB-MSCs-grafted rats, compared to the control 4 weeks or 8weeks after transplantation (p<0.05). However, cells positive for TIMP-2, an endogenous tissue inhibitor of MMP-2, were significantly increased in SNL and SNI models of hUCB-MSCs-grafted rats. Taken together, subcutaneous injection of hUCB-MSCs may have an antinociceptive effect via modulation of pain signaling during pain signal processing within the nervous system, especially for CCI model. Thus, subcutaneous administration of hUCB-MSCs might be beneficial for improving those patients suffering from neuropathic pain by decreasing neuropathic pain activation factors, while increasing neuropathic pain inhibition factor.

• Clinical and Experimental Pediatrics
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• v.56 no.7
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• pp.298-303
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• 2013

Purpose: The purpose of this study was to evaluate short-term thyroid dysfunction and related risk factors in pediatric patients who underwent hematopoietic stem cell transplantation (HSCT) during childhood. Methods: We studied 166 patients (100 boys and 66 girls) who underwent HSCT at the Catholic HSCT Center from January 2004 through December 2009. The mean age at HSCT was $10.0{\pm}4.8$ years. Thyroid function of the patients was tested before and during 3 months of HSCT. Results: Out of 166 patients, 165 (99.4%) underwent allotransplantation. Acute graft-versus-host disease (GVHD, grades II to IV) developed in 76 patients. Conditioning regimens before HSCT include total body irradiation (n=57), busulfan (n=80), and reduced intensity (n=29). Forty-five (27.1%) had thyroid dysfunction during 3 months after HSCT (29 euthyroid sick syndrome [ESS], 6 subclinical hyperthyroidism, 4 subclinical hypothyroidism, 3 hypothyroxinemia, 2 overt hyperthyroidism, and 1 high $T_4$ syndrome). In a univariate logistic regression analysis, age at HSCT (P=0.002) and acute GVHD (P=0.009) had statistically significant relationships with thyroid dysfunction during 3 months after HSCT. Also, in a univariate logistic regression analysis, ESS (P=0.014) showed a strong statistically significant association with mortality. Conclusion: In our study 27.1% patients experienced thyroid dysfunction during 3 months after HSCT. Increase in age and acute GVHD may be risk factors for thyroid dysfunction during 3 months after HSCT. There was a significant association between ESS and mortality.

• Journal of Microbiology and Biotechnology
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• v.23 no.8
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• pp.1154-1158
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• 2013

Ganciclovir resistance of human cytomegalovirus is associated with mutations in the viral UL97 gene and poses severe problems for immunocompromised patients. In this study, PCR-based restriction fragment length polymorphism and sequencing analyses detected the UL97 D605E mutation in all five clinical isolates from patients with ganciclovir-resistant human cytomegalovirus infection during prolonged ganciclovir therapy, whereas the M460V mutation was only present in 1 of 5 isolates. On the other hand, the detection rates of the D605E mutation in the stored available DNA samples from the donor and allogeneic stem cell transplantation recipients were 66.7% and 93.7%, respectively, suggesting that the presence of D605E mutation was not associated with the ganciclovir exposure. Although the D605E mutation may not be related to ganciclovir resistance, we suggest that this mutation could be an important molecular marker of human cytomegalovirus evolution in East Asian countries. Moreover, the restriction fragment length polymorphism method using the restriction enzyme HaeIII, which is generally used to detect the UL97 A591V mutation, could also detect the D605E mutation and may therefore be a useful tool for future research on the investigation of UL97 gene mutations.

• Journal of Microbiology and Biotechnology
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• v.15 no.2
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• pp.259-264
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• 2005

Transplantation of cultured chondrocytes can regenerate cartilage tissues in cartilage defects in humans. However, this method requires a long culture period to expand chondrocytes to a large number of cells for transplantation. In addition, chondrocytes may dedifferentiate during long-term culture. These problems can potentially be overcome by the use of undifferentiated or partially developed cartilage precursor cells derived from neonatal cartilage, which, unlike chondrocytes from adult cartilage, have the capacity for rapid in vitro cell expansion and may retain their differentiated phenotype during long-term culture. The purpose of this study was to compare the cell growth rate and phenotypic modulation during in vitro culture between adult chondrocytes and neonatal chondrocytes, and to demonstrate the feasibility of regenerating cartilage tissues in vivo by transplantation of neonatal chondrocytes expanded in vitro and seeded onto polymer scaffolds. When cultured in vitro, chondrocytes isolated from neonatal (immediately postpartum, 2 h of age) rats exhibited much higher growth rate than chondrocytes isolated from adult rats. After 5 days of culture, more neonatal chondrocytes were in the differentiated state than adult chondrocytes. Cultured neonatal chondrocytes were seeded onto biodegradable polymer scaffolds and transplanted into athymic mice's subcutaneous sites. Four weeks after implantation, neonatal chondrocyte-seeded scaffolds formed white cartilaginous tissues. Histological analysis of the implants with hematoxylin and eosin showed mature and well-formed cartilage. Alcian blue/ safranin-O staining and Masson's trichrome staining indicated the presence of highly sulfated glycosarninoglycans and collagen, respectively, both of which are the major extracellular matrices of cartilage. Immunohistochemical analysis showed that the collagen was mainly type II, the major collagen type in cartilage. These results showed that neonatal chondrocytes have potential to be a cell source for cartilage tissue engineering.