Hee Woo Lee

Transplantation of CTLA4Ig gene-transduced adipose tissue-derived mesenchymal stem cells reduces inflammatory immune response and improves Th1/Th2 balance in experimental autoimmune thyroiditis.

Autoimmune thyroiditis is one of common organ‐specific autoimmune disease. The aim of this study was to observe the effect of adipose tissue derived mesenchymal stem cells (ATMSC) and CTLA4Ig gene‐transduced ATMSC on autoimmune thyroiditis.

Anti-Tumor Effect of Adipose Tissue Derived-Mesenchymal Stem Cells Expressing Interferon-β and Treatment with Cisplatin in a Xenograft Mouse Model for Canine Melanoma

Adipose tissue-derived mesenchymal stem cells (AT-MSCs) are attractive cell-therapy vehicles for the delivery of anti-tumor molecules into the tumor microenvironment. The innate tropism of AT-MSCs for tumors has important implications for effective cellular delivery of anti-tumor molecules, including cytokines, interferon, and pro-drugs. The present study was designed to determine the possibility that the combination of stem cell-based gene therapy with low-dose cisplatin would improve therapeutic efficacy against canine melanoma. The IFN-β transduced canine AT-MSCs (cAT-MSC-IFN-β) inhibited the growth of LMeC canine melanoma cells in direct and indirect in vitro co-culture systems. In animal experiments using BALB/c nude mouse xenografts, which developed by injecting LMeC cells, the combination treatment of cAT-MSC-IFN-β and low-dose cisplatin significantly reduced tumor volume compared with the other treatment groups. Fluorescent microscopic analysis with a TUNEL (terminal deoxynucleotidyl transferase-mediated nick-end labeling) assay of tumor section provided evidence for homing of cAT-MSC-IFN-β to the tumor site and revealed that the combination treatment of cAT-MSC-IFN-β with low-dose cisplatin induced high levels of cell apoptosis. These findings may prove useful in further explorations of the application of these combined approaches to the treatment of malignant melanoma and other tumors.

Therapeutic effects of hepatocyte growth factor-overexpressing human umbilical cord blood-derived mesenchymal stem cells on liver fibrosis in rats

Fibrosis is a common end stage for a variety of liver diseases, including most chronic liver diseases, and results from an imbalance between collagen deposition and degradation. Mesenchymal stem cells (MSCs) have the ability to migrate into fibrotic livers and differentiate into hepatocytes. Hepatocyte growth factor (HGF) has potent anti‐apoptotic and mitogenic effects on hepatocytes during liver injury and plays an essential role in the development and regeneration of the liver. In this study, human HGF‐overexpressing human umbilical cord blood‐derived MSCs (hHGF‐HUCB‐MSCs) were prepared using the pMEX Expression System, and the upregulation of hHGF expression was confirmed by RT‐PCR and ELISA. HGF expressed by hHGF‐HUCB‐MSCs exerted a stimulatory effect on hepatocyte proliferation in vitro. hHGF‐HUCB‐MSCs were transplanted to investigate the therapeutic effects of these cells on carbon tetrachloride (CCL4)‐induced liver fibrosis in a rat model. After 4 weeks of cell treatment once per week with 2 × 106 cells, biochemical analysis of the serum and histopathological analysis of the liver tissue were performed. The results of the biochemical analysis of the serum show that the hHGF‐HUCB‐MSC‐treated group had higher levels of alanine aminotransferase, aspartate aminotransferase and alkaline phosphatase, indicating the improvement of liver function. Histopathology showed that the hHGF‐HUCB‐MSC‐treated group had reduction in the density of collagen fibres. Thus hHGF‐HUCB‐MSCs can enhance liver regeneration and could be useful for the treatment of patients with liver fibrosis or cirrhosis.

Genistein inhibits pro‑inflammatory cytokines in human mast cell activation through the inhibition of the ERK pathway

Anaphylaxis is a rapidly occurring allergic reaction to any foreign substance, including venom from insects, foods and medications, which may cause fatalities. To prevent anaphylaxis, these triggers must be avoided. However, avoidance of numerous triggers is difficult. For this reason, the development of immunotherapeutic adjuvants that suppress the allergic response is important for anaphylaxis control. Mast cells are one of the major inflammatory cells involved in the inflammatory response, which secrete several inflammatory cytokines, including tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β, and recruits other immune cells. Mast cells are also involved in a number of diseases, such as sinusitis, rheumatoid arthritis and asthma. Genistein, a phytoestrogen, has been reported to have anti-oxidative and anti-inflammatory activities. However, the effects of genistein on the anti-inflammatory response of mast cells remain unknown. In the present study, the anti-inflammatory effects of genistein on mast cells were investigated. Genistein significantly decreased IL-6 and IL-1β mRNA levels, as well as IL-6 production in PMA/A23187-induced mast cells activation. In addition, genistein inhibited the phosphorylation of ERK 1/2 in PMA/A23187-induced mast cell activation. However, phosphorylation of p38 was not altered. Thus, these findings indicate that genistein inhibited the inflammatory status of mast cells through inhibition of the ERK pathway.

Comparative Efficacies of Long-Term Serial Transplantation of Syngeneic, Allogeneic, Xenogeneic, or CTLA4Ig-Overproducing Xenogeneic Adipose Tissue-Derived Mesenchymal Stem Cells on Murine Systemic Lupus Erythematosus.

Allogeneic and xenogeneic transplantation are suitable alternatives for treating patients with stem cell defects and autoimmune diseases. The purpose of this study was to compare the effects of long-term serial transplantation of adipose tissue-derived mesenchymal stem cells (ASCs) from (NZB × NZW) F1 mice (syngeneic), BALB/c mice (allogeneic), or humans (xenogeneic) on systemic lupus erythematosus (SLE). The effects of transplanting human ASCs overproducing CTLA4Ig (CTLA4Ig-hASC) were also compared. Animals were divided into five experimental groups, according to the transplanted cell type. Approximately 500,000 ASCs were administered intravenously every 2 weeks from 6 to 60 weeks of age to all mice except for the control mice, which received saline. The human ASC groups (hASC and CTLA4Ig-hASC) showed a 13-week increase in average life spans and increased survival rates and decreased blood urea nitrogen, proteinuria, and glomerular IgG deposition. The allogeneic group also showed higher survival rates compared to those of the control, up to 40, 41, 42, 43, 44, 45, 52, and 53 weeks of age. Syngeneic ASC transplantation did not accelerate the mortality of the mice. The mean life span of both the syngeneic and allogeneic groups was prolonged for 6–7 weeks. Both human ASC groups displayed increased serum interleukin-10 and interleukin-4 levels, whereas both mouse ASC groups displayed significantly increased GM-CSF and interferon-γ levels in the serum. The strongest humoral immune response was induced by xenogeneic transplantation, followed by allogeneic, CTLA4Ig-xenogeneic, and syngeneic transplantations. Long-term serial transplantation of the ASCs from various sources displayed different patterns of cytokine expression and humoral responses, but all of them increased life spans in an SLE mouse model.

Antitumor effects of celecoxib in COX-2 expressing and non-expressing canine melanoma cell lines.

Cyclooxygenase-2 (COX-2) is a potential target for chemoprevention and cancer therapy. Celecoxib, a selective COX-2 inhibitor, inhibits cell growth of various types of human cancer including malignant melanoma. In dogs, oral malignant melanoma represents the most common oral tumor and is often a fatal disease. Therefore, there is a desperate need to develop additional therapeutic strategies. The purpose of this study was to investigate the anticancer effects of celecoxib on canine malignant melanoma cell lines that express varying levels of COX-2. Celecoxib induced a significant anti-proliferative effect in both LMeC and CMeC-1 cells. In the CMeC cells, treatment of 50 μM celecoxib caused an increase in cells in the G0/G1 and a decreased proportion of cells in G-2 phase. In the LMeC cells, 50 μM of celecoxib led to an increase in the percentage of cells in the sub-G1 phase and a significant activation of caspase-3 when compared to CMeC-1 cells. In conclusion, these results demonstrate that celecoxib exhibits antitumor effects on canine melanoma LMeC and CMeC-1 cells by induction of G1-S cell cycle arrest and apoptosis. Our data suggest that celecoxib might be effective as a chemotherapeutic agent against canine malignant melanoma.

Therapeutic effects of CTLA4Ig gene-transduced adipose tissue-derived mesenchymal stem cell transplantation on established autoimmune thyroiditis.

This study aimed to identify the beneficial effects of adipose tissue-derived mesenchymal stem cells (ASCs) and ASCs that overexpress the CTLA4Ig gene (CTLA4Ig-ASCs) on established autoimmune thyroiditis and to examine changes in clinical chemistry parameters and the presence of humoral responses upon repeated long-term administration of autologous ASCs. This study also aimed to acquire desirable results in a preclinical study by using large-sized lab animals and applying ASCs that overexpress therapeutic genes. Experimental autoimmune thyroiditis was induced by immunization with thyroglobulin. Experimental dogs were divided into five groups: (i) ASC IT + IV, (ii) ASC IV, (iii) CTLA4Ig-ASC IT + IV, (iv) CTLA4Ig-ASC IV, and (v) control IT + IV (saline only), and they received intrathyroidal (IT; 10 million cells/250 μl saline per thyroid) administration one time or intravenous (IV; 20 million cells/5 ml) administration seven times within a 101-day period. Blood samples were collected every week, and thyroids were harvested on days 104-106. After serial ASC or CTLA4Ig transplantation, the levels of canine thyroglobulin autoantibodies (TgAA) in serum and the infiltration of T-lymphocytes between the follicles of the thyroid glands were decreased. The expression of FoxP3 in submandibular lymph nodes was significantly increased. Repeated long-term administration of autologous ASCs or CTLA4Ig-ASCs did not generate changes in clinical chemistry parameters or humoral responses. The TgAA test can detect autoimmune thyroiditis years before clinical signs of hypothyroidism occur. Thus, ASC and CTLA4Ig-ASC transplantation in that period can be attractive candidates to ameliorate autoimmune thyroiditis and prevent the development of hypothyroidism.

Stimulatory effect of HGF-overexpressing adipose tissue-derived mesenchymal stem cells on thymus regeneration in a rat thymus involution model.

The thymus is the central lymphoid organ providing a unique and essential microenvironment for T‐cell precursor development into mature functionally competent T‐lymphocytes. Thus, it is important to develop the strategies for enhancing thymic regeneration from involution induced by a variety of clinical treatments and conditions. Hepatocyte growth factor (HGF) promotes proliferation in a variety of cell types. We have used stem cell‐based HGF gene therapy to enhance regeneration from acute thymic involution. HGF‐overexpressing human adipose tissue‐derived mesenchymal stem cells (HGF‐hATMSCs) were generated by liposomal transfection with the pMEX expression vector, constructed by inserting the HGF gene. Significantly increased HGF expression in these cells was confirmed by reverse transcription‐polymerase chain reaction and an enzyme‐linked immunosorbent assay. HGF produced by HGF‐hATMSCs enhanced the proliferation of a mouse thymic epithelial cell line and the expression of interleukin‐7 in vitro. We also examined the effect of HGF‐hATMSCs on thymic regeneration in rats with acute thymic involution. Significant increases in thymus size and weight, as well as the number of thymocytes (especially, early thymocyte progenitors), were seen in the HGF‐hATMSCs‐treated rats compared to saline‐treated control animals. A stimulatory effect of HGF‐hATMSCs on thymic regeneration has therefore been shown, highlighting the clinical value of HGF‐hATMSCs for treating thymic involution.