Yong Man Kim

Adipose-derived stem cells as a new therapeutic modality for ageing skin.

Abstract:  Stem cells are undifferentiated cells, which have the important properties of self‐renewal and differentiation. Adipose‐derived stem cells (ADSC) have relative advantages in accessibility and abundance compared to other kinds of stem cells. Regeneration therapy using ADSC has received attention in the treatment of various dermatologic diseases. In previous studies, ADSC were shown to have antioxidant, whitening and wound‐healing effects in the skin through secretion of growth factors and by activating fibroblasts. In this study, we investigated whether ADSC could be used as an anti‐ageing therapy, especially by dermal collagen synthesis and angiogenesis. Subcutaneous injection of ADSC significantly increased collagen synthesis in hairless mice, and dermal thickness, collagen density and fibroblast number also increased. In addition, procollagen type I protein and mRNA expression increased, which accounts for the increased dermal collagen density. Angiogenesis, which was visualized by CD31 and NG2 immunofluorescence stains, also increased in ADSC‐treated skin. Our results suggest that ADSC therapy may be useful in ageing skin. Its effects are mainly mediated by stimulating collagen synthesis in dermal fibroblasts and increasing angiogenesis.

Histological improvement following administration of autologous bone marrow-derived mesenchymal stem cells for alcoholic cirrhosis: a pilot study

In experimental models, bone marrow‐derived mesenchymal stem cells (BM‐MSCs) have the capacity to differentiate into hepatocytes and exhibit antifibrotic effects. However, there have been no studies in humans with alcoholic cirrhosis.

The role of growth factors in maintenance of stemness in bone marrow-derived mesenchymal stem cells

Mesenchymal stem cells (MSCs) are an active topic of research in regenerative medicine due to their ability to secrete a variety of growth factors and cytokines that promote healing of damaged tissues and organs. In addition, these secreted growth factors and cytokines have been shown to exert an autocrine effect by regulating MSC proliferation and differentiation. We found that expression of EGF, FGF-4 and HGF were down-regulated during serial passage of bone marrow-derived mesenchymal stem cells (BMSCs). Proliferation and differentiation potentials of BMSCs treated with these growth factors for 2 months were evaluated and compared to BMSCs treated with FGF-2, which increased proliferation of BMSCs. FGF-2 and -4 increased proliferation potentials at high levels, about 76- and 26-fold, respectively, for 2 months, while EGF and HGF increased proliferation of BMSCs by less than 2.8-fold. Interestingly, differentiation potential, especially adipogenesis, was maintained only by HGF treatment. Treatment with FGF-2 rapidly induced activation of AKT and later induced ERK activation. The basal level of phosphorylated ERK increased during serial passage of BMSCs treated with FGF-2. The expression of LC3-II, an autophagy marker, was gradually increased and the population of senescent cells was increased dramatically at passage 7 in non-treated controls. But FGF-2 and FGF-4 suppressed LC3-II expression and down-regulated senescent cells during long-term (i.e. 2month) cultures. Taken together, depletion of growth factors during serial passage could induce autophagy, senescence and down-regulation of stemness (proliferation via FGF-2/-4 and differentiation via HGF) through suppression of AKT and ERK signaling.

Transplantation with autologous bone marrow-derived mesenchymal stem cells for alcoholic cirrhosis: Phase 2 trial.

Bone marrow‐derived mesenchymal stem cell (BM‐MSC) transplantation has been suggested as an effective therapy for liver cirrhosis. The efficacy and safety of autologous BM‐MSC transplantation in the treatment of alcoholic cirrhosis were investigated. Seventy‐two patients with baseline biopsy‐proven alcoholic cirrhosis who had been alcohol‐abstinent for more than 6 months underwent a multicenter, randomized, open‐label, phase 2 trial. Patients were randomly assigned to three groups: one control group and two autologous BM‐MSC groups that underwent either one‐time or two‐time hepatic arterial injections of 5 × 107 BM‐MSCs 30 days after BM aspiration. A follow‐up biopsy was performed 6 months after enrollment, and adverse events were monitored for 12 months. The primary endpoint was improvement in fibrosis quantification based on picrosirius red staining. The secondary endpoints included liver function tests, Child‐Pugh score, and Model for End‐stage Liver Disease score. Outcomes were analyzed by per‐protocol analysis. In terms of fibrosis quantification (before versus after), the one‐time and two‐time BM‐MSC groups were associated with 25% (19.5 ± 9.5% versus 14.5 ± 7.1%) and 37% (21.1 ± 8.9% versus 13.2 ± 6.7%) reductions in the proportion of collagen, respectively (P < 0.001). In the intergroup comparison, two‐time BM‐MSC transplantation in comparison with one‐time BM‐MSC transplantation was not associated with improved results in fibrosis quantification (P > 0.05). The Child‐Pugh scores of both BM‐MSC groups (one‐time 7.6 ± 1.0 versus 6.3 ± 1.3 and two‐time 7.8 ± 1.2 versus 6.8 ± 1.6) were also significantly improved following BM‐MSC transplantation (P < 0.05). The proportion of patients with adverse events did not differ among the three groups. Conclusion: Autologous BM‐MSC transplantation safely improved histologic fibrosis and liver function in patients with alcoholic cirrhosis. (Hepatology 2016;64:2185‐2197)

Adipose tissue-derived mesenchymal stem cells cultured at high density express IFN-β and suppress the growth of MCF-7 human breast cancer cells.

Although it has been reported that mesenchymal stem cells (MSCs) suppress tumor growth in vitro and in vivo, little is known about the underlying molecular mechanisms. We found that type I interferon is expressed in adipose tissue-derived stem cells (ASCs) cultured at high density, and ASCs and their conditioned medium (ASC-CM) suppress the growth of MCF-7 cells in vitro. Growth inhibition was amplified by glucose deprivation that resulted from high density culture of ASCs after 3days. The cytotoxic effect of the ASC-CM obtained from high density culture of ASCs was neutralized by anti-IFN-β antibody. STAT1 was phosphorylated in MCF-7 cells treated with ASC-CM, and JAK1/JAK2 inhibitor treatment decreased STAT1 phosphorylation. The cytotoxic effect of ASC-CM was reduced especially by JAK1 inhibitors in MCF-7 cells. Our findings suggest that ASCs cultured at high density express type I interferons, which suppresses tumor growth via STAT1 activation resulting from IFN-β secretion in MCF-7 breast cancer cells.

Cytosine deaminase-producing human mesenchymal stem cells mediate an antitumor effect in a mouse xenograft model

Background and Aim:  Stem cell transplantation offers potential gene therapy for brain tumors. However, this approach has received little attention as a treatment for gastrointestinal tumors. In the present study, we explored the possibility of human bone marrow‐derived mesenchymal stem cells (hMSC) producing cytosine deaminase (CD), followed by systemic 5‐fluorocytosine (5‐FC) administration, showing an antitumor effect on a mouse gastric cancer xenograft.

The effect of adipose stem cell therapy on pulmonary fibrosis induced by repetitive intratracheal bleomycin in mice

ABSTRACT Adipose stem cells (ASCs) are detectable in the parenchyma and large airways of lungs after systemic administration, and ameliorate inflammatory infiltration and cell death in animal models of emphysema. We evaluated whether ASC treatment could attenuate lung fibrosis induced by repetitive intratracheal bleomycin administration. Male 8-week-old C57BL/6J mice (control group, bleomycin-only group, and bleomycin-plus-ASC group) were used. Eight biweekly doses of bleomycin were injected intratracheally via an intubation procedure at a dose of 0.04 units in a total volume of 100 μL of sterile saline. During the latter 2 months of the 4-month bleomycin exposure, human ASCs (3 × 105 cells) were administered repeatedly via intraperitoneal injection at the same time as bleomycin. Lung tissues were evaluated for histology, collagen content, TUNEL staining, and TGF-β levels. Bronchoalveolar lavage (BAL) was performed for cell counting. Administrations of ASCs ameliorated the deleterious effects of repetitive intratracheal instillation of bleomycin, namely hyperplasia of Club cells (Clara cells) and cuboidal alveolar epithelial cells, infiltration of the perialveolar ducts by inflammatory cells, septal thickening, enlarged alveoli, and extensive fibrosis. Addition of ASC led to suppression of bleomycin-induced epithelial cell apoptosis and expression of TGF-β. These results suggest a useful therapeutic effect of ASCs on pulmonary fibrosis induced by repetitive bleomycin administration. Further studies will be required to evaluate the efficacy of ASC therapy for the treatment of idiopathic pulmonary fibrosis.

l-Ascorbic acid 2-phosphate and fibroblast growth factor-2 treatment maintains differentiation potential in bone marrow-derived mesenchymal stem cells through expression of hepatocyte growth factor

Abstract l-ascorbic acid 2-phosphate (Asc-2P) acts as an antioxidant and a stimulator of hepatocyte growth factor (HGF) production. Previously, we reported that depletion of growth factors such as fibroblast growth factor (FGF)-2, epidermal growth factor (EGF), FGF-4 and HGF during serial passage could induce autophagy, senescence and down-regulation of stemness (proliferation via FGF-2/-4 and differentiation via HGF). In this study, we investigated the proliferation and differentiation potential of BMSCs by FGF-2 and Asc-2P. Co-treatment with FGF-2 and Asc-2P induced optimal proliferation of BMSCs and increased the accumulation rate of BMSC numbers during a 2-month culture period. Moreover, differentiation potential was maintained by co-treatment with FGF-2 and Asc-2P via HGF expression. Adipogenic differentiation potential by FGF-2 and Asc-2P was dramatically suppressed by c-Met inhibitors (SU11274). These data suggest that co-treatment with FGF-2 and Asc-2P would be beneficial in obtaining BMSCs that possess “stemness” during long-term culture.

VEGF therapeutic gene delivery using dendrimer type bio-reducible polymer into human mesenchymal stem cells (hMSCs).

The therapeutic potential of mesenchymal stem cells (MSCs) has garnered great attention in the expansive diversity of biomedical research. Despite this broad interest in stem cells, limited incorporation and poor viability are major disadvantages for accomplishing therapeutic success in the field of hMSC-based cell therapy, and an optimal approach for hMSC-based cell therapy using non-viral vectors has not been established. Hence, we examined the possibility of performing gene therapy using the biodegradable polymeric non-viral vector Arginine-grafted poly (cystaminebisacrylamide-diaminohexane) (ABP)-conjugated poly (amidoamine) (PAMAM) dendrimer (PAM-ABP) in hMSCs. PAM-ABP formed compact nanosized polyplexes and showed low cytotoxicity compared to bPEI 25k and Lipofectamine® 2000 in hMSCs. Although the cellular uptake was similar, the transfection efficiency and VEGF expression of PAM-ABP using gWiz-Luc and pβ-VEGF were higher than those of the control groups. Although hMSCs were transfected, their stem cell characteristics were retained. Our results suggest that PAM-ABP has the ability to deliver a therapeutic gene in hMSCs.

Adipose tissue-derived mesenchymal stem cells cultured at high cell density express brain-derived neurotrophic factor and exert neuroprotective effects in a 6-hydroxydopamine rat model of Parkinson’s disease

Mesenchymal stem cells (MSCs) secrete neurotrophic factors, and have been reported to improve functional outcomes in animal models of neurodegenerative diseases such as cerebral ischemia, stroke, spinal cord lesions, and Parkinson’s disease. Previously, we found that adipose tissue-derived mesenchymal stem cells (ASCs) cultured at high cell density (HD-ASCs) expressed interferon-beta (IFN-β). Here we demonstrate that ASCs expressing IFN-β also express brain-derived neurotrophic factor (BDNF). Growth rates of neuroblastoma cells (SK-N-BE(2)C) were increased when co-cultured with HD-ASCs or treated with concentrated medium obtained from HD-ASCs (HD-ASC-CM). The HD-ASC-CM induced AKT phosphorylation in SK-N-BE(2)C cells, and AKT inhibition by Ly294002 reduced cell viability of SK-N-BE(2)C cells. Additionally, a protective effect on SK-N-BE(2)C cells exposed to 6-hydroxydopamine (6-OHDA) was observed in the HD-ASC-CM or brain-derived neurotrophic factor (BDNF) treated cells. The protective effect of the HD-ASC-CM was neutralized by anti-BDNF antibody. In the 6-OHDA-induced Parkinson’s disease rat model, ASCs reduced amphetamine-induced rotations and a greater number of tyrosine hydroxylase (TH)-positive cells were observed in the HD-ASCs-injected group compared with sham controls and the low density cultured ASC-injected group. Moreover, the expression of BDNF, nerve growth factor (NGF), TH, and proliferating cell nuclear antigen (PCNA) in ipsilateral midbrain tissues including substantia nigra pars compacta (SNc) was increased by transplantation of HD-ASCs. These data indicate that HD-ASCs may induce neuroprotective effects through BDNF expression and subsequent increase of proliferation in neuronal cells both in vitro and in vivo.