• Development and Reproduction
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• v.13 no.3
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• pp.173-181
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• 2009

One of the most extensively studied populations of multipotent adult stem cells are mesenchymal stem cells (MSCs). MSCs derived from the human umbilical cord vein (HUC-MSCs) are morphologically and immunophenotypically similar to MSCs isolated from bone marrow. HUC-MSCs are multipotent stem cells, differ from hematopoietic stem cells and can be differentiated into neural cells. Since neural tissue has limited intrinsic capacity of repair after injury, the identification of alternate sources of neural stem cells has broad clinical potential. We isolated mesenchymal-like stem cells from the human umbilical cord vein, and studied transdifferentiation-promoting conditions in neural cells. Dopaminergic neuronal differentiation of HUC-MSCs was also studied. Neural differentiation was induced by adding bFGF, EGF, dimethyl sulfoxide (DMSO) and butylated hydroxyanisole (BHA) in N2 medium and N2 supplement. The immunoreactive cells for $\beta$-tubulin III, a neuron-specific marker, GFAP, an astrocyte marker, or Gal-C, an oligodendrocyte marker, were found. HUC-MSCs treated with bFGF, SHH and FGF8 were differentiated into dopaminergic neurons that were immunopositive for tyrosine hydroxylase (TH) antibody. HUC-MSCs treated with DMSO and BHA rapidly showed the morphology of multipolar neurons. Both immunocytochemistry and RT-PCR analysis indicated that the expression of a number of neural markers including NeuroD1, $\beta$-tubulin III, GFAP and nestin was markedly elevated during this acute differentiation. While the stem cell markers such as SCF, C-kit, and Stat-3 were not expressed after neural differentiation, we confirmed the differentiation of dopaminergic neurons by TH/$\beta$-tubulin III positive cells. In conclusion, HUC-MSCs can be differentiated into dopaminergic neurons and these findings suggest that HUC-MSCs are alternative cell source of therapeutic treatment for neurodegenerative diseases.

• Journal of Acupuncture Research
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• v.22 no.2
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• pp.93-101
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• 2005

Objective: We have evaluated UDHA into the joint for its effectiveness on immune responses to CII in the rat CIA. In an attempt to gain further insight into the mode of action of UDHA, we also investigated the effects of UDHA on the incidence and development of arthritis in rat CIA with 2 different regimens: (1) started prior to a primary immunization, (2) started on the day of a primary immunization. Methods : Male rats were immunized with an emulsion of $200\;{\mu}g/100g$ of CII and complete Freund's adjuvant (CFA). The rats were then given intraperitoneal(i.p) stimulation of Ulmus davidiana Planch herbal acupuncture(UDHA) or saline during the experiment. Lymph node cells were obtained from rats 14 days after immunization and cultured in vitro with CII. When compared with rats treated with saline as control, UDHA at doses of more than $20\;{\mu}g/100\;g$ rat once a day for 7 days inhibited the ability of inguinal lymph node cells to produce T cell cytokines interleukin-$1{\beta}$, tumor necrosis $factor-{\alpha}$ $(TNF-{\alpha})$. When rats were injected intraperitoneally with SRBC, hemaglutination titers in UD-treated and control rats did not differ significantly when low doses of UD was given to rats. However, i.p injection of UD at doses of more than $10\;{\mu}g/100\;g/day$ for 7 days slightly suppressed antibody production. Results : The present results show that treatment with UDHA can inhibit the onset and development of arthritis and the immune responses to collagen. Conclusion: Therapeutic i.p injection with UD affect the clinical course of the disease and the immune responses to CII.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.39
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• pp.8.1-8.8
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• 2017

Background: For an effective bone graft for reconstruction of the maxillofacial region, an adequate vascular network will be required to supply blood, osteoprogenitor cells, and growth factors. We previously reported that the secretomes of bone marrow-derived mesenchymal stem cells (MSC-CM) contain numerous growth factors such as insulin-like growth factor (IGF)-1, transforming growth factor $(TGF)-{\beta}1$, and vascular endothelial growth factor (VEGF), which can affect the cellular characteristics and behavior of regenerating bone cells. We hypothesized that angiogenesis is an important step for bone regeneration, and VEGF is one of the crucial factors in MSC-CM that would enhance its osteogenic potential. In the present study, we focused on VEGF in MSC-CM and evaluated the angiogenic and osteogenic potentials of MSC-CM for bone regeneration. Methods: Cytokines in MSC-CM were measured by enzyme-linked immunosorbent assay (ELISA). Human umbilical vein endothelial cells (HUVECs) were cultured with MSC-CM or MSC-CM with anti-VEGF antibody (MSC-CM + anti-VEGF) for neutralization, and tube formation was evaluated. For the evaluation of bone and blood vessel formation with micro-computed tomography (micro-CT) and for the histological and immunohistochemical analyses, a rat calvarial bone defect model was used. Results: The concentrations of IGF-1, VEGF, and $TGF-{\beta}1$ in MSC-CM were $1515.6{\pm}211.8pg/mL$, $465.8{\pm}108.8pg/mL$, and $339.8{\pm}14.4pg/mL$, respectively. Tube formation of HUVECs, bone formation, and blood vessel formation were increased in the MSC-CM group but decreased in the MSC-CM + anti-VEGF group. Histological findings suggested that new bone formation in the entire defect was observed in the MSC-CM group although it was decreased in the MSC-CM + anti-VEGF group. Immunohistochemistry indicated that angiogenesis and migration of endogenous stem cells were much more abundant in the MSC-CM group than in the MSC-CM + anti-VEGF group. Conclusions: VEGF is considered a crucial factor in MSC-CM, and MSC-CM is proposed to be an adequate therapeutic agent for bone regeneration with angiogenesis.

• Korean Journal of Acupuncture
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• v.24 no.3
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• pp.179-204
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• 2007

Objectives & Methods : The purpose of this study is to observe the effects of Angelicae Radix Pharmacopuncture(AR-P) at Joksamni(ST36) on collagen II induced arthritis in DBA/1J mice. The authors evaluated arthritis index, arthritis incidence and joint edema, and measured body weight, spleen size and stenosis rate, serum cytokine level, serum antibody level, immune cell populations In spleen, lymph node, and knee joint, and performed histological analysis of CIA mouse joint. Results : In the AR-P group, arthritis index, arthritis incidence and joint edema were decreased, and the enlargement, malformation and stenosis of spleen and the malformation of joint appeared milder than the control group. In AR-P group, the levels of $IL-1{\beta}$, IL-6, $TNF-{\alpha}$, $IFN-{\gamma}$ in serum were significantly decreased. And the level of anti-collagen II in serum was maintained lower in AR-P group than in the control group. In AR-P group, the level of $IFN-{\gamma}$ and IL-10, and $IFN-{\gamma}/IL-4$ ratio were significantly decreased, and the ratios of $CD3e^+$ cells to $CD45^+$ cells, $CD4^+$ cells to $CD8^+$ cells in spleen were similarly maintained as those of the normal group. In the AR-P group, the populations of $CD4^+/CD25^+$ cells in spleen and lymph nodes, $CD45R^+/CD69^+$ cells in lymph nodes, $CD3^+/CD69^+$ cells and $CD11b^+/Gr-1^+$ cells in knee joint were decreased. In the histological analysis, the cartilage destruction, synovial cell proliferation and the collagen fiber destruction were decreased in the AR-P group Conclusions : The results suggest that AR-P at right ST36 has a therapeutic effect on collagen-induced arthritis in mice.

• Journal of Physiology & Pathology in Korean Medicine
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• v.21 no.4
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• pp.982-991
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• 2007

The present study was done to assess the suppressive effects of Haedongpi-san(HDPS), a traditional herbal medicine, on collagen induced arthritis (CIA) in mice and to examined it's effects on immune system. Oral administration of HDPS (200 or 400 mg/Kg) significantly suppressed the progression of CIA, which extend is comparable to that of methotrexate (MTX, 30 mg/Kg), a positive control. Histological examinations reveled that HDPS inhibited infiltration of inflammatory cells into affected paw joint, and bone erosion and cartilage destruction were greatly reduced compared with control. In paw joint, the number of CD3+ cells and CD11b+/Gr-1+ cells were greatly reduced by HDPS. The levels of pathologic cytokines including TNF-a and IL-6 were significantly decreased in the serum by oral treatment with HDPS. The levels of $IFN-{\gamma}$ in the culture supernatant of splenocyte stimulated with CD3/CD28 or collagen were dramatically decreased, while those of IL-4 was increased. Rheumatoid factors including IgG, IgM and collagen specific antibody were present much lower in the serum of HDPS treated mice than control. In peripheral blood mononuclear cells of HDPS treated mice, the percentage of CD3+, CD3+/CD69+, CD4+, CD4+/CD25+ cells were significantly decreased, while CD19+ cells were slightly increased compared with control. The absolute number of CD19+, CD3+, CD3+/CD69+, CD4+/CD25+, CD49b+ cell in spleen from HDPS treated mice were significantly decreased. The absolute number of CD3+, CD3+/CD69+, CD4+, CD4+/CD25+ CD8+, CD49b+, CD3+/CD49b+ cells in draining lymph node were significantly increased compared with control. Taken together, HDPS has suppressive effects on rheumatoid arthritis by modulating immune system, and has potential to use as an therapeutic for rheumatoid arthritis.

• Journal of Biomedical Engineering Research
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• v.15 no.1
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• pp.41-50
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• 1994

The chemotherapy using macromolecules, i.e., monoclonal antibodies loaded with anticancer agents hasn't been successful in delivering therapeutic amount of the conjugates. The comprehensive evaluation of mass transfer factors in tumor is prerequisite for the development of the effective chemotherapy. Characterization of neuroblastoma implanted in immunosuppressed athymic nude rats was performed. Its growth kinetics, glucose metabolic rate (GMR) were measured along with the interstitial fluid pressure (IFP), blood perfusion rate (BPR) and pH distribution throughtout the tumor radius. Volume doubling time and GMR were 8.1 days(SD 0.44 day), 23.53 mg/min/100 g(SD 3.54 mg/min/100 g), respectively. The IFP in tumor center was increased with tumor volume, and approached to 3 mmHg (SD 2.6 mmHg) when the tumor was 3 cm high. The radial distribution of IFP, BPR and pH in 2 cm high tumors showed that BPR and pH were decreased, while IFP was increased as the ~ensors moved toward the tumor center. The elevated IFP, decreased BPR and pH in tumor center suggested that the delivery of conjugates might be increased by properly manipulating mass transfer factors.

• Archives of Plastic Surgery
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• v.37 no.6
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• pp.721-725
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• 2010

Purpose: The survival of composite graft is dependent on three steps, (1) plasmatic imbibitions, (2) inosculation, and (3) neovascularization. Among the many trials to increase the survival rate of composite graft, prostaglandin E1 (PGE1) has beneficial effects on the microcirculatory level with vasodilating, antithrombotic, anti-inflammatory and neoangiogenic properties. Lipo-PGE1 which is lipid microspheres containing PGE1 had developed to compensate the systemic and local side effects of PGE1. This study was proposed to determine whether Lipo-PGE1 administration enhanced the survival of composite graft through neovascularization quantitatively in a rabbit ear model. Methods: Fourteen New Zealand White Rabbits each weighing 3~4 kg were divided in two groups: (1) intravenous Lipo-PGE1 injection group and (2) control group. A $2{\times}1\;cm$ sized, full-thickness rectangular composite graft was harvested in each auricle. Then, the graft was reaaproximated in situ using a 5-0 nylon suture. For the experimental group, $3{\mu}g$/kg/day of Lipo-PGE1 ($5{\mu}g$/mL) was administered intravenously through the marginal vein of the ear for 14 days. The control group was received no pharmacologic treatment. On the 14th postoperative day, composite graft of the ear was harvested and immunochemistry staining used Monoclonal mouse anti-CD 31 antibody was performed. Neoangiogenesis was quantified by counting the vessels that showed luminal structures surrounded by the brown color-stained epithelium and counted from 10 random high-power fields (400x) by independent blinded observer. Statistical analysis (Wilcoxon Signed Ranks test for nonparametric data) was performed using SPSS v12.0, with values of p<0.05 considered significant. Results: The mean number of the microvessels was $15.48{\pm}8.65$ in the experimental group and $9.82{\pm}7.25$ in the control group (p=0.028). Conclusion: The use of Lipo-PGE1 facilitated the neoangiogenesis, resulted in the improvement of the survival rate of graft. On the basis of this results, we could support wider application of Lipo-PGE1 for more effective therapeutic angiogenesis and successful survival in various cases of composite graft in the human.

• Molecules and Cells
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• v.37 no.3
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• pp.226-233
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• 2014

Excessive reactive oxygen species (ROS) generated from abnormal cellular process lead to various human diseases such as inflammation, ischemia, and Parkinson's disease (PD). Sensitive to apoptosis gene (SAG), a RING-FINGER protein, has anti-apoptotic activity and anti-oxidant activity. In this study, we investigate whether Tat-SAG, fused with a Tat domain, could protect SH-SY5Y neuroblastoma cells against 1-methyl-4-phenylpyridinium ($MPP^+$) and dopaminergic (DA) neurons in the substantia nigra (SN) against 1-methyl-4-phenyl-1,2,3,6-tetra-hydropyridine (MPTP) toxicity. Western blot and immunohistochemical analysis showed that, unlike SAG, Tat-SAG transduced efficiently into SH-SY5Y cells and into the brain, respectively. Tat-SAG remarkably suppressed ROS generation, DNA damage, and the progression of apoptosis, caused by $MPP^+$ in SH-SY5Y cells. Also, immunohistochemical data using a tyrosine hydroxylase antibody and cresyl violet staining demonstrated that Tat-SAG obviously protected DA neurons in the SN against MPTP toxicity in a PD mouse model. Tat-SAG-treated mice showed significant enhanced motor activities, compared to SAG- or Tat-treated mice. Therefore, our results suggest that Tat-SAG has potential as a therapeutic agent against ROS-related diseases such as PD.

• Journal of Microbiology and Biotechnology
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• v.29 no.7
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• pp.1165-1176
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• 2019

Botulinum neurotoxins (BoNTs), produced by Clostridium botulinum, are the most toxic substances known. However, the number of currently approved medical countermeasures for these toxins is very limited. Therefore, studies on therapeutic antitoxins are essential to prepare for toxin-related emergencies. Currently, more than 10,000 Halla horses, a crossbreed between the native Jeju and Thoroughbred horses, are being raised in Jeju Island of Korea. They can be used for equine antitoxin experiments and production of hyperimmune serum against BoNT/A1. Instead of the inactivated BoNT/A1 toxoid, Halla horse was immunized with the receptor-binding domain present in the C-terminus of heavy chain of BoNT/A1 (BoNT/A1-HCR) expressed in Escherichia coli. The anti-BoNT/A1-HCR antibody titer increased rapidly by week 4, and this level was maintained for several weeks after boosting immunization. Notably, $20{\mu}l$ of the week-24 BoNT/A1-HCR(-immunized) equine serum showed an in vitro neutralizing activity of over 8 international units (IU) of a reference equine antitoxin. Furthermore, $20{\mu}l$ of equine serum and $100{\mu}g$ of purified equine $F(ab^{\prime})_2$ showed 100% neutralization of 10,000 $LD_{50}$ in vivo. The results of this study shall contribute towards optimizing antitoxin production for BoNT/A1, which is essential for emergency preparedness and response.

• Tuberculosis and Respiratory Diseases
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• v.46 no.2
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• pp.229-240
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• 1999

Background: The impact of the immune response on cancer gene therapy using viral vectors to deliver a "suicide gene" is currently unclear. A vigrous immune response targeted at viral proteins or transgene may enhance the efficacy of tumor destruction and even augment responses to tumor antigens. These responses may involve the release of cytokines and stimulation of tumor specific cytotoxic T-lymphocytes that enhance therapeutic efficacy. On the other hand, a vigorous rapid cellular immune response may destroy cells expressing the therapeutic gene and attenuate the response to therapy. Furthermore, development of neutralizing antibody responses may prevent readministration of virus, a potentially significant limitation. Evaluating the significance of these limitations in animal models and developing solutions are therefore of obvious importance. Methods: After retroviral transduction of mouse mesothelioma cell line(AB12) with Herpes Simplex Virus thymidine kinase (HSVtk) gene in vitro, subcutaneous flank tumors were established. To study the effect of intact immune system on efficacy of tumor erradication, the ability of the HSVtk/ganciclovir system to inhibit tumor growth was compared among normal Balb/c mice, immunodeficient Balb/c-nude and SCID mice, and Balb/c mice immunosuppressed with cyclosporin. Results: Ganciclovir treatment resulted in greater inhibition of tumor growth in Balb/c mice compared with immunodeficient Balb/c-nude mice and SCID mice(in immunodeficient mice, there were no growth inhibition by ganciclovir treatment). Ganciclovir treatment resulted in greater inhibition of tumor growth in noncyclosporin (CSA) treated Balb/c mice compared with CSA treated Balb/c mice. On day 8, mean ganciclovir-treated tumor volume were 65% of control tumor volume in Balb/c mice versus 77% control tumor volume in CSA-treated Balb/c mice. This effect was still evident during therapy (day 11 and 13). On day 13, non-CSA treated tumor volume was 35% of control tumor volume versus 60% of control tumor volume in CSA treated Balb/c mice. Duration of expression of HSVtk was not affected by the immunosuppression with CSA. Conclusion: These results indicate that the immune responses against retrovirally transduced cells enhance the efficacy of the HSVtk/ganciclovir system. These findings have important implications for clinical trials using currently available retrovirus vectors as well as for future vector design.