• 대한두개안면성형외과학회지
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• 제19권3호
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• pp.181-189
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• 2018

Background: Autogenous bone grafts have several limitations including donor-site problems and insufficient bone volume. To address these limitations, research on bone regeneration is being conducted actively. In this study, we investigate the effects of a three-dimensionally (3D) printed polycaprolactone (PCL)/tricalcium phosphate (TCP) scaffold on the osteogenic differentiation potential of adipose tissue-derived stem cells (ADSCs) and bone marrow-derived stem cells (BMSCs). Methods: We investigated the extent of osteogenic differentiation on the first and tenth day and fourth week after cell culture. Cytotoxicity of the 3D printed $PCL/{\beta}-TCP$ scaffold was evaluated by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay, prior to osteogenic differentiation analysis. ADSCs and BMSCs were divided into three groups: C, only cultured cells; M, cells cultured in the 3D printed $PCL/{\beta}-TCP$ scaffold; D, cells cultured in the 3D printed $PCL/{\beta}-TCP$ scaffold with a bone differentiation medium. Alkaline phosphatase (ALP) activity assay, von Kossa staining, reverse transcription-polymerase chain reaction (RT-PCR), and Western blotting were performed for comparative analysis. Results: ALP assay and von Kossa staining revealed that group M had higher levels of osteogenic differentiation compared to group C. RT-PCR showed that gene expression was higher in group M than in group C, indicating that, compared to group C, osteogenic differentiation was more extensive in group M. Expression levels of proteins involved in ossification were higher in group M, as per the Western blotting results. Conclusion: Osteogenic differentiation was increased in mesenchymal stromal cells (MSCs) cultured in the 3D printed PCL/TCP scaffold compared to the control group. Osteogenic differentiation activity of MSCs cultured in the 3D printed PCL/TCP scaffold was lower than that of cells cultured on the scaffold in bone differentiation medium. Collectively, these results indicate that the 3D printed PCL/TCP scaffold promoted osteogenic differentiation of MSCs and may be widely used for bone tissue engineering.

• Journal of Veterinary Science
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• 제25권3호
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• pp.36.1-36.15
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• 2024

Importance: The intravenous administration of adipose tissue-derived mesenchymal stem cells (AdMSCs) in veterinary medicine is an attractive treatment option. On the other hand, it can result in severe complications, including pulmonary thromboembolism (PTE). Objective: The present study assessed the occurrence of PTE after the intravenous infusion of canine AdMSCs (cAdMSCs) into experimental animals. Methods: Five-week-old male BALB/c hairless mice were categorized into groups labeled A to G. In the control group (A), fluorescently stained 2×10⁶ cAdMSCs were diluted in 200 µL of suspension and injected into the tail vein as a single bolus. The remaining groups included the following: group B with 5×10⁶ cells, group C with 3×10⁶ cells, group D with 1×10⁶ cells, group E with 1×10⁶ cells injected twice with a one-day interval, group F with 2×10⁶ cells in 100 µL of suspension, and group G with 2×10⁶ cells in 300 µL of suspension. Results: Group D achieved a 100% survival rate, while none of the subjects in groups B and C survived (p = 0.002). Blood tests revealed a tendency for the D-dimer levels to increase as the cell dose increased (p = 0.006). The platelet count was higher in the low cell concentration groups and lower in the high cell concentration groups (p = 0.028). A histological examination revealed PTE in most deceased subjects (96.30%). Conclusions and Relevance: PTE was verified, and various variables were identified as potential contributing factors, including the cell dose, injection frequency, and suspension volume.

• International Journal of Stem Cells
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• 제16권4호
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• pp.425-437
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• 2023

Obesity, which continues to increase worldwide, was shown to irreversibly impair the differentiation potential and angiogenic properties of adipose tissue mesenchymal stromal cells (ADSCs). Because these cells are intended for regenerative medicine, especially for the treatment of inflammatory conditions, and the effects of obesity on the immunomodulatory properties of ADSCs are not yet clear, here we investigated how ADSCs isolated from former obese subjects (Ex-Ob) would influence macrophage differentiation and polarization, since these cells are the main instructors of inflammatory responses. Analysis of the subcutaneous adipose tissue (SAT) of overweight (OW) and Ex-Ob subjects showed the maintenance of approximately twice as many macrophages in Ex-Ob SAT, contained within the CD68⁺/FXIII-A^- inflammatory pool. Despite it, in vitro, coculture experiments revealed that Ex-Ob ADSCs instructed monocyte differentiation into a M2-like profile, and under inflammatory conditions induced by LPS treatment, inhibited HLA-DR upregulation by resting M0 macrophages, originated a similar percentage of TNF-α⁺ cells, and inhibited IL-10 secretion, similar to OW-ADSCs and BMSCs, which were used for comparison, as these are the main alternative cell types available for therapeutic purposes. Our results showed that Ex-Ob ADSCs mirrored OW-ADSCs in macrophage education, favoring the M2 immunophenotype and a mixed (M1/M2) secretory response. These results have translational potential, since they provide evidence that ADSCs from both Ex-Ob and OW subjects can be used in regenerative medicine in eligible therapies. Further in vivo studies will be fundamental to validate these observations.

• Journal of Korean Neurosurgical Society
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• 제58권2호
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• pp.101-106
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• 2015

Objective : The aim of this study was to explore the immunity in rats transplanted with adipose-derived mesenchymal stem cells (ADSCs) and acellular nerve (ACN) for repairing sciatic nerve defects. Methods : ADSCs were isolated from the adipose tissues of Wistar rats. Sprague-Dawley rats were used to establish a sciatic nerve defect model and then divided into four groups, according to the following methods : Group A, allogenic nerve graft; Group B, allograft with ACN; Group C, allograft ADSCs+ACN, and Group D, nerve autograft. Results : At the day before transplantation and 3, 7, 14, and 28 days after transplantation, orbital venous blood of the Sprague-Dawley rats in each group was collected to detect the proportion of $CD3^+$, $CD4^+$, and $CD8^+$ subsets using flow cytometry and to determine the serum concentration of interleukin-2 (IL-2), tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$) and $interferon-{\gamma}$ ($IFN-{\gamma}$) using enzyme-linked immunosorbent assay (ELISA). At each postoperative time point, the proportion of $CD3^+$, $CD4^+$, and $CD8^+$ subsets and the serum concentration of IL-2, $TNF-{\alpha}$, and $IFN-{\gamma}$ in group C were all near to those in group B and group D, in which no statistically significant difference was observed. As compared with group A, the proportion of $CD3^+$, $CD4^+$, and $CD8^+$ subsets and the serum concentration of IL-2, $TNF-{\alpha}$, and $IFN-{\gamma}$ were significantly reduced in group C (p<0.05). Conclusion : The artificial nerve established with ADSCs and ACN has no obvious allograft rejection for repairing rat nerve defects.

• Journal of Veterinary Science
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• 제22권2호
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• pp.16.1-16.13
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• 2021

Background: Preconditioning with inflammatory stimuli is used to improve the secretion of anti-inflammatory agents in stem cells from variant species such as mouse, human, and dog. However, there are only few studies on feline stem cells. Objectives: This study aimed to evaluate the immune regulatory capacity of feline adipose tissue-derived (fAT) mesenchymal stem cells (MSCs) pretreated with interferon-gamma (IFN-γ). Methods: To assess the interaction of lymphocytes and macrophages with IFN-γ-pretreated fAT-MSCs, mouse splenocytes and RAW 264.7 cells were cultured with the conditioned media from IFN-γ-pretreated MSCs. Results: Pretreatment with IFN-γ increased the gene expression levels of cyclooxygenase-2, indoleamine 2,3-dioxygenase, hepatocyte growth factor, and transforming growth factor-beta 1 in the MSCs. The conditioned media from IFN-γ-pretreated MSCs increased the expression levels of M2 macrophage markers and regulatory T-cell markers compared to those in the conditioned media from naive MSCs. Further, prostaglandin E₂ (PGE₂) inhibitor NS-398 attenuated the immunoregulatory potential of MSCs, suggesting that the increased PGE₂ levels induced by IFN-γ stimulation is a crucial factor in the immune regulatory capacity of MSCs pretreated with IFN-γ. Conclusions: IFN-γ pretreatment improves the immune regulatory profile of fAT-MSCs mainly via the secretion of PGE₂, which induces macrophage polarization and increases regulatory T-cell numbers.

• Tuberculosis and Respiratory Diseases
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• 제77권3호
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• pp.116-123
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• 2014

Background: Mesenchymal stem cells (MSCs) obtained from bone marrow or adipose tissue can successfully repair emphysematous animal lungs, which is a characteristic of chronic obstructive pulmonary disease. Here, we describe the cellular distribution of MSCs that were intravenously injected into mice with elastase-induced emphysema. The distributions were also compared to the distributions in control mice without emphysema. Methods: We used fluorescence optical imaging with quantum dots (QDs) to track intravenously injected MSCs. In addition, we used a human Alu sequence-based real-time polymerase chain reaction method to assess the lungs, liver, kidney, and spleen in mice with elastase-induced emphysema and control mice at 1, 4, 24, 72, and 168 hours after MSCs injection. Results: The injected MSCs were detected with QD fluorescence at 1- and 4-hour postinjection, and the human Alu sequence was detected at 1-, 4- and 24-hour postinjection in control mice (lungs only). Injected MSCs remained more in mice with elastase-induced emphysema at 1, 4, and 24 hours after MSCs injection than the control lungs without emphysema. Conclusion: In conclusion, our results show that injected MSCs were observed at 1 and 4 hours post injection and more MSCs remain in lungs with emphysema.

• Journal of Korean Neurosurgical Society
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• 제60권4호
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• pp.404-416
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• 2017

Objective : Functional and neural tissue recovery has been reported in many animal studies conducted with stem cells. However, the combined effect of cytokines and stem cells has not yet been adequately researched. Here, we analyzed the additive effects of granulocyte colony-stimulating factor (GCSF) on adipose-derived stem cells (ADSCs) infusion in the treatment of acute spinal cord injury (SCI) in rats. Methods : Four days after intrathecal infusion tubes implantation in Sprague-Dawley rats, SCI was induced with an infinite horizon impactor. In the Sham group (n=5), phosphate-buffered saline was injected 3, 7, and 14 days after SCI. GCSF, ADSCs, and ADSCs with GCSF were injected at the same time in the GCSF (n=8), ADSC (n=8), and ADSC+GCSF groups (n=7), respectively. Results : The ADSC and ADSC+GCSF groups, but not the GCSF group, showed significantly higher Basso-Beattie-Bresnahan scores than the Sham group during 8 weeks (p<0.01), but no significant difference between the ADSC and ADSC+GCSF groups. In the ladder rung test, all four groups were significantly different from each other, with the ADSC+GCSF group showing the best improvement (p<0.01). On immunofluorescent staining (GAP43, MAP2), western blotting (GAP43), and reverse transcription polymerase chain reaction (GAP43, nerve growth factor), the ADSC and ADSC+GCSF groups showed higher levels than the Sham and GCSF groups. Conclusion : Our analyses suggest that the combination of GCSF and ADSCs infusions in acute SCI in the rat does not have a significant additive effect. Hence, when combination agents for SCI stem cell therapy are considered, molecules other than GCSF, or modifications to the methodology, should be investigated.

• Tissue Engineering and Regenerative Medicine
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• 제15권6호
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• pp.771-779
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• 2018

BACKGROUND: Mesenchymal stromal cells (MSCs) are multipotent stem cells that can differentiate into several cell types. In addition, many studies have shown that MSCs modulate the immune response. However, little information is currently available regarding the maintenance of immunomodulatory characteristics of MSCs through passages. Therefore, we investigated and compared cytokine and gene expression levels from adipose (AD) and bone marrow (BM)-derived MSCs relevant to immune modulation from early to late passages. METHODS: MSC immunophenotype, growth characteristics, cytokine expressions, and gene expressions were analyzed. RESULTS: AD-MSCs and BM-MSCs had similar cell morphologies and surface marker expressions from passage 4 to passage 10. Cytokines secreted by AD-MSCs and BM-MSCs were similar from early to late passages. AD-MSCs and BM-MSCs showed similar immunomodulatory properties in terms of cytokine secretion levels. However, the gene expressions of tumor necrosis factor-stimulated gene (TSG)-6 and human leukocyte antigen (HLA)-G were decreased and gene expressions of galectin-1 and -3 were increased in both AD- and BM-MSCs with repeated passages. CONCLUSION: Our study showed that the immunophenotype and expression of immunomodulation-related cytokines of AD-MSCs and BM-MSCs immunomodulation through the passages were not significantly different, even though the gene expressions of both MSCs were different.

• 한국임상수의학회지
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• 제28권1호
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• pp.122-127
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• 2011

후지마비와 심부통각의 소실을 보이는 5 년령의 수컷 Cocker Spaniel이 내원하였다. 신체검사, 신경검사, 방사선검사와 컴퓨터 단층촬영결과 심부통각의 소실을 동반한 요추 2번과 3번 사이의 디스크 탈출증으로 진단하였다. 편측 추궁 절제술을 시행하여 감압술과 탈출된 디스크 물질을 제거하였으며 $1{\times}10^6$ 개의 동종 지방유래 줄기세포를 $50{\mu}l$ 생리식염수에 희석하여 손상된 척수에 직접 주입하였다. 수술 10주 후, 양쪽 후지 모두에서 심부통각과 운동기능이 완전히 회복되었으며 세포이식과 관련된 부작용은 현재까지 발견되지 않았다. 본 증례를 통해 심각한 정도의 디스크질환을 가진 환축에서 감압술과 병행된 줄기세포의 이식방법은 좀더 나은 예후를 기대하기 위한 치료방법들 중 하나로 고려해볼 수 있을 것으로 생각된다.

• Journal of Veterinary Science
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• 제23권4호
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• pp.61.1-61.10
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• 2022

Background: Although there are growing demands for stem cell-based therapy for companion animals in various diseases, a few clinical trials have been reported. Moreover, most of them are the results from only one or a few times of stem cell injection. Objectives: The aim of this study is to describe a long-term treatment with allogeneic adipose-derived stem cells (ASCs) in a dog with rheumatoid arthritis (RA), which is a rare canine disease. Methods: The dog with RA received intravascular injection of allogeneic ASCs derived from two healthy donors once a month for 11 months. To assess therapeutic effects of ASCs, orthopedic examination and clinical evaluation was performed. Cytokines of tumor necrosis factor-α and interleukin-6 in the plasma were measured using ELISA analysis. Results: Despite this repeated and long-term administration of allogeneic ASCs, there were no side effects such as immunorejection responses or cell toxicity. The orthopedic examination score for the dog decreased after ASCs treatment, and the clinical condition of the dog and owner's satisfaction were very good Conclusions: Although ASCs has been suggested as one of the options for RA treatment because of its anti-inflammatory and immunosuppressive functions, it has never been used to treat RA in dogs. The present report describes a case of canine RA treated with allogeneic ASCs for long-term in which the dog showed clinical improvement without adverse effects.