• Clinical and Experimental Reproductive Medicine
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• 제36권4호
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• pp.311-319
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• 2009

목 적: 인간 배아줄기세포 (human embryonic stem cells; hESCs)는 미분화 상태로 무한 증식할 수 있는 자가 증식(self-renewal) 능력과 인체의 모든 세포로 분화할 수 있는 전분화능 (pluripotency)의 특징을 가진 세포로, 손상된 세포를 건강한 세포로 대체하고자 하는 세포치료 (cell therapy) 연구에 활용하기 위한 세포 공급원 (cell source)으로 제시되고 있다. 그러나 인간 배아줄기세포는 확립된 초기에 세포를 안정적으로 배양하고 유지하는 과정이 쉽지 않으며, 특히 동결보존되어 있던 세포를 해동한 후 배양할 때 자연 발생적 분화가 높기 때문에 세포주의 유지에 많은 어려움이 따른다. 본 연구에서는 동결보존되어 있던 초기 계대의 인간 배아줄기세포를 해동하여 다시 배양할 때 자연 발생적 분화 부분을 기계적 분리 방법으로 제거하여 미분화 상태의 세포를 보다 빠르게 확보하기 위한 효율적인 방법에 대해 알아보고자 하였다. 연구방법: 인간 배아줄기세포를 계대 배양한지 4일이 되는 날, 50% 이상의 자연 발생적 분화가 나타난 세포군에서 분화된 부분만을 절개용 유리 피펫 (drawn-out dissecting pasture pipette)을 사용하여 기계적인 방법으로 제거하였다. 이후 지속적으로 배양액을 교환해 주며 세포군의 제거된 부분을 7일째 되는 날까지 관찰하였다. 결 과: 기계적 분리 방법을 사용하여 인간 배아줄기세포의 자연 발생적인 분화 부분을 제거한 빈 공간에 미분화상태의 인간 배아줄기세포가 분열하여 채워지는 것을 관찰하였다. 또한, 이 실험 방법을 연속 두 번 적용하여 배양했을 때 미분화 세포로 회복되는 세포군의 비율이 조금 더 높아지는 것을 확인할 수 있었다. 결 론: 동결되어 있던 초기 계대 인간 배아줄기세포의 해동 후, 자연 발생적 분화에 의해 미분화 상태를 유지하는 세포의 수가 적어 계대를 유지 하기가 어려울 때 이와 같은 기계적 분리 방법을 사용하여 자연 발생적 분화 부분을 제거한 후 배양을 지속하는 것이 단기간 내에 미분화 상태를 유지하는 인간 배아줄기세포의 양적 확보를 위한 효율적인 방법이라고 사료된다.

• Archives of Plastic Surgery
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• 제35권3호
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• pp.229-234
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• 2008

Purpose: There are some reports presenting that peripheral blood contain circulating hematopoietic cells as well as, in significantly smaller quantities, mesenchymal stem cells. The purposes of this study is to isolate and characterize circulating mesenchymal progenitor cells with osteogenic potential from human peripheral blood. Methods: Human buffycoat containing mononuclear cells was harvested from peripheral blood of normal persons and isolated using a density gradient centrifugation and serially subcultured in osteogenic media for 1-4 weeks. The proliferation capability, phase-contrast microscopy, transmission electron microscopy, immunophenotype FACS analysis, Alizarin red staining and RT-PCR assays for osteogenic differentiation potential were performed. Results: The phenotype of cultured cells changed from small round or cuboidal cells at passage 1 into large spindle-shaped fibroblastic morphology cells at passage 4. Surface marker expressed CD14, but did not express CD34, CD80, CD83. Strong positive staining was observed for Alizarin reds in osteogenic medium on day 14, Using RT-PCR, the mRNA levels of bone- specific genes, such as ALP, c-bfa-1 and osteocalcin were detected. Conclusion: A new subset of peripheral blood derived progenitor cells described here has the ability to proliferate and differentiate into osteogenic cell lineages in vitro, and to be candidate for regenerative therapy.

• Journal of Korean Neurosurgical Society
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• 제29권5호
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• pp.701-705
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• 2000

Solitary plasmacytomas are rare and account for 5-10% of all plasma cell disorders. These tumors are categorized as solitary plasmacytomas of bone(osseous) or extramedullary plasmacytomas(non-osseous). About a half of solitary plasmacytomas of bone occur in the spine but rarely in the skull. We report a case of solitary plasmacytoma of the skull presented with a painless palpable left parietal calvarial mass in an otherwise asymptomatic 38- year-old man. Skull radiographs showed a large radiolucent lesion with well defined non-sclerotic margins. Computed tomograph scan demonstrated a markedly enhancing mass extending from the epidural to the subcutaneous space. The patient underwent surgery and tumor was completely excised. Pathological examination showed tumor to be a plasmacytoma synthesizing IgG. Postoperatively, the patient received radiotherapy. There was no evidence of systemic involvement on postoperative laboratory wokups. Our recommended treatment is a complete surgical excision combined with postoperative radiation therapy. The patient should be follwed carefully for more than 10 years because of either local recurrence or possible progression to multiple myeloma.

• BMB Reports
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• 제52권9호
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• pp.572-576
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• 2019

Bisphosphonates are the mainstay of therapy worldwide for osteoporosis. However, bisphosphonates also have limitations. The objective of this study was to determine the role of miR-101-3p/Rap1b signal pathway in osteoclast differentiation after treatment with bisphosphonates. Our results revealed that miR-101-3p was an important regulator in bisphosphonates treated-osteoclasts. When miR-101-3p was down-regulated in bone marrow-derived macrophage-like cells (BMMs), the development of mature osteoclasts was promoted, and vice versa. However, alendronate decreased multinucleated cell number regardless of whether miR-101-3p was knocked down or over-expressed. TRAP activity assay confirmed the above results. Luciferase assay indicated that miR-101-3p was a negative regulator of Rap1b. Western blot analysis revealed that protein expression level of Rap1b in BMMs transfected with OV-miR-101-3p was lower than that in BMMs transfected with an empty vector. Rap1b overexpression increased TRAP-positive multinucleated cells, while Rap1b inhibition decreased the cell numbers. In vivo data showed that miR-101-3p inhibited osteoclast differentiation in ovariectomized mice while overexpressed of Rap1b blocked the differentiation. Taken together, our data demonstrate that miR-101-3p/Rap1b signal pathway plays a key role in osteoclast differentiation after treatment with bisphosphonates.

• Natural Product Sciences
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• 제29권1호
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• pp.42-49
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• 2023

This study investigated the effect of ethanol extracts of horsetail, alfalfa, ortie, chêne and aleppo oak on blood coagulation in vitro. Extraction was performed by the maceration method. Extracts were mixed with platelet and plasma, then prothrombin time (PT), activated partial thromboplastin time (APTT) and platelet aggregation tests were conducted. Alfalfa extract had a dose-dependent effect on the PT. Ortie, and horsetail, reduced the PT significantly compared to control group. Alfalfa, horsetail, and ortie reduced the APTT, but their effect was insignificant compared to the control group. The pooled extract showed the highest effect compared to the single extracts in a dose-dependent manner. Horsetail and alfalfa induced platelet aggregation in response to arachidonic acid but not in response to collagen. In the case of ortie, no aggregation occurred regarding the arachidonic acid, and incomplete was observed in response to collagen. Interestingly, blood clotting occurred immediately after adding the chêne, aleppo oak and the pooled extract, and therefore platelet poor plasma (PPP) and platelet rich plasma (PRP) became jelly. Generally, chêne and aleppo oak, as well as pooled extract, were more effective in inducing both primary and secondary coagulation pathways via shortening the PT and APTT, and induction of platelet aggregation.

• The Korean Journal of Physiology and Pharmacology
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• 제16권3호
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• pp.167-174
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• 2012

Natural killer (NK) cells provide one of the initial barriers of cellular host defense against pathogens, in particular intracellular pathogens. Because bone marrow-derived hematopoietic stem cells (HSCs), lymphoid protenitors, can give rise to NK cells, NK ontogeny has been considered to be exclusively lymphoid. Here, we show that porcine c-$kit^+$ bone marrow cells (c-$kit^+$ BM cells) develop into NK cells in vitro in the presence of various cytokines [interleukin (IL)-2, IL-7, IL-15, IL-21, stem cell factor (SCF), and fms-like tyrosine kinase-3 ligand (FLT3L)]. Adding hydrocortisone (HDC) and stromal cells greatly increases the frequency of c-$kit^+$ BM cells that give rise to $CD2^+CD8^+$ NK cells. Also, intracellular levels of perforin, granzyme B, and NKG2D were determined by RT-PCR and western blotting analysis. It was found that of perforin, granzyme B, and NKG2D levels significantly were increased in cytokine-stimulated c-$kit^+$ BM cells than those of controls. And, we compared the ability of the cytotoxicity of $CD2^+CD8^+$ NK cells differentiated by cytokines from c-$kit^+$ BM cells against K562 target cells for 28 days. Cytokines-induced NK cells as effector cells were incubated with K562 cells as target in a ratio of 100 : 1 for 4 h once a week. In results, $CD2^+CD8^+$ NK cells induced by cytokines and stromal cells showed a significantly increased cytotoxicity 21 days later. Whereas, our results indicated that c-$kit^+$ BM cells not pretreated with cytokines have lower levels of cytotoxicity. Taken together, this study suggests that cytokines-induced NK cells from porcine c-$kit^+$ BM cells may be used as adoptive transfer therapy if the known obstacles to xenografting (e.g. immune and non-immune problems) were overcome in the future.

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2002년도 국제심포지엄
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• pp.95-95
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• 2002

Human embryonic stem (hES) cells can be induced to differentiate into tyrosine hydroxylase expressing (TH+) cells that may serve as an alternative for cell replacement therapy for Parkinson's disease (PD). To examine in vitro differentiation of hES (MB03, registered in NIH) cells into TH+ cells, hES cells were induced to differentiate according to the 4-/4+ protocol using retinoic acid (RA), ascorbic acid (AA), and/or lithium chloride (LiCl) followed by culture in N2 medium for 14 days, during which time the differentiation occurs. Immunocytochemical stainings of the cells revealed that approximately 21.1% of cells treated with RA plus AA expressed TH protein that is higher than the ratio of TH+ cells seen in any other treatment groups (RA, RA+LiCl or RA+AA+LiCl). In order to see the differentiation pattern in vivo and the ability of in vitro differentiation-induced cells in easing symptomatic motor function of PD animal model, cells (2 $\times$ 10$^{5}$ cells/2${mu}ell$) undergone 4-/4+ protocol using RA plus AA without any further treatment were transplanted into unilateral striatum of MPTP-lesioned PD animal model (C57BL/6). Following the surgery, motor behavior of the animals was examined by measuring the retention time on an accelerating rotar-rod far next 10 weeks. No significant differences in retention time of the animals were noticed until 2 weeks post-graft; however, it increased markedly at 6 weeks and 10 weeks time point after the surgery. Immunohistochemical studies confirmed that a reasonable number of TH+ cells were found at the graft site as well as other remote sites, showing the migrating nature of embryonic stem cells. These results suggest that in viかo differentiated hES cells relieve symptomatic motor behavior of PD animal model and should be considered as a promising alternative for the treatment of PD.

• Asian Pacific Journal of Cancer Prevention
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• 제14권8호
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• pp.4583-4589
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• 2013

Objective: This study aimed to investigate the effect of multimodality treatment of advanced paediatric hepatoblastoma (HB) and the factors affecting prognosis. Methods: A total of 35 children underwent multimodality treatments consisting of chemotherapy, surgery, interventional therapy, and autologous peripheral blood stem cell transplantation. The patients were followed up every month. Results: Serum AFP levels in 33 out of 35 patients in this study were significantly increased (P = 0.0002). According to the statistical scatter plot, the values of serum AFP on the 25th, 50th, and 75th percentages were 1,210, 1,210 and 28,318 ng/dl, respectively. Of the 35 cases, 21 were stage IV. 18 cases were treated with systemic chemotherapy before surgery, and 3 cases with locally interventional chemotherapy before surgery. Statistical analysis showed that the preferred interventional treatment affected prognosis, and that there was a statistically significant difference (P = 0.024). Some 33 patients completed the follow-up, of which 17 were in complete remission (CR), 5 were in partial remission (PR), 1 became disease progressive (DP), and 10 died. The remission and overall survival rates were 66.7% (22/33) and 69.7% (23/33), respectively. Patients with the mixed HB phenotypes had worse prognoses than the epithelial phenotype (P < 0.001), and patients in stage IV had a lower survival rate than those in stage III (P < 0.001). Conclusion: Multimodality treatment can effectively improve remission rate and prolong the survival of children with advanced HB. In addition, alpha-fetoprotein (AFP), a tumor marker of liver malignant tumors, HB pathological classification, and staging are highly useful in predicting prognosis.

• 한국보건간호학회지
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• 제14권2호
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• pp.191-202
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• 2000

This study was conducted for 39 patients who are recipients of allogeneic hemopoietic stem cell transplantation at BMT ward of St. Mary's hospital affiliated to Catholic University of Korea from April to September 1999. The subjects were devided into two groups; those who received both TEl and chemo therapy as conditioning regimen (TEl group). and those who used chemo agents as singular conditioning regimen (chemo group). The oral intake status of the two groups were compared through physical assessment and blood chemistry exam of the subjects, and factors influencing their nutritional change and oral intake were explored in each stage of the transplantation (six stages: admission, conditional stage, date of transplantation, one week after transplantation, two weeks after transplantation, and three weeks after transplantation). The prior aim of the study was to provide baseline data to minimize delayed treatment from nutritional deficiency of the subjects. The results were as follows: 1. TBI group was significantly decreased of oral calorie intake in two weeks after transplantation compared to admission and conditioning stage while that of chemo group was significantly decreased on the date of transplantation. 2. TBI group was significantly decreased of protein intake in two weeks after transplantation compared to admission and conditioning stage. In chemo group, protein intake was significantly decreased on the date of transplantation compared to admission. It was remarkable that TBI group showed lesser protein intake than chemo group. 3. Both group were significantly decreased of BMI in one week and three weeks after transplantation compared to admission. TBI group showed significantly higher BMI than chemo group. 4. Both group were significantly decreased of Triceps Skinfold Thickness (TST)on the date of transplantation compared to admission stage. 5. TBI group was significantly decreased of mid-arm muscle circumference (MAMC) in two weeks after transplantation compared to admission, conditioning, date of transplantation. 6. TBI group was significantly decreased of albumin level in two weeks after transplantation compared admission stage. In chemo group, it was significantly decreased on the date of transplantation compared to admission, three weeks after the transplantation. 7. TBI group was significantly decreased of transferrin level in two weeks after transplantation compared admission, conditioning, date of transplantation and one week after transplantation. In chemo group, it was decreased of transferrin level in 3 weeks after transplantation. 8. Oral intake of TEl group was impacted by vomiting before transplantation and gingivitis after transplantation. In chemo group, it was impacted by vomiting before transplantation and by two factors, gingivitis and nausea, after transplantation. The results showed oral calorie intake was not different between the two groups while protein intake was significantly lower in TBI group than chemo group. Oral intake was significantly impacted by vomiting before transplantation in both groups, but affected by oral gingivitis in TBI group and gingivitis and nausea in chemo group after transplantation. This findings present that standardized strategies to manage nutrition and gingivitis more effectively are desperately needed to enhance oral intake and protein intake of patients who receive TBI as conditioning regimen.

• 의료법학
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• 제22권4호
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• pp.159-184
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• 2021

첨단재생바이오법의 제정 의의는 본질적으로 '이식'이라는 의료행위인 재생의료를 약사법 규제에서 벗어나 의료기술적 접근으로 환자치료기회를 확대하는 데 있다. 그러나, 법이 시행된 지 1년여가 지난 상황에서 식약처가 승인하는 고위험연구는 1건도 승인되지 않고 있는 등 임상연구가 활성화되지 않고 있다. 그 이유는 환자치료기회 확대를 위한 법률 취지에도 불구하고 법률적 근거가 미흡함에도 임상연구 승인을 위한 자료요건을 의약품 개발과 연결하여 정하고 있어 많은 연구자들이 자료요건을 맞추기가 어려운 실정이다. 법 제정 이전 약사법 체계 내의 세포치료제 임상연구를 위한 제출자료는 품질 및 비임상시험 자료가 상당히 면제되었지만, 첨단재생바이오법이 시행되면서 임상연구 계획 승인신청 시 품질 및 비임상시험자료를 의약품에 준해 요구하고 있다. 이를 바로잡으려면 치료기회 확대를 위한 첨단재생의료 임상연구의 정체성을 고려할 때 의약품 개발과 연결하는 데는 제한점이 있음을 인식하고 첨단재생의료 임상연구의 정체성을 지키고, 한편으로 품목허가 시 임상연구 결과를 활용할 수 있도록 하면서 활용요건을 구체화하여 시장의 힘으로, 임상연구자의 자발적인 동기로 임상연구 승인을 위한 기본요건보다 필요한 자료를 스스로 준비할 수 있도록 해야 한다.