Duration and magnitude of extracellular signal-regulated protein kinase phosphorylation determine adipogenesis or osteogenesis in human bone marrow-derived stem cells.

PURPOSE: Imbalances between osteogenic and adipogenic differentiation leads to diseases such as osteoporosis. The aim of our study was to demonstrate the differences in extracellular signal-regulated kinase (ERK) phosphorylation during both adipogenesis and osteogenesis of human bone marrow-derived stem cells (BMSCs). MATERIALS AND METHODS: Using troglitazone, GW9662 and U0126, we investigated their role in hBMSC differentiation to adipogenic and osteogenic fates. RESULTS: ERK1/2 inhibition by U0126 suppressed proliferator-activated receptor (PPAR)γ expression and lipid accumulation, while it decreased the mRNA expression of adipogenic genes (lipoprotein lipase, PPARγ, and adipocyte protein) and osteogenic genes (type I collagen and osteopontin). ERK phosphorylation was transient and decreased during adipogenesis, whereas it occurred steadily during osteogenesis. Troglitazone, a PPARγ agonist, induced adipogenesis by inhibiting ERK phosphorylation even in an osteogenic medium, suggesting that ERK signaling needs to be shut off in order to proceed with adipose cell commitment. Cell proliferation was greatly increased in osteogenesis but was not changed during adipogenesis, indicating that ERK might play different roles in cellular proliferation and differentiation between the two committed cell types. CONCLUSION: The duration and magnitude of ERK activation might be a crucial factor for the balance between adipogenesis and osteogenesis in human bone marrow-derived stem cells.ope

The role of microRNAs involved in mesenchymal stem cell differentiation

ope

Peroxisome proliferator-activated receptor gamma/signal transducers and activators of transcription 5A pathway plays a key factor in adipogenesis of human bone marrow-derived stromal cells and 3T3-L1 preadipocytes

Adipogenesis is largely dependent on the signal transducers and activators of transcription (STAT) pathway. However, the molecular mechanism of the STAT pathway in the adipogenesis of human bone marrow-derived stromal cells (hBMSCs) remains not well understood. The purpose of this research was to characterize the transcriptional regulation involved in expression of STAT5A and STAT5B during adipogenesis in hBMSCs and 3T3-L1 cells. The expression of STAT5A and STAT5B increases with the onset of adipogenesis in hBMSCs and 3T3-L1 cells. The PPAR response elements regulatory element of STAT5A exists at a promoter region ranging from -346 to -101, and the CCAAT/enhancer-binding protein (C/EBP) regulatory element is located at -196 to -118 of the STAT5B promoter. C/EBPβ and C/EBPα bound to the STAT5B promoter region, whereas peroxisome proliferator-activated receptor γ (PPARγ) bound to STAT5A. RNA interference of STAT5A completely blocked differentiation, whereas the inhibition of STAT5B only partially blocked differentiation. We propose that C/EBPα, C/EBPβ, and PPARγ control adipogenesis by regulating STAT5B and STAT5A and that STAT5A is necessary, whereas STAT5B plays a supplementary role during adipogenesis. Further, the regulation of PPARγ-STAT5 by C/EBPβ signaling seems to be the crucial adipogenesis pathway-initiating cascade of the various adipogenic genes.ope

Differential Expression of STATs and Its Function in Osteogenesis of Mesenchymal Stem Cells

PURPOSE: To investigate the role of STAT5 during osteogenesis differentiation of human mesenchymal stem cells(hMSC). MATERIALS AND METHODS: To assess the expression pattern of STATs during osteogenic differentiation, we performed western blot analysis and RT-PCR. By RNA interference, we have performed effect of STAT5A and STAT5B in osteogenesis. As a result of Luciferase assay, the promoter activity of DLX5 was decreased by STAT5A. RESULTS: To assess the expression pattern of STATs during osteogenic differentiation, we have performed western blot and RT-PCR after 14 day differentiation period. STAT1, 2, 3 and 4 showed no change in expression level during differentiation, while STAT5A and STAT5B displayed steady increase compared to the control. When STAT5A was knock down, the level of osteogenesis was increased. The transcriptional activity of DLX5 was decreased about 70% by STAT5A. CONCLUSION: The expression of STAT5A and STAT5B increased during osteogenesis of hMSC. Also, we shown that STAT5A regulated the transcriptional activity of DLX5. These results indicate that STAT5A acts as a pivotal transcription factor in osteogenesis of hMSC.ope

The regulation of STAT5 by C/EBPa, C/EBPb, and PPARr in adipogenesis

Dept. of Medical Science/박사Adipogenesis is a complex process that includes the proliferation of precursor cells, their commitment to the adipogenic lineage, and terminal differentiation. During the differentiation of preadipocytes, the expression of adipocyte-specific genes depends on the transcriptional activation and the expression of families of transcription factors, C/EBPs and PPARs. In addition, adipogenesis is largely dependent on the STAT (Signal Transducer and Activator of Transcription) pathway. However, the molecular mechanism of the STAT pathway’s role on the adipogenesis of human bone marrow derived-stromal cells (BMSCs) is still not well known. In this study, we investigated transcriptional regulation involved in the expression of STAT5A and STAT5B during adipogenesis.We determined that the expression of STAT5A and STAT5B increases from the onset of adipogenesis of hBMSCs and 3T3-L1 cells and is maintained throughout adipogenesis. In addition, we successfully determined that STAT5A is regulated at the transcriptional level by PPAR? and RXR?, whereas STAT5B is regulated by C/EBP? and C/EBP?. The PPRE regulatory element of STAT5A exists at a promoter region ranging from nucleotides -346 to -101, and the C/EBP regulatory element was located at the -196 to -118 nucleotide region of the STAT5B promoter. We determined that C/EBP? and C/EBP? collectively bind to the STAT5B promoter region, whereas PPAR? binds to that of STAT5A using EMSA and ChIP assays. RNA interference with STAT5A resulted in a complete blockage of the differentiation, whereas inhibition of STAT5B only partially blocked the differentiation. We propose that C/EBP?, C/EBP? and PPAR? control adipogenesis by regulating STAT5B and STAT5A, respectively, and that STAT5A is absolutely necessary; STAT5B only plays a supplemental role during adipogenesis. Furthermore, PPAR?-STAT5 regulation by C/EBP? signaling seems to be crucial in the adipogensis pathway-initiating cascade of various adipogenic genes.restrictio

Analysis of the success factors of smoking cessation of new and re-registrants of smoking cessation clinics at public health centers.

Background and Purpose Smoking cessation clinics at public health centers play a key role in community smoking cessation projects, but the size of re-registrants and the success factors of smoking cessation are not well known compared to new registrants. The study aims to understand the number of new and re-registered smoking clinics nationwide from 2010 to 2020, and to find out the relationship between demographic characteristics, smoking-related characteristics, and 6-month smoking success of new and re-registered health centers nationwide from 2015 to 2020. Methods From 2010 to 2020, 4,478,300 new, re-registration, and age-specific registration trends were identified, and 2,172,038 users from 2015 to 2020, when the integrated smoking cessation service information system was established and managed, were analyzed. Frequency analysis and descriptive statistics were conducted to understand the characteristics of the study subjects, and the relationship between the success of smoking cessation of new registration and re-registration was analyzed. Logistic regression analysis was conducted by correcting nicotine dependence, first smoking age, average daily smoking amount, smoking confidence, and smoking preparation due to demographic characteristics such as gender, age, social security, and service use. Result From 2010 to 2020, the total number of registrants per year was the highest in 2015 and decreased in 2020. Over the past 11 years, new registrations have been on the decline, and re-registration has been on the rise recently. Among the 2,172,038 subjects analyzed between 2015 and 2020, 1,930,536 males, 88.9% were 1,580 and 138 newly registered and 591,900 re-registered. Compared to new registrants, re-registrants often received behavioral therapy and supplements together, the number of consultations, smoking cessation confidence, and the degree of preparation for smoking cessation were high, and the amount of smoking per day was high. The four-week smoking cessation success rate was higher, but the six-month smoking cessation success rate was 37.9%, lower than 39.3% of new registrants. In the analysis of six-month smoking cessation success factors, the odds ratio for re-registrants to succeed was 0.84 (95% CI 0.83-0.84) lower than that of new registrants. The factors for smoking cessation success of new registrants and re-registrants were the same, with men, high age, health insurance subscribers, and anti-smoking assistance provided alone (OR 1.255,95% CI 1.23-1.28; re-registrants OR 1.30,95% CI 1.26-1.34; non-smoking confidence and 7.75%.) Conclusion Amid a decrease in the number of registered smoking cessation clinics at public health centers, the number of re-registrants is increasing. Re-registrants are more prepared to quit smoking than new registrants, but they consume more resources and are difficult to succeed. Smoking cessation clinics The factors for improving the success rate of smoking cessation were the same for new and re-registrants, and the increase in the number of consultations was the most characteristic. When the number of consultations increases, it is necessary to prepare an action plan after confirming the effect on improving the success rate. Research is needed to see if smoking cessation confidence and smoking cessation preparation also have the effect of improving the success rate during intervention, and further research is needed to develop a successful intervention strategy for the increasing number of re-registrants. 배경 및 목적 보건소 금연 클리닉은 지역사회 금연사업의 핵심 중추 역할을 하고 있으나, 신규등록자보다 재등록자의 규모와 금연 성공 요인에 대해서 잘 알려지지 않았다. 이 연구에서는 2010년부터 2020년까지 전국 보건소 금연 클리닉의 신규등록 및 재등록자 수와 이들의 성공률 추이를 파악하고, 2015년부터 2020년 전국 보건소 금연 클리닉 서비스 대상자, 신규등록 및 재등록자의 인구 사회학적 특성, 흡연 관련 특성, 서비스 이용 특성과 6개월 금연 성공의 관련성을 알아보고자 한다. 연구방법 2010년부터 2020년까지 전국 258개 보건소 금연 클리닉을 방문한 4,478,300명을 대상으로 신규, 재등록과 연령별 등록현황 추이를 파악하였고, 금연서비스 통합정보시스템이 구축, 관리된 2015년부터 2020년까지의 이용자 2,172,038명을 대상으로는 금연 성공 관련 요인을 분석하였다. 연구 대상자의 제 특성을 파악하기 위해 빈도 분석과 기술 통계를 시행하였고, 신규등록 및 재등록의 금연 성공과의 관련성을 분석하였다. 금연 성공 관련 요인을 파악하기 위해 인구 사회학적 특성으로 성별, 연령, 사회보장, 서비스 이용 특성으로 서비스 제공 내용, 상담 횟수, 서비스 등록 경로, 흡연 관련 특성으로 니코틴 의존도, 첫 흡연 나이, 하루 평균 흡연량, 금연 준비 정도 중 금연 자신감, 금연 준비 정도의 영향을 보정하여 로지스틱 회귀 분석을 시행하였다. 연구결과 2010년부터 2020년까지 연도별 총등록자 수는 2015년에 최다였고 2020년에는 감소하였다. 11년간 신규등록은 감소 추세, 재등록은 최근 증가 추세를 보였다. 2015-2020년 분석 대상자 2,172,038명(남자 1,930,536명, 88.9%) 중, 신규등록자 1,580,138명, 재등록자 591,900명이었다. 재등록 자는 신규등록자보다 행동요법과 보조제를 같이 받은 경우가 많았고, 상담횟수, 금연자신감, 금연 준비 정도가 높았으며, 하루 흡연량이 많았다. 4주 금연 성공률은 더 높았으나, 6개월 금연 성공률은 37.9%로 신규등록자의 39.3%보다 낮았다. 6개월 금연 성공 요인 분석에서, 재등록 자는 신규등록자보다 성공할 오즈비가 0.84(95%CI 0.83-0.84)로 낮았다. 신규등록자와 재등록 자의 금연 성공 요인은 다음과 같으며 동일하였는데, 남자, 높은 연령, 건강보험 가입자, 금연보조제 단독 제공(신규등록자 OR 1.25,95%CI 1.23-1.28; 재등록자 OR 1.30,95%CI 1.26-1.34),금연 자신감과 금연 준비 정도가 높은 경우, 7.75회 초과 상담 횟수(신규등록자 OR 7.90,95%CI 7.84-7.96; 재등록자 OR 6.96,95%CI 6.87-7.05), 낮은 니코틴 의존도, 높은 첫 흡연 연령, 적은 하루 평균 흡연량이었다. 결론 보건소 금연 클리닉 등록자가 줄어든 가운데 재등록 자는 증가하고 있다. 재등록 자는 신규등록자보다 금연 준비도는 높으나 자원을 더 많이 소모하고, 성공하기는 어렵다. 금연 클리닉 금연 성공률 향상을 위한 요인은 신규등록자와 재등록 자에게서 동일하였으며, 상담 횟수 증가가 가장 특징적이었다. 상담 횟수 증가 시 성공률 향상에 대한 효과를 확인한 후, 실행 방안 마련이 필요하다. 금연 자신감과 금연 준비 정도 역시 개입 시 성공률 향상 효과가 있는지 연구가 필요하며, 그 외 증가하는 재등록 자에 대한 성공적 개입 전략 개발을 위한 지속적인 연구가 필요하다.open석

The Role of MAP Kinase in Adipogenesis from Human Bone Marrow-Derived Stromal Cells

Adipogenesis is a complex process that includes the proliferation of precursor cells, their commitment to the adipogenic lineage and terminal differentiation. This study examined whether or not the ERK signaling pathway regulates the expression or activity of the adipogenic transcription factors during human adipogenesis. The bone marrow-derived stromal cells (BMSCs) were differentiated into adipocytes with the 25 μM troglitazone for 14 days, and aP2 and LPL mRNA were detected. In order to determine the role of the activated ERK during adipogenesis, the BMSCs were exposed to troglitazone and U0126. After inducing differentiation, ERK activity was transiently decreased from 30 min to 6 h in the adipogenic medium and the adipogenic medium plus troglitazone. Those with U0126 markedly inhibited ERK activation. In the presence of troglitazone, the expression of the marker genes increased in a time-dependent manner, and the expression of aP2, LPL and PPARγ genes decreased in the adipogenic BMSCs treated with U0126 in RNA level. The treatment of the adipogenic BMSCs with troglitazone activated C/EBPα and PPARγ, and the U0126 treatment did not affect C/EBPα expression but inhibited PPARγ expression in protein level. In addition, U0126 blocked the change in the adipocyte phenotype. These results suggest that ERK activation is related to PPAR expression but not C/EBPα, and C/EBPα might mediate another pathway in the adipogenesis of BMSCs.restrictio

Construction of Information Database in Research Equipment

funder : 과학기술부agency : 과학기술부agency : Ministry of Science & Technology본 연구는 대학, 연구소 등이 보유하고 있는 고가 연구장비의 DB구축을 통해 연구장비의 효율적 취득․관리, 공동활용 활성화 및 장비관련예산의 효율적 사용 도모하고 미래의 지식기반 사회에 대응할 만한 성장기반을 확충하기 위해서는 연구장비의 확충에 노력해야 하나, 연구장비의 대부분이 수입된 고가 연구장비로서 이러한 수입장비의 양적인 확충에 대한 투자효율의 문제가 제기되고 있어, 기존 장비를 대외개방하여 공동활용함으로써 연구개발 투자에 대한 효율을 높이고, 연구장비의 중복구매를 방지하는데 있다.The purpose of the this project is to supply research equipment information resource to the researchers. The ultimate goal of the study is to support researchers for the development of science and technology. This database will serve end-users such as institutes, universities, industries and personal researchers via Korea Equipment On-Line

Evaluation of Efficacy and Safety of Stem Cells as a Therapeutics, Especially Focusing on Bone Marrow Derived Stromal Cells

Bone marrow derived-stromal cells (BMSCs) are pluriplotent progenitors for a variety of cell types, including osteoblast, chondrocyte, adipocyte, and so on. Stem cell research has enormous potential in the future clinical treatment of a wide range of diseases. Tracking stem cell localization, survival, differentiation, and proliferation after transplantation in living subjects is essential for understanding stem cell biology and physiology. However, we don’t have exact evaluation methods and safeguards for clinical application. In this study, we investigated tracking BMSCs differentiation, localization, toxicity, and migration in vivo. The fluorescent vector used in our studies did not affect BMSCs viability or their ability to undergo osteogenic and adipogenic differentiation in vitro. During differentiation, EGFP-BMSCs by Oil Red O and Alizarin Red S were stained. EGFP-BMSCs were transplanted into the femoral region in autologous rabbit. After one month, these cells were detectable by confocal microscopy and RTPCR. Transplanted EGFP-BMSCs were not detected another organs (spleen, kidney, liver, and muscle) in immunohistochemistry and RT-PCR. In organ-function test and cell-toxicity examination, there is no difference between the before and after EGFP-BMSCs transplantation. we observed that transplanted EGFP-BMSCs were not affected cell-toxicity and migration. This results offer some evaluation methods and safeguards for clinical application using BMSCs. In further study, it will be needed to test reproductive and developmental toxicity after transplantation and observe migration of EGFP-BMSCs after transplantation.ope

Importance of Sox2 in maintenance of cell proliferation and multipotency of mesenchymal stem cells in low-density culture.

OBJECTIVES: This study has aimed to repopulate 'primitive' cells from late-passage mesenchymal stem cells (MSCs) of poor multipotentiality and low cell proliferation rate, by simply altering plating density. MATERIALS AND METHODS: Effects of low density culture compared t high density culture on late-passage bone marrow (BM)-derived MSCs and pluripotency markers of multipotentiality were investigated. Cell proliferation, gene expression, RNA interference and differentiation potential were assayed. RESULTS AND CONCLUSIONS: We repopulated 'primitive' cells by replating late-passage MSCs at low density (17 cells/cm(2) ) regardless of donor age. Repopulated MSCs from low-density culture were smaller cells with spindle shaped morphology compared to MSCs from high-density culture. The latter had enhanced colony-forming ability, proliferation rate, and adipogenic and chondrogenic potential. Strong expression of osteogenic-related genes (Cbfa1, Dlx5, alkaline phosphatase and type Ι collagen) in late-passage MSCs was reduced by replating at low density, whereas expression of three pluripotency markers (Sox2, Nanog and Oct-4), Osterix and Msx2 reverted to levels of early-passage MSCs. Knockdown of Sox2 and Msx2 but not Nanog, using RNA interference, showed significant decrease in colony-forming ability. Specifically, knockdown of Sox2 significantly inhibited multipotentiality and cell proliferation. Our data suggest that plating density should be considered to be a critical factor for enrichment of 'primitive' cells from heterogeneous BM and that replicative senescence and multipotentiality of MSCs during in vitro expansion may be predominantly regulated through Sox2.ope