Aim The aim of this study was to confirm the multilineage differentiation ability of dental pulp stem cells （DPSCs） from green fluorescent protein （GFP） transgenic mice. The expression of GFP in DPSCs was also obs...Aim The aim of this study was to confirm the multilineage differentiation ability of dental pulp stem cells （DPSCs） from green fluorescent protein （GFP） transgenic mice. The expression of GFP in DPSCs was also observed during differentiation. Methodology DPSCs were harvested from the dental pulp tissue of transgenic nude mice, and then transferred to osteogenic, adipogenic, and chondrogenic media. The morphological characterization of induced cells was observed by microscopy and histological staining. The expression of marker genes was measured by RT-PCR. Results The endogenous GFP and multilineage potential of transgenic DPSCs had no influence on each other. Moreover, the results of fluorescence microscopic imaging suggest that there was no significant decline of GFP expression during DPSCs differentiation. Conclusion As the population of GFP labeled DPSCs can be easily identified, this will be a promising method for tracking DPSCs in vivo.展开更多
AIM:To characterize single-cell-derived mouse clonal mesenchymal stem cells (mcMSCs) established with bone marrow samples from three different mouse strains. METHODS:We established mcMSC lines using subfractionation ... AIM:To characterize single-cell-derived mouse clonal mesenchymal stem cells (mcMSCs) established with bone marrow samples from three different mouse strains. METHODS:We established mcMSC lines using subfractionation culturing method from bone marrow samples obtained from long bones.These lines were characterized by measuring cell growth, cell surface epitopes, differentiation potential, lineage-specific gene expression and T-cell suppression capability. Nonclonal MSCs isolated by the conventional gradient centrifugation method were used as controls. RESULTS:All mcMSC lines showed typical nonclonal MSC-like spindle shape morphology. Lines differed inoptimal growth density requirement.Cell surface epitope prof iles of these mcMSC lines were similar to those of nonclonal MSCs. However, some lines exhibited different expression levels in a few epitopes, such as CD44 and CD105. Differentiation assays showed that 90% of the mcMSC lines were capable of differentiating into adipogenic and/or chondrogenic lineages, but only 20% showed osteogenic lineage differentiation. T-cell suppression analysis showed that 75% of the lines exhibited T-cell suppression capability. CONCLUSION:mcMSC lines have similar cell morphology and cell growth rate but exhibit variations in their cell surface epitopes, differentiation potential, lineage-specifi c gene expression and T-cell suppression capability.展开更多
Although widely applied in treating hematopoietic malignancies,transplantation of hematopoietic stem/progenitor cells(HSPCs)is impeded by HSPC shortage.Whether circulating HSPCs(cHSPCs)in steady-state blood could be u...Although widely applied in treating hematopoietic malignancies,transplantation of hematopoietic stem/progenitor cells(HSPCs)is impeded by HSPC shortage.Whether circulating HSPCs(cHSPCs)in steady-state blood could be used as an alternative source remains largely elusive.Here we develop a three-dimensional culture system(3DCS)including arginine,glycine,aspartate,and a series of factors.Fourteen-day culture of peripheral blood mononuclear cells(PBMNCs)in 3DCS led to 125-and 70-fold increase of the frequency and number of CD34+cells.Further,3DCS-expanded cHSPCs exhibited the similar reconstitution rate com-pared to CD34+HSPCs in bone marrow.Mechanistically,3DCS fabricated an immunomodulatory niche,secreting cytokines as TNF to support cHSPC survival and proliferation.Finally,3DCS could also promote the expansion of cHSPCs in patients who failed in HSPC mobilization.Our 3DCS successfully expands rare cHSPCs,providing an alternative source for the HSPC therapy,particularly for the patients/donors who have failed in HSPC mobilization.展开更多
目的:探讨单细胞克隆肝癌干细胞(liver cancer stem cell,LCSCs)向间充质样细胞分化的潜能。方法:通过有限稀释法获得单个细胞来源的LCSC克隆,采用RT-PCR法鉴定干细胞标志物;将该单细胞克隆分别用成骨、软骨和脂肪诱导分化培养基培养3周...目的:探讨单细胞克隆肝癌干细胞(liver cancer stem cell,LCSCs)向间充质样细胞分化的潜能。方法:通过有限稀释法获得单个细胞来源的LCSC克隆,采用RT-PCR法鉴定干细胞标志物;将该单细胞克隆分别用成骨、软骨和脂肪诱导分化培养基培养3周后,采用Real-time PCR及特殊染色技术比较诱导前后LCSC表达成骨、软骨及脂肪细胞特异标志物的差异。结果:单细胞克隆的LCSC表达多种干细胞标志物、干细胞因子(stem cell factor,SCF)、干细胞因子受体、C-kit、巢蛋白(nestin)、CD34、三磷酸腺苷结合转运蛋白G超家族成员2(ATP-binding cassette sub-family G member 2,ABCG2)、CD133。分化诱导培养3周后,成骨方向诱导的细胞茜素红染色呈现橘红色钙结节形成,软骨方向诱导的细胞阿尔新蓝染色显示蓝色蛋白多糖沉积,脂肪方向诱导的细胞油红O染色显示大量脂滴形成。Real-time PCR结果显示,诱导后成骨细胞特异标志物骨钙素和Ⅰ型胶原、软骨细胞特异标志物蛋白聚糖和Ⅱ型胶原、脂肪细胞特异标志物脂联素和过氧化物酶体增殖物激活受体γ的mRNA表达均较对照组显著上调(Ⅰ型及Ⅱ型胶原间为P<0.05,其他指标间均为P<0.01)。结论:LCSC具有可塑性,在特定的微环境下具有向间充质样细胞分化的潜能。展开更多
基金funded by The Peabody Foundation Inc.,the Anthony and Constance Franchi Fund for Pediatric Orthopaedics at the Mass General Hospital for Children, and the National Natural Science Foundation of China (30801304)Foundation for the Author of National Excellent Doctoral Dissertation of PR China (FANEDD 200977)
文摘Aim The aim of this study was to confirm the multilineage differentiation ability of dental pulp stem cells （DPSCs） from green fluorescent protein （GFP） transgenic mice. The expression of GFP in DPSCs was also observed during differentiation. Methodology DPSCs were harvested from the dental pulp tissue of transgenic nude mice, and then transferred to osteogenic, adipogenic, and chondrogenic media. The morphological characterization of induced cells was observed by microscopy and histological staining. The expression of marker genes was measured by RT-PCR. Results The endogenous GFP and multilineage potential of transgenic DPSCs had no influence on each other. Moreover, the results of fluorescence microscopic imaging suggest that there was no significant decline of GFP expression during DPSCs differentiation. Conclusion As the population of GFP labeled DPSCs can be easily identified, this will be a promising method for tracking DPSCs in vivo.
基金Supported by A Grant from the Korea Health 21 R&D ProjectMinistry of Health and Welfare,Republic of Korea(A092142)+1 种基金a research grant from Homeo Therapy Co.Ltd.(39856-01)by the Brain Korea 21 Project in 2010
文摘 AIM:To characterize single-cell-derived mouse clonal mesenchymal stem cells (mcMSCs) established with bone marrow samples from three different mouse strains. METHODS:We established mcMSC lines using subfractionation culturing method from bone marrow samples obtained from long bones.These lines were characterized by measuring cell growth, cell surface epitopes, differentiation potential, lineage-specific gene expression and T-cell suppression capability. Nonclonal MSCs isolated by the conventional gradient centrifugation method were used as controls. RESULTS:All mcMSC lines showed typical nonclonal MSC-like spindle shape morphology. Lines differed inoptimal growth density requirement.Cell surface epitope prof iles of these mcMSC lines were similar to those of nonclonal MSCs. However, some lines exhibited different expression levels in a few epitopes, such as CD44 and CD105. Differentiation assays showed that 90% of the mcMSC lines were capable of differentiating into adipogenic and/or chondrogenic lineages, but only 20% showed osteogenic lineage differentiation. T-cell suppression analysis showed that 75% of the lines exhibited T-cell suppression capability. CONCLUSION:mcMSC lines have similar cell morphology and cell growth rate but exhibit variations in their cell surface epitopes, differentiation potential, lineage-specifi c gene expression and T-cell suppression capability.
基金Data and materials availability:Processed and raw data can be downloaded from NCBI GEO(#GSE122682,and#GSE153421).
文摘Although widely applied in treating hematopoietic malignancies,transplantation of hematopoietic stem/progenitor cells(HSPCs)is impeded by HSPC shortage.Whether circulating HSPCs(cHSPCs)in steady-state blood could be used as an alternative source remains largely elusive.Here we develop a three-dimensional culture system(3DCS)including arginine,glycine,aspartate,and a series of factors.Fourteen-day culture of peripheral blood mononuclear cells(PBMNCs)in 3DCS led to 125-and 70-fold increase of the frequency and number of CD34+cells.Further,3DCS-expanded cHSPCs exhibited the similar reconstitution rate com-pared to CD34+HSPCs in bone marrow.Mechanistically,3DCS fabricated an immunomodulatory niche,secreting cytokines as TNF to support cHSPC survival and proliferation.Finally,3DCS could also promote the expansion of cHSPCs in patients who failed in HSPC mobilization.Our 3DCS successfully expands rare cHSPCs,providing an alternative source for the HSPC therapy,particularly for the patients/donors who have failed in HSPC mobilization.
文摘目的:探讨单细胞克隆肝癌干细胞(liver cancer stem cell,LCSCs)向间充质样细胞分化的潜能。方法:通过有限稀释法获得单个细胞来源的LCSC克隆,采用RT-PCR法鉴定干细胞标志物;将该单细胞克隆分别用成骨、软骨和脂肪诱导分化培养基培养3周后,采用Real-time PCR及特殊染色技术比较诱导前后LCSC表达成骨、软骨及脂肪细胞特异标志物的差异。结果:单细胞克隆的LCSC表达多种干细胞标志物、干细胞因子(stem cell factor,SCF)、干细胞因子受体、C-kit、巢蛋白(nestin)、CD34、三磷酸腺苷结合转运蛋白G超家族成员2(ATP-binding cassette sub-family G member 2,ABCG2)、CD133。分化诱导培养3周后,成骨方向诱导的细胞茜素红染色呈现橘红色钙结节形成,软骨方向诱导的细胞阿尔新蓝染色显示蓝色蛋白多糖沉积,脂肪方向诱导的细胞油红O染色显示大量脂滴形成。Real-time PCR结果显示,诱导后成骨细胞特异标志物骨钙素和Ⅰ型胶原、软骨细胞特异标志物蛋白聚糖和Ⅱ型胶原、脂肪细胞特异标志物脂联素和过氧化物酶体增殖物激活受体γ的mRNA表达均较对照组显著上调(Ⅰ型及Ⅱ型胶原间为P<0.05,其他指标间均为P<0.01)。结论:LCSC具有可塑性,在特定的微环境下具有向间充质样细胞分化的潜能。
