Spinal cord injury（SCI）is a devastating ailment that results in drastic life style alterations for the patients and their family members（Mc Donald and Sadowsky,2002）.Damage post injury causes necrosis,edema,hemorr...Spinal cord injury（SCI）is a devastating ailment that results in drastic life style alterations for the patients and their family members（Mc Donald and Sadowsky,2002）.Damage post injury causes necrosis,edema,hemorrhage and vasospasm.Post injury,secondary damage is caused by ischemia,展开更多
Objective:To obtain the bone marrow mesenchymal stem cells(BMSCs).complete phenotypic identification and successfully transfecl rat BMSCs by recombinant plasmid pF.GFP/Ang-1.Methods:BMSCs were isolated from bone marro...Objective:To obtain the bone marrow mesenchymal stem cells(BMSCs).complete phenotypic identification and successfully transfecl rat BMSCs by recombinant plasmid pF.GFP/Ang-1.Methods:BMSCs were isolated from bone marrow using density gradient centrifugation method and adherence screening method,and purified.Then the recombinant plasmid pEGFP/Ang-1was used to transfect BMSCs and the positive clones were obtained by the screen of C418 and observed under light microscopy inversely.Green fluorescent exhibited by protein was enhanced to measure the change time of the expression amount of Ang-1.Results:BMSCs cell lines were obtained successfully by adherence screening method and density gradient ccntrifugation.Ang-1 recombinant plasmid was transfected smoothly into rat BMSCs,which can express Ang-1 for 3 d and decreased after 7 d.Conclusions:Adherence screening method und density gradient ceiilrifugation can be effective methods lo obtain BMSCs with high purity and rapid proliferation.Besides,the expression of transfected recombinant plasmid pEGFP/Ang-1 in rat BMSCs is satisfactory.展开更多
We report here a method for the use of poly-L-lysine （PLL） to markedly improve the adenoviral transduction efficiency of primary murine osteoblasts and bone marrow stromal cells （BMSCs） in culture and in situ, whi...We report here a method for the use of poly-L-lysine （PLL） to markedly improve the adenoviral transduction efficiency of primary murine osteoblasts and bone marrow stromal cells （BMSCs） in culture and in situ, which are typically difficult to transduce. We show by fluorescence microscopy and flow cytometry that the addition of PLL to the viral-containing medium significantly increases the number of green fluorescence protein （GFP）-positive osteoblasts and BMSCs transduced with an enhanced GFP-expressing adenovirus. We also demonstrate that PLL can greatly enhance the adenoviral transduction of osteoblasts and osteocytes in situ in ex vivo tibia and calvaria, as well as in long bone fragments. In addition, we validate that PLL can improve routine adenoviral transduction studies by permitting the use of low multiplicities of infection to obtain the desired biologic effect. Ultimately, the use of PLL to facilitate adenoviral gene transfer in osteogenic cells can provide a cost-effective means of performing efficient gene transfer studies in the context of bone research.展开更多
Non-adherent bone marrow cell-derived mesenchymal stem cells from C57BL/6J mice were sepa- rated and cultured using the ＂pour-off＂ method. Non-adherent bone marrow cell-derived mesen- chymal stem ceils developed col...Non-adherent bone marrow cell-derived mesenchymal stem cells from C57BL/6J mice were sepa- rated and cultured using the ＂pour-off＂ method. Non-adherent bone marrow cell-derived mesen- chymal stem ceils developed colony-forming unit-fibroblasts, and could be expanded by supple- mentation with epidermal growth factor. Immunocytochemistry showed that the non-adherent bone marrow cell-derived mesenchymal stem cells exposed to basic fibroblast growth factor/epidermal growth factor/nerve growth factor expressed the neuron specific markers, neurofilament-200 and NeuN, in vitro. Non-adherent bone marrow cell-derived mesenchymal stem cells from 13-galactosidase transgenic mice were also transplanted into focal ischemic brain （right corpus striatum） of C57BL/6J mice. At 8 weeks, cells positive for LacZ and 13-galactosidase staining were observed in the ischemic tissues, and cells co-labeled with both 13-galactosidase and NeuN were seen by double immunohistochemical staining. These findings suggest that the non-adherent bone marrow cell-derived mesenchymal stem cells could differentiate into neuronal-like cells in vitro and in vivo.展开更多
Cerebral ischemia remains the most frequent cause of death and quality-of-life impairments due to neurological deficits, and accounts for the majority of total healthcare costs. However, treatments for cerebral ischem...Cerebral ischemia remains the most frequent cause of death and quality-of-life impairments due to neurological deficits, and accounts for the majority of total healthcare costs. However, treatments for cerebral ischemia are limited. Over the last decade, bone marrow stromal cell （BMSC） therapy has emerged as a particularly appealing option, as it is possible to help patients even when initiated days or even weeks after the ischemic insult. BMSCs are a class of multipotent, self-renewing cells that give rise to differentiated progeny when implanted into appropriate tissues. Therapeutic effects of BMSC treatment for ischemic stroke, including sensory and motor recovery, have been reported in pre-clinical studies and clinical trials. In this article, we review the recent progress in BMSC-based therapy for ischemic stroke, focusing on the route of delivery and pre-processing of BMSCs. Selecting an optimal delivery route is of particular importance. The ideal approach, as well as the least risky, for translational applications still requires further identification. Appropriate pre- processing of BMSCs or combination therapy has the benefit of achieving the maximum possible restoration. Further pre-clinical studies are required to determine the time-window for transplantation and the appropriate dosage of cells.展开更多
文摘Spinal cord injury（SCI）is a devastating ailment that results in drastic life style alterations for the patients and their family members（Mc Donald and Sadowsky,2002）.Damage post injury causes necrosis,edema,hemorrhage and vasospasm.Post injury,secondary damage is caused by ischemia,
基金Supported by National Natural Science Foundation of China(Grant No.20114020)
文摘Objective:To obtain the bone marrow mesenchymal stem cells(BMSCs).complete phenotypic identification and successfully transfecl rat BMSCs by recombinant plasmid pF.GFP/Ang-1.Methods:BMSCs were isolated from bone marrow using density gradient centrifugation method and adherence screening method,and purified.Then the recombinant plasmid pEGFP/Ang-1was used to transfect BMSCs and the positive clones were obtained by the screen of C418 and observed under light microscopy inversely.Green fluorescent exhibited by protein was enhanced to measure the change time of the expression amount of Ang-1.Results:BMSCs cell lines were obtained successfully by adherence screening method and density gradient ccntrifugation.Ang-1 recombinant plasmid was transfected smoothly into rat BMSCs,which can express Ang-1 for 3 d and decreased after 7 d.Conclusions:Adherence screening method und density gradient ceiilrifugation can be effective methods lo obtain BMSCs with high purity and rapid proliferation.Besides,the expression of transfected recombinant plasmid pEGFP/Ang-1 in rat BMSCs is satisfactory.
基金supported by grants, R01-AR063631 (JPS) and F31-AR064673 (AMB), from the National Institutes of Health/National Institute for Arthritis, Musculoskeletal and Skin Diseases
文摘We report here a method for the use of poly-L-lysine （PLL） to markedly improve the adenoviral transduction efficiency of primary murine osteoblasts and bone marrow stromal cells （BMSCs） in culture and in situ, which are typically difficult to transduce. We show by fluorescence microscopy and flow cytometry that the addition of PLL to the viral-containing medium significantly increases the number of green fluorescence protein （GFP）-positive osteoblasts and BMSCs transduced with an enhanced GFP-expressing adenovirus. We also demonstrate that PLL can greatly enhance the adenoviral transduction of osteoblasts and osteocytes in situ in ex vivo tibia and calvaria, as well as in long bone fragments. In addition, we validate that PLL can improve routine adenoviral transduction studies by permitting the use of low multiplicities of infection to obtain the desired biologic effect. Ultimately, the use of PLL to facilitate adenoviral gene transfer in osteogenic cells can provide a cost-effective means of performing efficient gene transfer studies in the context of bone research.
基金supported by the National Natural Science Foundation of China,No.30471836
文摘Non-adherent bone marrow cell-derived mesenchymal stem cells from C57BL/6J mice were sepa- rated and cultured using the ＂pour-off＂ method. Non-adherent bone marrow cell-derived mesen- chymal stem ceils developed colony-forming unit-fibroblasts, and could be expanded by supple- mentation with epidermal growth factor. Immunocytochemistry showed that the non-adherent bone marrow cell-derived mesenchymal stem cells exposed to basic fibroblast growth factor/epidermal growth factor/nerve growth factor expressed the neuron specific markers, neurofilament-200 and NeuN, in vitro. Non-adherent bone marrow cell-derived mesenchymal stem cells from 13-galactosidase transgenic mice were also transplanted into focal ischemic brain （right corpus striatum） of C57BL/6J mice. At 8 weeks, cells positive for LacZ and 13-galactosidase staining were observed in the ischemic tissues, and cells co-labeled with both 13-galactosidase and NeuN were seen by double immunohistochemical staining. These findings suggest that the non-adherent bone marrow cell-derived mesenchymal stem cells could differentiate into neuronal-like cells in vitro and in vivo.
基金supported by the Natural Science Foundation of Heilongjiang Province, China (LC201040)the Science Foundation of Education Department of Heilongjiang Province, China (1155h005)
文摘Cerebral ischemia remains the most frequent cause of death and quality-of-life impairments due to neurological deficits, and accounts for the majority of total healthcare costs. However, treatments for cerebral ischemia are limited. Over the last decade, bone marrow stromal cell （BMSC） therapy has emerged as a particularly appealing option, as it is possible to help patients even when initiated days or even weeks after the ischemic insult. BMSCs are a class of multipotent, self-renewing cells that give rise to differentiated progeny when implanted into appropriate tissues. Therapeutic effects of BMSC treatment for ischemic stroke, including sensory and motor recovery, have been reported in pre-clinical studies and clinical trials. In this article, we review the recent progress in BMSC-based therapy for ischemic stroke, focusing on the route of delivery and pre-processing of BMSCs. Selecting an optimal delivery route is of particular importance. The ideal approach, as well as the least risky, for translational applications still requires further identification. Appropriate pre- processing of BMSCs or combination therapy has the benefit of achieving the maximum possible restoration. Further pre-clinical studies are required to determine the time-window for transplantation and the appropriate dosage of cells.
