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1.
AIM: To investigate the effects of cotransplantation of mesenchymal stem cells (MSCs) and umbilical cord blood (UCB) by intra-bone marrow (IBM) injection on the hematopoietic reconstitution and recovery of bone marrow MSCs in the recipients. METHODS: Wistar female rats were transplanted with fetal and neonatal peripheral blood (FNPB) and BrdU-labeled MSCs separated from BMNCs of F344 rats. The MSCs were infused by IBM injection in bilateral tibiae or intravenous injection (IV), while the FNPB was all via IBM route. The survival rate, reconstitution of hematopoietic and immunological function, engraftment level of HSCs and recovery of bone marrow (BM)-MSCs in recipients were monitored. The origins of BM-MSCs of recipients were examined by immunofluorescence assay. RESULTS: (1)The survival rate in the two cotransplantation groups was 100% at day 60, while that in FNPB group was only 66.7%. (2)The counts of peripheral blood cells and BM hematopoietic stem/progenitor cell colonies of the recipients were better in cotransplantation groups than those in FNPB group, especially in the FNPB (IBM)+MSC (IBM) group. (3)No significant difference between of engraftment level of HSCs in the two cotransplantation groups was observed. The percentage of RT1A1 cells subset in FNPB (IBM)+MSC (IBM) group was much higher than that in FNPB group (P<0.05). (4)At day 30, the growth characteristic of recipient BM-MSCs was still below normal, but that in FNPB (IBM)+MSC (IBM) group was the best of all the experiment groups (P<0.05). (5)The donor MSCs coexisted with host MSCs in only a few recipient rats. CONCLUSION: The cotransplantation of MSCs and FNPB can accelerate the recovery of recipient BM-MSCs and hematopoietic reconstitution, promote the engraftment level of HSCs. Cotransplantation by IBM route is safe and has better effects on hematopoietic reconstitution than by IV route.  相似文献   

2.
AIM:To investigate effects of rBMMSC on he matopoiesis and immune reconstitution after allo-hematopoietic stem cells transp lantation (HSCT).METHODS:Allogeneic BMT model from Fischer344 rats (RT-1Al) to W istar rats (RT-1Au) was established.The effects of MSCs on hematopoietic recons titution and immune reconstitution were studied by observing the survival rate,peripheral blood counts,thymus counts,spleen counts,bone marrow counts and im mune function analysis at 30 days after transplantation.RESULTS:1.Cotransplantation of MSCs and bone marrow (BM) was d emonstrated to improve hematopoietic reconstitution.Lymphocyte and platelet cou nts in peripheral blood in cotransplantation groups were higher than those in co ntrol groups.More bone marrow neucleated cells were also observed in cotranspla ntation groups.2.Cotransplantation of MSCs and BM improved immune reconstituti on.First,overall thymic cellularity and spleen cellularity significantly incre ased in cotransplantation groups at day 30.Secondly,cotransplantation improved immune functional recovery.Non-specific lymphocytes proliferation reaction ind uced by ConA and LPS increased in cotransplantation group,and so did for alloge neic mixed-lymphocyte reaction.CONCLUSION:Hematopoietic reconstitution and immune reconstituti on were significantly enhanced by MSCs cotransplanted with BM.  相似文献   

3.
AIM: To evaluate the effects of bone marrow-derived mesenchymal stem cells (MSCs) on engraftment of hematopoietic stem/progenitor cells in sensitized mice. METHODS: Mouse bone marrow-derived MSCs were cultured by adherent culture method. MSCs combined with or without hematopoietic stem/progenitor cells were implanted into the sensitized mouse model, which was established by allogeneic splenocyte transfusion, and were divided into 6 groups: MSC intervention groups, including sensitized mice with MSCs on day 11, sensitized mice with MSCs on day 0 and sensitized-mice with MSCs both on day 11 and day 0; control groups, including sensitized mice without MSC intervention, non-sensitized mice without MSC intervention and non-sensitized mice without MSCs or transplantation of hematopoietic stem/progenitor cells. The survivors were assessed after transplantation and hematopoietic recovery was monitored weekly including hematological change, immune function reconstruction, bone marrow cell recovery, chimera analysis and graft-versus-host disease development. RESULTS: Compared with different control groups, MSC intervention did not prolong the survival rates of the sensitized model mice after lethal irradiation. CONCLUSION: Under the experimental conditions, MSC combined with C57BL/6 bone marrow hematopoietic stem/progenitor cells fail to promote the growth of engraftment in C57BL/6 allogeneic splenocyte-sensitized BALB/c mice in vivo.  相似文献   

4.
AIM：To compare the biological characteristics, surface markers and multi-differentiation potential of the mesenchymal stem cells (MSCs) derived from the umbilical cord and bone marrow in the green fluorescent protein (GFP) transgenic mice. METHODS：Umbilical cord MSCs (UCMSCs) were isolated by collagen type II enzymatic digestion and bone marrow MSCs (BMSCs) were isolated by density gradient centrifugation. The growth of the 2 types of MSCs was observed under inverted microscope. The cell proliferation was detected by determining the growth curve and MTT assay. The Trypan blue method was performed to analyze the cell viability rate. The cell cycle and cell surface markers were measured by flow cytometry. The differentiation potentials of the 2 types of MSCs were tested by the differentiation kits toward adipocytes and osteoblasts. RESULTS：The UCMSCs attached to the culture surface 1 d after the isolation, and the cells showed spiral shape with notable growth and proliferation after 2 d of culture. After 3 d, the cell arrived sub-confluent and was ready for passage. BMSCs still showed circular shape and started to attach to the surface 4 d after culture. They formed the small colony shape only after 5 d with obvious proliferative potential. The cells became confluent 7 d after the culture. The original generation of cultivating UCMSCs growth curve was shown typically an “S” shape. But the BMSCs growth was slower than the UCMSCs. The cell proliferation was obvious for UC-MSCs in 3~5 d. BMSCs proliferated significantly only after 7 d. The viability rate arrived more than 96% for both types of MSCs. The cell cycle of both MSCs did not show significant difference (G0/G1 phases were above 85%, P>0.05). Both MSCs positively expressed CD44, CD90 and CD105 (60.7%±2.3%) but the expression of CD45, CD19, CD14 and CD79 was negative (less than 25.6%±4.8%, P>0.05). More than 90% of the MSCs from the umbilical cord and bone marrow differentiated towards the adipocytes and osteoblasts without significant difference (P<0.05). CONCLUSION：UCMSCs have stronger ability of proliferation and multi-directional differentiation potentials. UCMSCs in GFP transgenic mice as a high-quality tracer can serve for tracking the stem cells in vivo.  相似文献   

5.
AIM： To study the telomere maintenance mechanism in mesenchymal stem calls (MSCs).〖WT5"HZ〗 METHODS：MSCs were isolated from healthy human bone marrow by their adherence to plastic and then were checked with CD14-FITC,CD45-FITC,CD44-FITC,HLA-DR-FITC,CD34-PE,CD29-PE and CD166-PE.Telomere length and ECTR DNA in MSCs were detected by Southern blotting.The localization of TRF1 and promyelocytic leukemia (PML) in MSCs were detected with immunofluorescence staining.TRAP protocol was performed to detect the telomerase activity in MSCs and MSCs-derived adipocytes.Western blotting and TRAP protocol were applied to measure telomerase activity of MSCs,which were synchronized by serum starvation and aphidicolin treatment.〖WT5"HZ〗RESULTS：The telomere in length seemed shorter and relatively more homogeneous in MSCs and HeLa cells than that in WI-38-2RA cells.TRF1 did not concide with PML nuclear body in MSCs and HeLa cells while it exclusively did in WI-38-2RA cells.ECTR DNA was negative in MSCs and HeLa cells but positive in WI-38-2RA cells.Telomerase was negative in MSCs but it was positive in MSCs-derived adipocytes detected by TRAP.Moreover,a cell cycle-dependent expression profile of telomerase was found in MSCs when they were synchronized by serum starvation and aphidicolin treatment.Untreated MSCs expressed very low level of telomerase probed by Western blotting with 2C4 mAb,but the telomerase level had significantly increased when these cells were trapped in S phase.〖WT5"HZ〗CONCLUSION：The telomere of MSCs is maintained by telomerase pathway instead of alternative lengthing of telomere(ALT) and the level of telomerase expression is associated with cell cycle stage.  相似文献   

6.
AIM：To study the effect of calcitonin gene-related peptide (CGRP) gene transfection mediated by lentivirus on the differentiation of rat bone marrow mesenchymal stem cells (MSCs) to endothelial cells. METHODS：Rat bone marrow MSCs were isolated by density gradient centrifugation combined with adherence method. Recombinant lentivirus vector carrying CGRP gene (Lenti-CGRP) was transfected into the MSCs. The secretion of CGRP in culture supernatants of the transfected MSCs was detected using ELISA method. The cells at passage 3 were divided into three groups: CGRP group (MSCs transfected with Lenti-CGRP), CGRP+CGRP8-37 (an antagonist of CGRP receptor) group and control group (MSCs transfected with PBS). The differentiation of the MSCs was detected by immunocytochemical staining for CD31 and factor Ⅷ-related antigen. The proliferation of the cells was measured by cell counting, and the angiogenic ability of the cells was analyzed using Matrigel assay. RESULTS：The proportion of CD31-and factor Ⅷ-related antigen-positive cells in CGRP and CGRP+CGRP8-37 groups was larger than that in control group (P<0.05). The numbers of the cells in CGRP and CGRP+CGRP8-37 groups were significantly increased compared with control group (P<0.05). Lumen-like structures were observed in CGRP and CGRP+CGRP8-37 groups. The above indexes in CGRP+CGRP8-37 group were reduced compared with CGRP group. CONCLUSION: Transfection with CGRP gene induces rat bone marrow MSCs to differentiate into endothelial cells and enhances their proliferation, suggesting that CGRP may play a role in the regulation of angiogenesis.  相似文献   

7.
AIM: To explore the supportive and expansive effects of aorta-gonad-mesonephros(AGM) region derived stromal cells on hematopoietic stem cells (HSC) in vitro.METHODS: The murine stromal cells were separated and cultured from AGM region of a 11 day postcoitum (dpc) mouse embryo and 6 week mouse. After identification by Wrights staining and flow cytometry, the stromal cells were co-cultured with the embryonic stem cell(ESC)-derived, cytokine-induced HSCs, and the maintenance and expansion of HSCs were evaluated by detecting CD34+，CD34+Sca-1+cells with flow cytometry.Blast colony-forming cell (BL-CFC) and high proliferative potential colony-forming cells(HPP-CFC) were determined by semi-solid medium clonal culture.RESULTS: AGM-derived and bone marrow(BM)-derived stromal cells were similar in morphology and phenotype, and had common character of stromal cells. Supported by AGM stromal cells or by BM stromal cells, more primitive progenitor cells HPP-CFC were expanded, but BL-CFC expansion was only detected in AGM-derived stromal cells. In the supporting of BM stromal cells CD34+ hematopoietic stem/progenitor cells were expanded 3-4 times, but no significant expansion in CD34+Sca-1+ cells was observed. While in the supporting of AGM stromal cells, both CD34+ hematopoietic stem/progenitor cells and CD34+Sca-1+ cells were expanded significantly from 4 to 5 times, respectively (P<0.05).CONCLUSION: AGM-derived stromal cells significantly support the expansion of HSC, and also maintain the self-renewal activity and multi-lineage differentiation of HSC in vitro.  相似文献   

8.
南京不同类型梅花品种香气成分的比较研究   总被引：10，自引：1，他引：9  
金荷仙  陈俊愉  金幼菊 《园艺学报》2005,32(6):1139-1142
1 材料与方法 采用活体植株动态顶空套袋采集法和TCT／GC／MS（热脱附／气相色谱／质谱联用）技术分析南京梅花山真梅种系直枝梅类宫粉型‘玉露宫粉’、玉蝶型‘宇治里’、黄香型‘黄金鹤’、朱砂型‘姬千鸟’、绿萼型‘变绿萼’、洒金型‘复瓣晚跳’及垂枝梅类白碧垂枝型‘双碧垂枝’的香气组成。2002年3月513采样，将采样后的吸附管套上聚四氟乙烯套，放在干燥器中低温保存。样品分析时间为2002年4月27日。同时采集和分析空气作对照。采用Xcalibur 1．2版本软件及NIST98谱图库进行梅花香气成分的检索，兼顾挥发物出峰的保留时间鉴定。  相似文献   

9.
AIM: To evaluate the biological characteristics and differentiating potentials of bone marrow-derived mesenchymal stem cells (MSCs) from sensitized mice by allogeneic splenocyte transfusion in vitro. METHODS: Adherent culture method was applied for culturing the bone marrow-derived MSCs from sensitized mice. The cell morphology was examined and the surface marker profiles were analyzed by flow cytometry. The differentiating potentials of the MSCs into osteogenic, adipogenic and myogenic lineages were explored. The bone marrow-derived MSCs from the normal mice were collected and served as controls. RESULTS: Both the bone marrow-derived MSCs from sensitized and normal mice were exhibited a homogeneous distinctive morphology and were positive for the surface markers CD29, CD105, CD44 and Sca-1, negative in CD 34 and CD11b. The abilities of both MSCs to differentiate into osteogenic, adipogenic and myogenic pathways in the same condition were also observed. CONCLUSION: There is no difference in the biological characteristics and induced differentiating potentials between the sensitized mouse bone marrow-derived MSCs by allogeneic splenocytes transfusion and the MSCs from normal mice.  相似文献   

10.
AIM: Our purpose was to induce MSCs differentiating into endothelial cells (EC) in vitro and to provide the seed cells for study of cardiovascular tissue-engineering. METHODS: MSCs were separated by gradient centrifugation on Percoll (density 1 073 g/L) from human bone marrow (HBM), and incubated for purification and amplification in DMEM (low glucose) with 10% fetal bovine serum (FBS). Then, the MSCs were incubated for orientation differentiated into EC in DMEM (high glucose) with 20% FBS, VEGF (10 μg/L), bFGF (5 μg/L), L-glutamine (2 mmol/L), penicillin (1×105U/L) and streptomycin (100 mg/L) for about 14-21 days and their phenotypic characteristics were analyzed by flow cytometry. Afterwards, the differentiating cells were evaluated by histology and immunohistochemistry. RESULTS: The quantity of MSCs was increased from 5.0×105 in the primary culture to 8.0×1012, or to increase to 1.6×107 times after 15 generations of incubation. The purity of MSCs was above 95% and 98% homogeneous at passages 2 and 3, respectively. About 80%-90% of the differentiating cells from MSCs after 14-21 days were positively stained for Ⅷ factor (vWF) related antigen by immunohistochemistry assay, and Weible-palade corpuscle was also observed by transmission electron microscopy in the cytoplasm. CONCLUSION: MSCs from HBM have the capability of differentiation into ECs in vitro, which may be a potential source of seed cells for fabrication of tissue-engineering heart valve, particularly in children with congenital heart disease.  相似文献   

11.
AIM：To investigate the effects of caspase-3 gene silencing on proliferation, cell cycle and apoptosis of rat bone marrow mesenchymal stem cells (MSCs). METHODS：A lentiviral vector expressing caspase-3 shRNA was constructed and transfected into rat bone marrow MSCs．The expression of caspase-3 at mRNA and protein levels was detected by real-time PCR and Western blotting, respectively. Cell proliferation and cell cycle were evaluated by MTS assay and flow cytometry, respectively. The expression of bcl-2 and bax mRNA was detected by real-time PCR. The apoptosis of the cells was evaluated by Hoechst 33258 staining. RESULTS：Recombinant lentivirus was successfully transfected into MSCs. The proliferation of the MSCs transfected with caspase-3 shRNA was significantly promoted (P<0.05) and the proportion of the cells in S phase was increased to (52.66±0.30) %. Compared with control groups, caspase-3 silencing up-regulated the mRNA level of bcl-2 and down-regulated the mRNA level of bax, and the ratio of bcl-2 to bax increased (P<0.05). The apoptotic rate in MSCs-shRNA group was (15.01±1.73) %, which was significantly lower than those in MSCs and MSCs-vector group ［(23.67±1.16) % and (25.67±3.05) %, respectively; P<0.05］. CONCLUSION: Caspase-3 silencing regulates cell cycle, promotes the proliferation and attenuates the apoptosis of rat bone marrow MSCs.  相似文献   

12.
AIM：To estimate the neural differentiation efficiency of bone marrow mesenchymal stem cells (MSCs) derived from amyloid precursor protein (APP) transgenic mice and to investigate the correlation with Notch1 signaling and the autophagy activity during the differentiation. METHODS：The MSCs were divided into APP group (MSCs from APP transgenic mice) and WT group (MSCs from wild-type mice). MSCs were treated with β-mercaptoethanol as an inducer for differentiating into neurons. The levels of Aβ40 and Aβ42 were measured using enzyme-linked immunosorbent assay kits. The expression of neural cell-specific markers, neuron-specific enolase (NSE) and microtubule-associated protein 2 (MAP-2), was measured by immunocytochemistry and Western blotting. The expression levels of Notch1, Notch intracellular domain (NICD), Hes5, LC3 and p62 (a selective substrate of autophagy) were also detected by Western blotting. RESULTS：The neural differentiation capacity and the Aβ expression level of the MSCs in APP group were higher than those in WT group, and stronger inhibition of Notch1 signaling pathway in the MSCs from APP group was observed. However, the process of autophagy, which is essential for the survival and function of the neural cells, was impaired in the neural differentiated counterpart of the MSCs in APP group. CONCLUSION:Over-expression of APP might contribute to the high neural differentiation capacity of MSCs by inhibiting Notch1 signaling pathway in vitro. However, autophagy is impaired in the differentiated MSCs from APP transgenic mice.  相似文献   

13.
AIM:To investigate the changes of biological characteristics of GFP transgenic mouse bone marrow mesenchymal stem cells (MSCs), which were transfected with human β-nerve growth factor (β-NGF) gene in a recombinant eukaryotic expression vector. METHODS:MSCs obtained from GFP transgenic mice were isolated, cultured and purified by the whole bone marrow adherence methods. Human β-NGF gene was transfected into the MSCs by a recombinant eukaryotic expression vector. The β-NGF expression in the MSCs was detected by the method of immunocytochemistry. Hippocampal neurons from neonatal mice were cultured with culture supernatant of the MSCs transfected with pcDNA3-β-NGF and the biological characteristics of the MSCs were investigated 3 d after culture under inverted phase-contrast microscope. RESULTS:The β-NGF positive rate of MSCs in pcDNA3-β-NGF transfection group ［(37.12±2.14)%］ was significantly higher than that in MSCs control group ［(2.36±0.62)%］ and blank control group ［(1.43±0.76)%］.The neurite length of neonatal mouse hippocampal neurons cultured with culture supernatant from pcDNA3-β-NGF-transfected MSCs ［(31±3)μm］ was significantly longer than that in negative control group ［(23±4)μm］, suggesting that MSCs transfected with β-NGF gene maintained better biological characteristics. CONCLUSION: The constructed recombinant eukaryotic expression vector of human β-NGF gene can be transfected into MSCs efficiently and NGF can be effectively expressed in MSCs. MSCs transfected with β-NGF gene are capable of stable expression and secretion of β-NGF, and maintenance of better biological characteristics.  相似文献   

14.
AIM：To investigate the regulatory function of bone marrow-derived mesenchymal stem cells (MSCs) on T helper 17 cells (Th17) and regulatory T cells (Treg) in peripheral blood of severe asthmatic children. METHODS：MSCs were isolated, cultured and identified in vitro. MSCs digested with mitomycin were cocultured with T lymphocytes (TLC) at different ratios (1∶1, 1∶2, 1∶10 and 1∶20) from severe asthmatic children for 72 h. The proliferation of TLC was measured by CCK-8 method. In the coculture system of the 1∶2 ratio and the single TLC system, the supernatant levels of interleukin-17 (IL-17) and transforming growth factor-β (TGF-β) were measured by ELISA. The mRNA expression of retinoic acid-related orphan nuclear receptor C (RORC) and forkhead box protein 3 (Foxp3) in TLC was detected by qRT-PCR. RESULTS：After cocultured with MSCs, the proliferation of TLC decreased significantly in a dose-dependent manner (P<0.05). It also showed decreases in IL-17 (3 799±441 vs 4 890 ±373, P<0.05) and RORC mRNA level (1.21±0.14 vs 3.85±0.48, P<0.05), while an increase in TGF-β level (209±32 vs 117±26, P<0.05) was observed. No influence on the mRNA expression of Foxp3 was found (P>0.05). CONCLUSION：MSCs suppresses Th17 polarization of naive peripheral blood CD4+ T cells and matures Th17 cells secreting IL-17, which may effectively revise Th17/Treg imbalance of asthma.  相似文献   

15.
AIM: To investigate the isolation, purification, expansion and multilineage differentiation of mesenchymal stem cells (MSCs) derived from human umbilical cord vein in vitro.METHODS: By 1% collagenase Ⅱ digestion, endothelial cells were isolated from human umbilical cord vein and cultured in IMDM medium.The morphology of the cells was observed by Wright’s staining and electron microscope.Cell cycle and immunophenotype were investigated by flow cytometry.Assays of adipogenic and osteogenic differentiation were performed in vitro.von Kossa staining, Oil Red O staining and mRNA expression of osteopontin and lipoprotein lipase were studied in the induced cells.RESULTS: The cells from the cord vein displayed a fibroblast-like morphology adhering to the culture plate.FACS showed that the cells expressed several MSCs-related antigens such as CD29, CD44 and CD105, while CD13, CD31, CD45, CD34, and HLA-DR were negative.Adipocyte and osteocyte differentiation were induced successfully.CONCLUSION: The morphology, growth characteristics, immunophenotype and pluripotentiality of the MSCs from human umbilical cord vein are similar to the MSCs from bone marrow (BM).They could potentially be an excellent source of MSCs for experiments and clinics.  相似文献   

16.
AIM: To investigate adrenomedullin gene transfection enhances the therapeutic effects of homogeneous transplantation of bone marrow mesenchymal stem cells (MSCs) on cardiac function and ventricle remodeling in acute myocardial infarction rats. METHODS: MSCs were isolated and expanded using the preplating method. The infection efficiency of adenovirus vector to MSCs was tested by X-gal staining. Ad-ADM expression in MSCs and its secretion in culture medium were measured by ELISA. The left anterior descending branch of rats was ligated to establish a myocardial infarction model. The MSCs were labeled by DAPI, and were directly implanted into the acute infarct site via focal injection. Four weeks later, cardiac function was evaluated using physiological recorder. Hearts were harvested and sliced to be analyzed by immunohistochemistry (factor Ⅷ and ADM) and the DAPI-labeled cells were identified. Sirius red staining was used to identify interstitial collagen on slides. Analysis of collagen type I and III was performed using a polarized filter on sections stained for collagen with Sirius red, and the ratio of collagen type I and III were detected. RESULTS: With X-gal staining, MSCs were effectively transfected by adenovirus in vitro. The transfection efficiency showed the dose-effect relationship with multiplicities of infection (MOI). When MOI was 150, the infection efficiency was 95.4%. The expression of ADM was traced in culture medium and expressed in the time-dependent manner. A maximum production of ADM was observed at 7 d after infection ［(26.53±1.42) ng/L vs (1.34±0.08) ng/L, P<0.05］, and ADM secretion reduced to normal level at 15 d ［(2.20±1.44) ng/L vs (1.52±0.33) ng/L, P>0.05］. DAPI-labeled MSCs transplantation was found in the hearts of the recipients. Immunohistochemical studies demonstrated that intense immunostaining for ADM was higher in Ad-ADM plus MSCs group, compared to other groups. Compared with control, MSCs transplantation significantly increased capillary density in infarct area (P<0.01). A combination of Ad-ADM trensfection and MSCs transplantation demonstrated a further increase in capillary density compared with Ad-ADM or MSCs alone. MSCs transplantation decreased the ratio of collagen type I and III, obviously improved the left ventricular functions. Furthermore the combination treatment resulted in further decrease in the ratio of collagen type I and III, and significantly improved the left ventricular functions. CONCLUSION: Ad-ADM transfection enhances the angiogenic potency of MSCs transplantation and decreases the ratio of collagen type I and III through increasing ADM expression in infarct area, thus contributes to reverse the ventricular remodeling and improves the cardiac function.  相似文献   

17.
AIM：To explore the effects of rat mesenchymal stem cells (MSCs) modified by calcitonin gene-related peptide (CGRP) on the proliferation and phenotype transformation of rat vascular smooth muscle cells (VSMCs) in vitro. METHODS：Rat MSCs and VSMCs were isolated, cultured and identified. The MSCs were infected by lentivirus which carried genes encoding enhanced green fluorescent protein (EGFP) and CGRP. The expression levels of CGRP in CGRP-modified MSCs were detected using real-time PCR and enzyme-linked immunosorbent assay (ELISA). The prolife-ration and migratory abilities of VSMCs were evaluated by MTT assay, Trypan blue staining and scratch test. The expression levels of α-smooth muscle actin (α-SMA) and osteopontin (OPN) were assessed by Western blotting. RESULTS：Compared with MSCs and MSCs-EGFP groups, the expression levels of CGRP in MSCs-CGRP group were markedly increased (P<0.01). The results of MTT assay and scratch test demonstrated that the proliferation and migratory abilities of VSMCs in MSCs-CGRP group were significantly inhibited compared with MSCs and MSCs-EGFP groups (P<0.05). Furthermore, cell viability was >90% shown by Typan blue staining. Western blotting showed that the expression of α-SMA was increased and that of OPN was decreased in MSCs-CGRP group compared with MSCs and MSCs-EGFP groups (P<005). CONCLUSION：CGRP-modified MSCs could secrete CGRP protein and inhibit the proliferation and migration of VSMCs, which may be associated with deterring the phenotype transformation from contractile type to synthetic type. These results lay a foundation for gene therapy in vivo.  相似文献   

18.
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19.
WANG Yue-chun  ZHANG Yuan 《园艺学报》2007,23(11):2205-2209
AIM: To separate and identify the mesenchymal stem cells (MSCs) from human fetal bone and to study their differentiation to hepatocyte like cells under the action of chemical induction.METHODS: The MSCs from human fetal bone were isolated and purified according to the different growth characteristic of attaching to the wall of cell culture flask.The cell cycle and surface markers of MSCs were identified using flow cytometry.The MSCs were pre-induced by adding DMSO,β-Me and 5-aza for 24 h,then adding the inductive medium of H-DMEM and rh-HGF to induce their differentiation to hepatocyte like cells (HLCs).HLCs were identified by the typical morphological change and the expression of special protein with the method of immunocytochemistry.RESULTS: The MSCs derived from human fetal bone expressed adhesion molecules CD29+,CD44+,but not antigens of hematopoietic CD34,CD45,and not antigens related to GVHD,such as HLA-DR,CD80 and CD86.Exposure of these cells to above-mentioned inductive agents resulted in obvious morphological change and an increase in expression of AFP and ALB.CONCLUSION: The results suggest the existence of plentiful MSCs in human fetal bone.MSCs derived from human fetal bone can easily differentiate to HLCs,and they have a lower immunogenic nature,which may provide the ideal source for tissue engineering (bioartificial liver) for cellular therapeutics.  相似文献   

20.
AIM:To induce lymphoid stem cells and/or T-cell precursors to differentiate into functional mature T lymphocyte, and to increase the surface marker of T lymphocytes such as CD3+, while embryonic stem(ES) cells differentiated into hematopoietic stem/progenitor cells(HSPCs) in vitro. When they were injected into lethally irradiated mice, these differentiated cells had the advantage in immune reconstitution. METHODS:Embryonic stem cells formed embryoid bodies(EBs) in the medium containing methycellulose, hematopoietic growth factors(HGFs) was added to the culture system on the 6th day, thymopeptide was added at the same time. Flow cytometry were performed to detect the surface marker CD34+ and CD3+ of the differentiated cells. Finally the differentiated cells were injected into lethally irradiated mice, 60 days later, the incidence rate of graft versus host disease(GVHD) was taken as the mark of cell mediated immunity, PCR was performed to detect the sex determining region of the Y-chromosome(Sry) in bone marrow cells and spleen cells of the survival host female mice. RESULTS:The percentage of CD3+ T lymphocytes was 10.52% and the incidence rate of GVHD was 0% on the 13th day, but they respectively rose up to 22.93% and 100% if thymopeptide was added in the procedure of inducing ES cells to differentiate into HSPC in vitro. CONCLUSION:The quantity of CD3+ T lymphocytes increased in medium containing thymopeptide when ES cells differentiated into CD34+ HSPC.  相似文献   

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