Objective To evaluate the effects endothelial progenitor cells （EPCs） survival in vitro of serum deprivation （SD） and hypoxia on the bone marrow Methods Rat bone marrow EPCs were exposed for 48 h to 02 deprivation...Objective To evaluate the effects endothelial progenitor cells （EPCs） survival in vitro of serum deprivation （SD） and hypoxia on the bone marrow Methods Rat bone marrow EPCs were exposed for 48 h to 02 deprivation, serum deprivation （ SD ） , and prolonged （ 120 h） hypoxia concomitant with serum deprivation. Cell death was assessed by Live/Dead staining and image analysis. Reaulta The EPCs death rate seemed not affected by 48 h hypoxia （ P 〉 0. 05 ）, but affected by SD （ P 〈 0. 01 ）. Prolonged hypoxia concomitant with SD resulted in the death of nearly all EPCs from 72 h, but this rate was remarkably reduced when with 20% fetal bovine serum（ P 〈 0. 01 ）. Conclusion Our findings indicate that EPCs are sensitive to hypoxia/SD stimuli, while serum may be the more important factor for EPCs survival.展开更多
Oligodendrocyte lineage gene-1 expressed in oligodendrocytes may trigger the repair of neuronal myelin impairment, and play a crucial role in myelin repair. Hypoxia-inducible factor la, a transcription factor, is of g...Oligodendrocyte lineage gene-1 expressed in oligodendrocytes may trigger the repair of neuronal myelin impairment, and play a crucial role in myelin repair. Hypoxia-inducible factor la, a transcription factor, is of great significance in premature infants with hypoxic-ischemic brain damage There is little evidence of direct regulatory effects of hypoxia-inducible factor le on oligodendrocyte lineage gene-l. In this study, brain slices of Sprague-Dawley rats were cultured and subjected to oxygen-glucose deprivation. Then, slices were transfected with hypoxia-inducible factor la or oligodendrocyte lineage gene-1. The expression levels of hypoxia-inducible factor la and oligodendrocyte lineage gene-1 were significantly up-regulated in rat brains prior to transfection, as detected by immunohistochemical staining. Eight hours after transfection of slices with hypoxia-inducible factor la, oligodendrocyte lineage gene-1 expression was upregulated, and reached a peak 24 hours after transfection. Oligodendrocyte lineage gene-1 transfection induced no significant differences in hypoxia-inducible factor la levels in rat brain tissues with oxygen-glucose deprivation. These experimental findings indicate that hypoxia-inducible factor la can regulate oligodendrocyte lineage gene-1 expression in hypoxic brain tissue, thus repairing the neural impairment.展开更多
Background The proliferation and apoptosis property of mesenchymal stem cells derived from peripheral blood （PB-MSCs） were investigated under hypoxia and serum deprivation conditions in vitro so as to evaluate the f...Background The proliferation and apoptosis property of mesenchymal stem cells derived from peripheral blood （PB-MSCs） were investigated under hypoxia and serum deprivation conditions in vitro so as to evaluate the feasibility for autologous PB-MSCs applications in cartilage repair. Methods MSCs were mobilized into peripheral blood by granulocyte colony stimulating factor （G-CSF） and AMD3100. The blood samples were collected from central ear artery of rabbits. Adhered cells were obtained by erythrocyte lysis buffer and identified as MSCs by adherence to plastic, spindle shaped morphology, specific surface markers, differentiation abilities into osteoblasts, adipocytes and chondroblasts in vitro under appropriate conditions. MSCs were cultured in four groups at different oxygen tension （20% 02 and 2% O2）, with or without 10% fetal bovine serum （FBS） conditions： 20% 02 and 10% FBS complete medium （normal medium, N）, 20% 02 and serum deprivation medium （D）, 2% 02 and 10% FBS complete medium （hypoxia, H）, 2% 02 and serum deprivation （HD）. Cell proliferation was determined by CCK-8 assay. Apoptosis was detected by Annexin V/PI and terminal deoxynucleotide transferase dUTP nick end labeling （TUNEL） staining. Results Spindle-shaped adherent cells were effectively mobilized from peripheral blood by a combined administration of G-CSF plus AMD3100. These cells showed typical fibroblast-like phenotype similar to MSCs from bone marrow （BM-MSCs）, and expressed a high level of typical MSCs markers CD29 and CD44, but lacked in the expression of hematopoietic markers CD45 and major histocompatibility complex Class II （MHC II）. They could also differentiate into osteoblasts, adipocytes and chondroblasts in vitro under appropriate conditions. No significant morphological differences were found among the four groups. It was found that hypoxia could enhance proliferation of PB-MSCs regardless of serum concentration, but serum deprivation inhibited proliferation at the later stage of culture. Apart from that, hypoxia or serum deprivation could promote the apoptosis of PB-MSCs after 48 hours; the effect was stronger when these two conditions combined together. Furthermore, the effect of serum deprivation on apoptosis was stronger compared with that of hypoxia. Conclusions PB-MSCs possess similar phenotypes as BM-MSCs. Their differentiation and proliferation abilities make them a new source of seed cells for ischemia-related cell therapy and tissue engineering in the field of the articular cartilage repair.展开更多
BACKGROUND： Intercellular adhesion molecule-5 （ICAM-5） relieves the damage of beta-amyloid protein to PAJU cells, However, little is known about how ICAM-5 works as a neurotrophic factor, or whether ICAM-5 lessens ...BACKGROUND： Intercellular adhesion molecule-5 （ICAM-5） relieves the damage of beta-amyloid protein to PAJU cells, However, little is known about how ICAM-5 works as a neurotrophic factor, or whether ICAM-5 lessens neuronal damage under ischemic conditions following cerebral infarction. OBJECTIVE： To investigate the effects of ICAM-5 on PAJU cells growth in serum-free medium under ischemic conditions following cerebral infarction. DESIGN, TIME AND SETTING： The cytological in vitro study was performed at the Central Laboratory, Second Xiangya Hospital, Central South University, China, in June 2009. MATERIALS： Human ICAM-5 gene transfected into PAJU-TLN cells was supplied by the Life Science College, Helsinki University, Finland. Empty vector transfected PAJU-NEO cells were established by the Gene Center, Second Xiangya Hospital, Central South University, China. METHODS： PAJU-TLN cells transfected with human ICAM-5 or empty vector were incubated in serum-free medium. MAIN OUTCOME MEASURES： Phase contrast microscopy was used to observe changes in PAJU cell morphology. 3-（4, 5-dimethylthiazolzyl）-2, 5-diphenyltetrazolium bromide was used to determine cell viability. Hoechst 33258 was used to stain cell nuclei. Flow cytometry was utilized to measure the apoptosis rate of both PAJU-TLN and PAJU-NEO cells. RESULTS： Both PAJU-TLN and PAJU-NEO cells were injured by cultivating in serum-free medium, but the survival rate of PAJU-TLN cells was significantly higher. CONCLUSION： ICAM-5 protects PAJU-TLN cells from serum deprivation-induced apoptosis, induces the outgrowth of PAJU cells, and diminishes their morphologic impairment.展开更多
基金Supported by National Nature Science Foundation of China(30170930)Shanghai Nature Science Foundation,China(044119715,08ZR1413600)
文摘Objective To evaluate the effects endothelial progenitor cells （EPCs） survival in vitro of serum deprivation （SD） and hypoxia on the bone marrow Methods Rat bone marrow EPCs were exposed for 48 h to 02 deprivation, serum deprivation （ SD ） , and prolonged （ 120 h） hypoxia concomitant with serum deprivation. Cell death was assessed by Live/Dead staining and image analysis. Reaulta The EPCs death rate seemed not affected by 48 h hypoxia （ P 〉 0. 05 ）, but affected by SD （ P 〈 0. 01 ）. Prolonged hypoxia concomitant with SD resulted in the death of nearly all EPCs from 72 h, but this rate was remarkably reduced when with 20% fetal bovine serum（ P 〈 0. 01 ）. Conclusion Our findings indicate that EPCs are sensitive to hypoxia/SD stimuli, while serum may be the more important factor for EPCs survival.
基金supported by the National Natural Science Foundation of China,No. 81241022the Natural Science Foundation of Beijing,No. 7072023,7122045
文摘Oligodendrocyte lineage gene-1 expressed in oligodendrocytes may trigger the repair of neuronal myelin impairment, and play a crucial role in myelin repair. Hypoxia-inducible factor la, a transcription factor, is of great significance in premature infants with hypoxic-ischemic brain damage There is little evidence of direct regulatory effects of hypoxia-inducible factor le on oligodendrocyte lineage gene-l. In this study, brain slices of Sprague-Dawley rats were cultured and subjected to oxygen-glucose deprivation. Then, slices were transfected with hypoxia-inducible factor la or oligodendrocyte lineage gene-1. The expression levels of hypoxia-inducible factor la and oligodendrocyte lineage gene-1 were significantly up-regulated in rat brains prior to transfection, as detected by immunohistochemical staining. Eight hours after transfection of slices with hypoxia-inducible factor la, oligodendrocyte lineage gene-1 expression was upregulated, and reached a peak 24 hours after transfection. Oligodendrocyte lineage gene-1 transfection induced no significant differences in hypoxia-inducible factor la levels in rat brain tissues with oxygen-glucose deprivation. These experimental findings indicate that hypoxia-inducible factor la can regulate oligodendrocyte lineage gene-1 expression in hypoxic brain tissue, thus repairing the neural impairment.
基金the National Natural Science Foundation of China，the Key Doctoral Foundation of Chinese Education Commission，the Beijing Natural Science Foundation
文摘Background The proliferation and apoptosis property of mesenchymal stem cells derived from peripheral blood （PB-MSCs） were investigated under hypoxia and serum deprivation conditions in vitro so as to evaluate the feasibility for autologous PB-MSCs applications in cartilage repair. Methods MSCs were mobilized into peripheral blood by granulocyte colony stimulating factor （G-CSF） and AMD3100. The blood samples were collected from central ear artery of rabbits. Adhered cells were obtained by erythrocyte lysis buffer and identified as MSCs by adherence to plastic, spindle shaped morphology, specific surface markers, differentiation abilities into osteoblasts, adipocytes and chondroblasts in vitro under appropriate conditions. MSCs were cultured in four groups at different oxygen tension （20% 02 and 2% O2）, with or without 10% fetal bovine serum （FBS） conditions： 20% 02 and 10% FBS complete medium （normal medium, N）, 20% 02 and serum deprivation medium （D）, 2% 02 and 10% FBS complete medium （hypoxia, H）, 2% 02 and serum deprivation （HD）. Cell proliferation was determined by CCK-8 assay. Apoptosis was detected by Annexin V/PI and terminal deoxynucleotide transferase dUTP nick end labeling （TUNEL） staining. Results Spindle-shaped adherent cells were effectively mobilized from peripheral blood by a combined administration of G-CSF plus AMD3100. These cells showed typical fibroblast-like phenotype similar to MSCs from bone marrow （BM-MSCs）, and expressed a high level of typical MSCs markers CD29 and CD44, but lacked in the expression of hematopoietic markers CD45 and major histocompatibility complex Class II （MHC II）. They could also differentiate into osteoblasts, adipocytes and chondroblasts in vitro under appropriate conditions. No significant morphological differences were found among the four groups. It was found that hypoxia could enhance proliferation of PB-MSCs regardless of serum concentration, but serum deprivation inhibited proliferation at the later stage of culture. Apart from that, hypoxia or serum deprivation could promote the apoptosis of PB-MSCs after 48 hours; the effect was stronger when these two conditions combined together. Furthermore, the effect of serum deprivation on apoptosis was stronger compared with that of hypoxia. Conclusions PB-MSCs possess similar phenotypes as BM-MSCs. Their differentiation and proliferation abilities make them a new source of seed cells for ischemia-related cell therapy and tissue engineering in the field of the articular cartilage repair.
文摘目的在体外用低氧无血清条件模拟缺血心肌微环境,探讨心肌细胞皮肤桥蛋白(DPT)表达的变化。方法以低氧无血清处理乳鼠心肌细胞0、6、12和24 h,用实时反转录聚合酶链式反应检测DPT mRNA的表达变化,用Western blot检测DPT的蛋白表达变化。用酶联免疫吸附实验(ELISA)证实低氧无血清处理乳鼠心肌细胞24 hDPT蛋白的表达变化。结果与0 h组相比,低氧无血清处理心肌细胞6、12、24 h DPT的mRNA和蛋白水平均显著升高(P<0.05)。用ELISA法也检测到低氧无血清处理心肌细胞24 h DPT的表达水平高于正常对照组(P<0.05)。结论低氧无血清可促进乳鼠心肌细胞DPT的表达,DPT可能在缺血缺氧引起的心室重构中发挥一定的作用。
文摘BACKGROUND： Intercellular adhesion molecule-5 （ICAM-5） relieves the damage of beta-amyloid protein to PAJU cells, However, little is known about how ICAM-5 works as a neurotrophic factor, or whether ICAM-5 lessens neuronal damage under ischemic conditions following cerebral infarction. OBJECTIVE： To investigate the effects of ICAM-5 on PAJU cells growth in serum-free medium under ischemic conditions following cerebral infarction. DESIGN, TIME AND SETTING： The cytological in vitro study was performed at the Central Laboratory, Second Xiangya Hospital, Central South University, China, in June 2009. MATERIALS： Human ICAM-5 gene transfected into PAJU-TLN cells was supplied by the Life Science College, Helsinki University, Finland. Empty vector transfected PAJU-NEO cells were established by the Gene Center, Second Xiangya Hospital, Central South University, China. METHODS： PAJU-TLN cells transfected with human ICAM-5 or empty vector were incubated in serum-free medium. MAIN OUTCOME MEASURES： Phase contrast microscopy was used to observe changes in PAJU cell morphology. 3-（4, 5-dimethylthiazolzyl）-2, 5-diphenyltetrazolium bromide was used to determine cell viability. Hoechst 33258 was used to stain cell nuclei. Flow cytometry was utilized to measure the apoptosis rate of both PAJU-TLN and PAJU-NEO cells. RESULTS： Both PAJU-TLN and PAJU-NEO cells were injured by cultivating in serum-free medium, but the survival rate of PAJU-TLN cells was significantly higher. CONCLUSION： ICAM-5 protects PAJU-TLN cells from serum deprivation-induced apoptosis, induces the outgrowth of PAJU cells, and diminishes their morphologic impairment.
