| 摘要: |
| 目的 模拟体内急性心肌梗死缺血缺氧微环境，探讨血清剥夺是否能激活心肌成纤维细胞向内皮细胞转分化。方法 取新鲜分离的C57BL/6新生小鼠心脏组织，获取成纤维细胞进行培养，用白血病抑制因子（LIF）促进成纤维细胞自我更新、长程维持干性和抑制其向终末分化。将第3代成纤维细胞分为：对照组[DMEM+10%胎牛血清（FBS）培养]、血清剥夺24 h组（不含FBS的DMEM培养24 h）、血清剥夺48 h组（不含FBS的DMEM培养48 h）、血清剥夺72 h组（不含FBS的DMEM培养72 h）。经过不同处理后，采用qPCR法检测各组细胞谱系特异性基因的表达变化，采用体外血管新生实验和Dil-Ac-LDL吞噬实验检测血清剥夺后成纤维细胞的功能学变化，采用ELISA法检测血清剥夺后成纤维细胞分泌血管内皮生长因子（VEGF）的情况。结果 血清剥夺不同时间组成纤维细胞形成血管分叉的数目与对照组相比均增加（P<0.05），但均未发现诱导细胞吞噬Dil-Ac-LDL的现象。与对照组比较，血清剥夺48 h组成纤维细胞中内皮特异性基因CD31和VE-herin的表达均升高（P<0.05）；其中血清剥夺48 h组成纤维细胞中CD31的表达是对照组的13.7倍（P<0.05），血清剥夺72 h组是对照组的2 倍（P<0.05）。ELISA检测结果显示，血清剥夺48 h组成纤维细胞分泌的VEGF是对照组的7倍（P<0.05），血清剥夺72 h组是对照组的3.7倍（P<0.05）。结论 血清剥夺可以激活心肌成纤维细胞向内皮细胞谱系的转分化。 |
| 关键词: 成纤维细胞 内皮细胞 细胞转分化 缺血性心脏病 血管新生 血清剥夺 |
| DOI：10.16781/j.0258-879x.2017.05.0602 |
| 投稿时间：2017-02-24修订日期：2017-04-16 |
| 基金项目:国家自然科学基金（9153910092，91539118，81473445，8151101216）. |
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| Serum deprivation promoting transdifferentiation of fibroblasts into endothelial cells: an in vitro study |
| ZHAO Xiao-feng,HE Zhi-qing,WU Zong-gui,LIANG Chun* |
(Department of Cardiology, Changzheng Hospital, Second Military Medical University, Shanghai 200003, China
*Corresponding author) |
| Abstract: |
| Objective To determine whether serum deprivation can induce transdifferentiation of cardiac fibroblasts into endothelial cells in the ischemic and hypoxic microenvironment of acute myocardial infarction. Methods Fibroblasts were separated and cultured from hearts of new born C57BL/6 mice, and leukemia inhibitory factor (LIF) was used to accelerate the self-renewal, keep plasticity and inhibit differentiation to the terminal fates. The fibroblasts were exposed to five different intervention conditions:control group (DMEM+10% fetal bovine serum[FBS]) and serum deprivation (DMEM without FBS) for 24 h, 48 h and 72 h groups. qPCR was used to detect the expression of cell lineage specific genes; the in vitro angiogenesis test and Dil-Ac-LDL phagocytic function test were used to observe function of fibroblasts; and ELISA assay was used to examine the level of vascular endothelial growth factor (VEGF) secreted by fibroblasts. Results Compared with the control group, the number of capillaries-like bifurcation formed by fibroblasts was significantly increased at different serum deprivation time points (P<0.05), but no induced engulfment of Dil-Ac-LDL was noticed. Compared with the control group, the expressions of endothelial specific genes CD31 and VE-herin in fibroblasts were significantly increased at 48 h of serum deprivation (P<0.05); the expression of CD31 at 48 h of serum deprivation was 13.7 times that of the control group (P<0.05), and at 72 h of serum deprivation it was two times that of the control group (P<0.05). ELISA results showed that VEGF level at 48 h of serum deprivation was seven times that of the control group, and at 72 h of serum deprivation it was 3.7 times (P<0.05). Conclusion The serum deprivation can stimulate the transdifferentiation of cardiac fibroblasts into endothelial cell lineages. |
| Key words: fibroblasts endothelial cells cell transdifferentiation ischemic heart disease angiogenesis serum deprivation |
