siRNA介导的GDF11沉默对人脐静脉内皮细胞衰老的影响

Effect of siRNA-mediated GDF11 Silencing on the Senescence of Human Umbilical Vein Endothelial Cells

中文关键词: 生长分化因子11  细胞衰老  细胞周期

英文关键词: growth differentiation factor 11 (GDF11)  cell senescence  cell cycle

中文摘要:

      目的 基于细胞周期机制探索生长分化因子11(growth differentiation factor 11,GDF11)与细胞衰老的关系.方法 应用D-半乳糖构建人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)衰老模型,应用siRNA构建GDF11低表达模型,观察GDF 11对HUVEC的β-半乳糖苷酶(senescence associated β-galactosidase,SA-β-Gal)的活性及周期相关P300、C-myc和TGFβ表达的影响.实验分组与处理:(1)对照组:HUVEC不作处理;(2)模型组:用10 g/L的D-半乳糖成功构建HUVEC衰老模型;(3)siRNA-NC组:siRNA的内部对照组不干预GDF11表达,转染24h后再应用D-半乳糖诱导24 h后进行指标检测;(4) siRNA组:与siRNA-NC组步骤相同但可降低GDF 11的表达;(5) GDF11前处理组:先用GDF11处理24 h后再用D-半乳糖诱导24 h;(6) GDF11后处理组:先应用D-半乳糖诱导24 h后再用GDF 11处理24 h.结果 10 g/L D-半乳糖可成功诱导HUVEC细胞衰老模型;10 ng/μLGDF11可促进P300和C-myc表达;GDF11可减少衰老相关SA-β-Gal的表达;GDF11可能经P300/C-myc通路干预细胞周期进而干预细胞衰老进程;GDF11处理后可明显降低SA-β-Gal活性,且GDF11前处理,细胞抵抗D-半乳糖诱导衰老的能力越好.结论 外源性GDF11参与调节HUVEC细胞周期并延缓细胞衰老.

英文摘要:

      Objective To explore the relationship between growth differentiation factor 11 (GDF 11) and cell senescence based on cell cycle mechanism.Methods D-galactose was applied in building up the senile model of human umbilical vein endothelial cells (HUVECs) while siRNA was applied in building up the low-expression model of GDF11;observation was made to the effect of GDF11 on the expressions of P300,C-myc and TGFβ correlated to the activity of senescence associated β-galactosidase (SA-β-Gal) and the cell cycle in HUVECs;6 groups were set up:(1) control group (with no treatment to HUVECs),(2) model group (HUVEC senile model built up with 10 g/L of D-galactose),(3) siRNA-NC group (control of siRNA with no effect on the expression of GDF11),(4) siRNA group (same procedures with siRNA-NC group but with GDF11 expression decreasing),(5) GDF11 pretreatment group (treated with GDF11 for 24 hours and then induced by D-galactose for 24 hours) and GDF11 post-treatment group (induced by D-galactose for 24 hours and then treated by GDF11 for 24 hours).Results HUVEC senile model was built up successfully with the induction of 10g/L of D-galactose;10ng/μL of GDF11 promoted the expression of P300 and C-myc;the expression of SA-β-Gal related to senescence was reduced by GDF11;GDF11 interfered with cell cycle via the P300/C-myc pathway and then interfered with cell senescence;the activity of SA-β-Gal in GDF11 post-treatment group decreased obviously while the cell ability against senescence induced by D-galactose was stronger in GDF11 pretreatment group.Conclusions Exogenous GDF11 involves in regulating the cycle of HUVECs and in delaying the senescent process of cells.