血管内皮细胞生长因子促进角质形成细胞中CD34的表达

Vascular endothelial growth factor promotes the expression of vascular endothelial cell factor in hair follicle keratinocytes

目的观察血管内皮细胞生长因子(VEGF)是否促进角质形成细胞(KCs)中血管内皮细胞因子CD34的表达。方法 2020年10月至2021年7月,15只C57BL/6小鼠背部制作全皮层缺损创面,给予及时护理,分别取正常组织及第1、3、5、7天创面组织进行蛋白免疫印迹法(Western blot)检测VEGF表达。细胞实验分为KCs组和KCs+VEGF组,采用实时荧光定量聚合酶链反应(RT-qPCR)检测CD34信使RNA(mRNA)水平,Western blot检测CD34、Delta样配体4(DLL4)、肝配蛋白B2 (Ephrin B2)蛋白水平的表达;加入VEGF组和VEGF+VEGF抑制剂L-685458组采用Western blot检测CD34蛋白表达。应用图像处理软件Image J和统计软件GraphPad Prism6分析,两组间比较采用t检验,多组间采用单因素方差分析。结果随着时间的推移,创面组织中VEGF蛋白表达水平呈动态升高(N=0.344±0.037,d1=0.158±0.019,d3=0.298±0.006,d5=0.509±0.066,d7=1.056±0.172,F=172.8,P<0.01)。加入VEGF后,角质形成细胞CD34 mRNA表达增加(KCs=1.33±0.36,KCs+VEGF=5.04±0.51,t=10.25,P<0.01),CD34、DLL4、Ephrin B2蛋白表达水平升高(KCs=0.268±0.026,KCs+VEGF=0.693±0.005,tCD34=27.78,PCD34<0.01;KCs=0.172±0.063,KCs+VEGF=0.609±0.171,tDLL4=4.785,PDLL4<0.05;KCs=0.293±0.031,KCs+VEGF=0.835±0.049,tEphrin B2=16.26,PEphrin B2<0.05),而加入VEGF抑制剂L-685458后,角质形成细胞中CD34表达低于VEGF组(VEGF=0.777±0.053,VEGF+L-685458=0.263±0.053,t=11.83,P<0.01)。结论 VEGF促进角质形成细胞中血管内皮细胞相关因子CD34、DLL4、Ephrin B2的表达。

Objective To investigate whether vascular endothelial growth factor(VEGF)promotes the expression of vascular endothelial cell-related factor CD34 in keratinocytes.Methods From October 2020 to July 2021,15 C57BL/6 mice(purchased from Three Gorges University)were used to make full-thickness defect wounds on the back,and timely care was given.The normal tissues and the wound tissues on the 1,3,5 and 7 days were taken for Western blotting to detect the expression of VEGF.Cell experiments were divided into KCs group and KCs+VEGF group.The level of CD34 messenger RNA(mRNA)was detected by real-time quantitative polymerase chain reaction(RT-qPCR).The protein expression levels of CD34,Delta-like ligand 4(DLL4)and liver ligand B2(Ephrin B2)were detected by Western blotting.The protein expression of CD34 was detected by Western blotting in VEGF group and VEGF+VEGF inhibitor L-685458 group.The image processing software Image J and statistical software GraphPad Prism 6 were used for analysis.The t test was used for comparison between the two groups,and one-way ANOVA was used for multiple groups.Results With the passage of time,the protein expression level of VEGF in the wound tissue increased dynamically(N=0.344±0.037,d1=0.158±0.019,d3=0.298±0.006,d5=0.509±0.066,d7=1.056±0.172,F=172.8,P<0.01).After VEGF was added,the mRNA expression of CD34 in keratinocytes increased(KCs=1.33±0.36,KCs+VEGF=5.04±0.51,t=10.25,P<0.01),and the protein expression levels of CD34,DLL4 and Ephrin B2 increased(KCs=0.268±0.026,KCs+VEGF=0.693±0.005,tCD34=27.78,PCD34<0.01;KCs=0.172±0.063,KCs+VEGF=0.609±0.171,tDLL4=4.785,PDLL4<0.05;KCs=0.293±0.031,KCs+VEGF=0.835±0.049,tEphrin B2=16.26,PEphrin B2<0.05).And after adding VEGF inhibitor,the expression of CD34 in keratinocytes was lower than that in VEGF group(VEGF=0.777±0.053,VEGF+L-685458=0.263±0.053,t=11.83,P<0.01).Conclusion VEGF promotes the expression of vascular endothelial cell-related factor CD34 in keratinocytes.