目的探讨人脑原代微血管周细胞(HBVPs)和人肾脏原代微血管周细胞(HRVPs)差异表达基因及其对脑微血管周细胞迁移和增殖功能的影响。方法利用RNA-seq检测对HBVPs和HRVPs进行转录组测序,qPCR及Western blot检测HAND2 mRNA及蛋白水平,构建HAND2基因的sh-RNA下调脑微血管周细胞HAND2表达,分别利用CCK-8检测HBVPs增殖活性和Transwell小室迁移实验检测迁移能力。结果 RNA-seq检测结果显示胚胎期HBVPs相对HRVPs有1 906个高表达基因,进一步的聚类分析发现这些基因可显著地聚类为4类功能富集的亚群;与血管发育密切相关的第Ⅱ群中HAND2在HBVPs中显著高表达(lg2FoldChange=11.93,P<0.05)。此外,qPCR和/或Western blot检测发现在小鼠胚胎第10.5天HAND2表达量较高,在第14.5天时最高,出生后几乎不表达(P<0.05),并且HAND2主要表达在HBVPs(P<0.05)。当sh-RNA下调HBVPs的HAND2表达后,发现其在24 h时迁移能力增强(P<0.05),48 h和72 h时增殖活性显著增加(P<0.05)。结论 HAND2对胚胎脑微血管发育和成熟期周细胞的增殖和迁移能力具有重要的调控作用。
Compared with renal microvascular pericytes, cerebral microvascular pericytes play a unique role in promoting the development and maturation of microvascular barrier structure, which play an important role in inhibiting the damage of peripheral blood immune inflammatory cells and inflammatory molecules on neurons and maintaining the stability of central nervous system. Thus, this study was designed to investigate the gene differentially expressed in human brain primary microvascular pericytes(HBVPs) and human renal microvascular pericytes(HRVPs) and its effect on the cerebral microvascular pericytes migration and proliferation. Genes differentially expressed in HBVPs and HRVPs were measured by RNA-seq;the expression of HAND2 mRNA and protein were measured by qPCR and Western blot. Down-regulation of HAND2 in cerebral microvascular pericytes was conducted by HAND2 sh-RNA. RNA-seq data indicated that there were 1 906 genes with high expression in embryonic HBVPs compared with HRVPs,and cluster analysis indicated that there were four gene functional enrichment subgroups. HAND2 in the second subgroup(closely related to vascular development)was significantly higher expressed in HBVPs compared with HRVPs(lg2FoldChange= 11.93, P<0.05). In addition, HAND2 in mice started to increase at E10.5, peaked at E14.5, hardly expressed after birth(P<0.05), and mainly expressed in HBVPs(P<0.05). After down-regulation of HAND2 expression, the proliferation activity of HBVPs increased at 48 h and 72 h(P<0.05), and the migration ability increased at 24 h(P<0.05). Taken together, HAND2 regulates pericytes proliferation and migration in the stages of embryonic cerebral microvascular development and maturation.