哮喘发作与细胞骨架重建相关基因的差异表达

Differentially expression of genes associated with cytoskeletal remodeling of peripheral blood eosinophils during asthmatic attack

目的:研究外周血嗜酸性粒细胞(EOS)在哮喘发作时与细胞骨架相关基因的差异表达.方法:应用SuperSMARTcDNAsynthesis技术从总RNA合成足量的双链cD-NA.经液相杂交消减两组共同序列,以抑制性PCR方式扩增差异表达序列.构建上调与下调cDNA文库,菌落PCR扩增差异片段,应用差异筛选技术进一步鉴定差异表达基因并测序,结果通过核酸数据库进行同源性比较.结果:构建了高效的上调与下调消减cDNA文库,经鉴定及同源性分析有6个与细胞骨架重建有关,包括上调基因slingshot磷酸酶(SSH-2L),蛋白磷酸酶1β亚型(PP1Cβ),无功能糖基转移酶样蛋白(DOCK-8),蛋白酪氨酸磷酸酶非受体型6(PTPN6)与非受体型12(PTPN12);下调基因,肌凝蛋白调节轻链结合蛋白(MIR).结论:这些骨架重建相关基因的差异表达可能参与了哮喘发作,进一步进行功能研究可能为哮喘治疗提供新的作用靶点.

AIM： To determine the differentially expressed genes associated with cytoskeletal remodeling of peripheral blood eosinophils （ EOS ） in patients during asthmatic attack. METHODS： Double-stranded cDNA was amplified from total RNA of EOS at the time of asthmatic attack and after improvement respectively by Super SMART cDNA Synthesis techology, Suppression subtractive hybridization （SSH） and PCR- select differential screening technology were used to detect differentially expressed genes and then differentially expressed genes were sequenced. Homology analysis was conducted by comparing these gene sequences based on GenBank database. RESULTS： Highefficiently up-regulated and down-regulated subtractive cDNA libraries were constructed successfully; differential screening and homology analysis identified 6 differentially expressed genes associated with cyteskeletal remodeling, including slingshot 2L（ SSH- 2L）, PP1 catalytic subunit, beta isoform（PP1Cβ）, dedicator of cytokinesis 8 （ DOCK-8 ） , Homo sapiens protein tyrosine phosphatase non-receptor type 6 and 12（ PTPN6, PTPN12）, myosin regulatory light chain interacting protein （MIR）. CONCLUSION： The differentially expressed genes associated with cytoskeletal remodeling may participate in exacerbation of asthma. Further study on the genes＇ function may provide new gene target for asthma treatment.