基础医学与临床2026，Vol.46Issue(4)：498-503,6.DOI:10.16352/j.issn.1001-6325.2026.04.0498
构建LILRA2过表达慢病毒载体及建立THP-1-LILRA2稳定转染细胞株
Construction of LILRA2-overexpressing lentiviral vector and establishment of THP-1-LILRA2 stable transfected cell strain
摘要
Abstract
Objective To construct a recombinant lentiviral vector overexpressing leukocyte immunoglobulin-like receptor subfamily A member 2(LILRA2),establish a stable LILRA2-overexpressing human myeloid leukemia mononuclear cell strain(THP-1),and preliminarily investigate the impact of LILRA2 on the expression of pro-in-flammatory cytokines.Methods The pLV-SFFV-MCS-EF1-ZsGreen1-T2A-Puro backbone vector was double-diges-ted with BamH Ⅰ/Xba Ⅰ to construct the LILRA2-overexpressing recombinant lentiviral vector.After lentiviral packaging,concentration,and titer determination,the optimal multiplicity of infection(MOI=30)and puromycin screening concentration(2 μg/mL)for THP-1 cells were determined.Following lentiviral infection,puromycin se-lection was performed to obtain stable transfectants.LILRA2 expression was verified by RT-qPCR and Western blot.TNF-α and IL-1β expression was determined by RT-qPCR.Results Sequencing confirmed the successful construc-tion of the LILRA2-overexpressing recombinant lentiviral vector.The fluorescence-positive rate in stably transfected THP-1 cells exceeded 90％.LILRA2 mRNA(P<0.001)and protein(P<0.05)expression levels in the THP-1-LILRA2 group were significantly higher than those in the THP-1-NC negative control group.TNF-α(P<0.05)and IL-1β(P<0.05)mRNA levels in the THP-1-LILRA2 group were significantly higher than those in the THP-1-NC negative control group.Conclusions The LILRA2-overexpressing lentiviral vector was successfully constructed and a stable LILRA2-overexpressing THP-1 cell strain was established.LILRA2 overexpression upregulates the expres-sion of pro-inflammatory cytokines in monocytes.关键词
白细胞免疫球蛋白样受体亚家族A成员2(LILRA2)/THP-1细胞/慢病毒载体/促炎性因子Key words
leukocyte immunoglobulin like receptor subfamily A member 2(LILRA2)/THP-1 cells/lentiviral vector/pro-inflamma-tory cytokines分类
医药卫生引用本文复制引用
黄高翔,魏冬梅,杨耀,郭夏宁,周凯,栾晶..构建LILRA2过表达慢病毒载体及建立THP-1-LILRA2稳定转染细胞株[J].基础医学与临床,2026,46(4):498-503,6.基金项目
国家自然科学基金(8230060948,32260239) （8230060948,32260239）
陕西省重点研发计划(2024SF-YBXM-347) （2024SF-YBXM-347）
