Differential expression of P2rx7 splice variants in microglial cells under different polarization states

AN Xiao-meng; ZHOU Fang-yu; ZHANG Ji; XU Xue-bing

doi:10.13820/j.cnki.gdyx.20242028

Abstract：Objective P2X7 receptor(P2rx7)alternative splicing affects exon expression of the P2rx7 gene and is a primary regulator of P2rx7 function.This study aims to investigate the differential expression of various P2rx7 splice variants in microglial cells under M1/M2 polarization states.Methods BV2 immortalized murine microglial cells were cultured and treated with lipopolysaccharide(LPS,100 ng/mL)or interleukin-4(IL-4,40 ng/mL)for48 hours to in-duce M1 or M2 polarization,respectively.Flow cytometry confirmed polarization states,and real-time quantitative PCR was employed to quantify expression levels of five P2rx7 splice variants(P2rx7-v1,-v2,-v3,-v4,and-vk).Results All P2rx7 transcripts were detectable at the mRNA level in BV2 cells under standard culture conditions,with P2rx7-v1 as the predominant variant.Other splice variants,including P2rx7-v2,-v3,-v4,and-vk,were present at relatively low levels or nearly undetectable.Following LPS treatment,expression of the M2 marker CD206 was signifi-cantly reduced(P＜0.01),while the M1 marker CD86 showed a non-significant increase(13.0％vs.78.8％,P=0.054).Notably,expression levels of all P2rx7 variants increased significantly(P＜0.01),except for P2rx7-v4,which exhibited a relatively modest increase(15.8％,P＜0.05).In contrast,IL-4 stimulation led to a significant upregula-tion of CD206(280.0％,P＜0.01)and a reduction in CD86(47.6％,P=0.035).Further analysis revealed de-creased expression of P2rx7-v1 and-v4(64.9％vs.52.9％,P＜0.01),while P2rx7-v3 expression showed a slight increase(61.7％,P＜0.01).Conclusion The five P2rx7 splice variants exhibit differential expression under M1/M2 polarization states in microglial cells.These findings underscore the need for further research on the structural and func-tional impacts of P2rx7 splice variants on microglial polarization.