摘要
目的:探究长链非编码RNA Gm26688对巨噬细胞RAW264.7的免疫调控功能,使用中国大鲵(CGS)胶原蛋白肽处理LPS诱导的巨噬细胞,探究Gm26688在CGS骨胶原蛋白肽调控炎症反应中的作用。方法:RT-PCR克隆Gm26688基因,RNA-FISH分析Gm26688在巨噬细胞中的定位;qRT-PCR检测LPS和CGS骨胶原蛋白肽处理后Gm26688和炎症因子表达;使用小干扰RNA(siRNA)沉默Gm26688表达,qRT-PCR检测Gm26688干扰后LPS和CGS骨胶原蛋白肽对炎症因子表达的影响;Western blot检测NF-κB p65表达及其磷酸化水平。结果:LPS显著促进Gm26688表达。RT-PCR扩增得到Gm26688序列,RNA-FISH显示Gm26688主要位于细胞核。CGS骨胶原蛋白肽抑制LPS诱导的Gm26688表达,且<3 kD的CGS骨胶原蛋白肽对Gm26688的抑制效果最显著。CGS骨胶原蛋白肽(<3 kD)显著抑制LPS诱导的炎症因子IL-6、IL-1β和TNF-α表达。Gm26688干扰显著下调LPS或LPS+CGS骨胶原蛋白肽刺激的RAW264.7细胞中炎症因子IL-6和TNF-α表达。Western blot结果显示,CGS骨胶原蛋白肽通过Gm26688抑制NF-κB p65磷酸化水平。结论:CGS骨胶原蛋白肽通过Gm26688/NF-κB路径抑制LPS诱导的RAW264.7细胞炎症。
Objective:To explore immunoregulatory function of long non-coding RNA Gm26688 on macrophages RAW264.7,macrophages stimulated by LPS were treated with Chinese giant salamander(CGS)collagen peptides to explore role of Gm26688 in mechanism of CGS collagen peptides in regulating inflammatory reflection.Methods:Gm26688 sequence was cloned by RT-PCR,Gm26688 localization in macrophage was analyzed by RNA-FISH;qRT-PCR was performed to detect expressions of Gm26688 and inflammatory factors after LPS and CGS bone collagen peptides treatment;expression of Gm26688 was silenced by small interfering RNA(siRNA),and qRT-PCR was used to detect effects of LPS and CGS bone collagen peptides on expression of inflammatory factors after Gm26688 interference;Western blot was used to detect protein expression of NF-κB p65 and its phosphorylation level.Results:LPS significantly promoted expression of Gm26688.RT-PCR amplified Gm26688 sequence,and RNA-FISH showed that Gm26688 was mainly expressed in nucleus.CGS bone collagen peptides inhibited expression of Gm26688 induced by LPS,and<3 kD CGS bone collagen peptides had the most significant inhibitory effect on Gm26688.CGS bone collagen peptides(<3 kD)significantly inhibited LPS-induced expressions of inflammatory factors IL-6,TNF-αand IL-1β.Interference of Gm26688 significantly down-regulated expressions of inflammatory factors IL-6 and TNF-αin LPS or LPS+CGS bone collagen peptides-stimulated RAW264.7 cells.Western blot results showed that CGS bone collagen peptides down-regulated level of NF-κB p65 phosphorylation through Gm26688.Conclu-sion:CGS bone collagen peptides inhibit LPS-induced inflammation in RAW264.7 cells via Gm26688/NF-κB pathway.
作者
苏淑贤
黎琴容
商文聪
SU Shuxian;LI Qinrong;SHANG Wencong(College of Biological and Environmental Engineering,Guiyang University,Guizhou Key Laboratory for Rare Animal and Economic Insect of the Mountainous Region,Guiyang 550005,China)
出处
《中国免疫学杂志》
北大核心
2025年第8期1920-1927,共8页
Chinese Journal of Immunology
基金
贵州省基础研究计划项目(黔科合基础-ZK[2022]一般011)
贵州省普通高等学校青年科技人才成长项目(黔教合KY字[2020]091)
贵阳市科技局贵阳学院专项资金(GYU-KY-[2021])
贵阳学院硕士研究生科研基金项目(GYU-YJS[2021]-67)。
