呼吸道合胞病毒对上呼吸道感染儿童呼吸道上皮细胞自噬和mTOR信号通路基因的影响

Regulation of Autophagy and mTOR Signaling Pathway Genes in Respiratory Epithelial Regulation of Autophagy and mTOR Signaling Pathway Genes in Respiratory Epithelial Cells of Children with Upper Respiratory Tract Infection by Respiratory Syncytial Virus

目的 探讨呼吸道合胞病毒(RSV)对上呼吸道感染儿童呼吸道上皮细胞自噬和mTOR信号通路相关基因的影响。方法 选择2022年11月至2023年12月唐山职业技术学院附属医院收治的145例上呼吸道感染儿童为研究对象,同时纳入同期体检的年龄和性别匹配的健康儿童(n=10)为健康对照组。采集所有儿童鼻咽分泌物标本(NPA),利用RT-PCR进行RSV筛查,通过G(黏附)基因扩增测定RSV毒株的分子特征并进行系统发育分析。利用qRT-PCR检测25例RSV阳性NPA样本(RSV组)和10例健康对照NPA样本(健康对照组)的呼吸道上皮细胞自噬相关基因(BECN1、ATG3、ATG5、NPC1与GABARAP)和mTOR信号通路相关基因(AKT1、mTOR、PDPK1、RICTOR与TSC1)的表达水平。结果 在145份上呼吸道感染儿童NPA样本中,69份(47.6%)为RSV阳性。对31株有代表性的RSV进行循环基因型检测,经BLAST分析,31株RSV均为RSV-A型。通过靶向G基因的胞外域部分,系统发育分析发现31株RSV-A聚类为具有72 bp核苷酸重复的ON1基因型。此外,毒株分别属于谱系1(51.6%),其次是谱系3(29%)和谱系2(19.4%)。RSV组呼吸道上皮细胞中ATG3和NPC1 mRNA表达水平显著高于健康对照组(均P<0.001)。RSV组呼吸道上皮细胞中AKT1、mTOR和TSC1 mRNA表达水平显著低于健康对照组(均P<0.001)。结论 RSV感染可以提高上呼吸道感染儿童呼吸道上皮细胞中自噬相关基因(ATG3和NPC1)的表达,抑制mTOR信号通路相关基因(AKT1、mTOR和TSC1)的表达,进而逃避宿主免疫系统,以实现其在宿主内的生存方式。

Objective To investigate the effects of respiratory syncytial virus(RSV)on autophagy and mTOR signaling pathway-related genes in respiratory epithelial cells of children with upper respiratory tract infection.Methods A total of 145 children with upper respiratory tract infection admitted to Tangshan Vocational and Technical College Affiliated Hospital from November 2022 to December 2023 were selected as the study objects.At the same time,age-and sex-matched healthy children(n=10)who underwent physical examination in our hospital during the same period were included as the healthy control group.Nasopharyngeal aspirate(NPA)were collected from all children.RSV was screened by RT-PCR,and the molecular characteristics of RSV strains were determined by G(attachment)gene amplification and phylogenetic analysis was performed.The expression levels of respiratory epithelial cell autophagy-related genes(BECN1,ATG3,ATG5,NPC1 and GABARAP)and mTOR signaling pathway-related genes(AKT1,mTOR,PDPK1,RICTOR,and TSC1)were detected using qRT-PCR in 25 RSV-positive NPA samples(RSV Group)and 10 healthy control NPA samples(Healthy Control Group).Results Of 145 NPA samples from children with upper respiratory tract infection,69(47.6%)were RSV positive.The circulating genotypes of 31 representative RSV strains were detected and BLAST analysis showed that all 31 RSV strains were RSV-A type.By targeting the ectodomain portion of the G gene,phylogenetic analysis showed that 31 strains of RSV-A were clustered into ON1 genotypes with 72 bp nucleotide repeats.In addition,the strains belonged to lineage 1(51.6%),followed by lineage 3(29%)and lineage 2(19.4%).The mRNA expression levels of ATG3 and NPC1 in respiratory epithelial cells of the RSV group were significantly higher than those of the healthy control group(P<0.001).The mRNA expression levels of AKT1,mTOR and TSC1 in respiratory epithelial cells of the RSV group were significantly lower than those of the healthy control group(P<0.001).Conclusion RSV infection can increase the expression of autophagy-related genes(ATG3 and NPC1)in respiratory epithelial cells of children with upper respiratory tract infection,inhibit the expression of mTOR signaling pathway-related genes(AKT1,mTOR and TSC1),and then evade the host immune system to achieve their survival mode in the host.