背景:皮肤癣菌病是人类和动物的一种顽固性浅部真菌病,主要由毛癣菌引起(T。植叶植物),全球患病率约为20%。角质形成细胞是皮肤免疫中最丰富的参与者,它们还在对抗T.mentagrophytes的一线防御中发挥作用。然而,尚未报道基于整个转录组的角质形成细胞对T.mentagrosphytes感染的反应。
目标:这里,我们使用全转录组测序技术,系统地分析了受T.mentagrosphytes感染的角质形成细胞的变化。
方法:1×105分生孢子/mL感染后角质形成细胞的表型变化。通过光学显微镜观察到植叶植物,扫描电子显微镜,透射电镜和末端脱氧核苷酸转移酶dUTP缺口末端标记。RNA测序(RNA-seq),小RNA-seq技术和相关的生物信息学方法被用来系统地分析角质形成细胞在T.
结果:我们发现T.膜损伤,不规则细胞器的形成和角质形成细胞的凋亡。总共204个差异表达(DE)环状RNA(circRNAs),868DE长链非编码RNA(lncRNAs),在未感染的角质形成细胞和T.mentagrosphytes感染的角质形成细胞之间鉴定了2973个DEmRNA和209个DE微RNA(miRNA)。通过定量实时聚合酶链反应(qRT-PCR)验证所选RNA的表达水平。功能富集分析表明,DEcircRNAs的亲本基因与细胞反应有关,细胞死亡和皮肤屏障的建立。miRNA靶向的基因参与调节免疫应答的启动。基于circRNAs的表达水平,lncRNAs,mRNA和miRNA,circRNA-miRNA-mRNA竞争内源性(ceRNA)网络由159个DEmiRNA组成,141DEcircRNAs和2307DEmRNAs,和由790个DElncRNA组成的lncRNA-miRNA-mRNAceRNA网络,190个DEmiRNAs和2663个DEmRNAs被构建。使用qRT-PCR验证了两个选择的ceRNA网络的可靠性。进一步的功能富集分析显示,在ceRNA网络中与circRNAs和lncRNAs相互作用的DEmRNAs主要参与真菌识别,炎症,先天免疫反应和角质形成细胞的死亡。
结论:我们的研究结果可能提供了新的证据,说明了T.这对于确定皮肤癣菌病治疗的新治疗靶点至关重要。
BACKGROUND: Dermatophytosis is an intractable superficial mycosis in humans and animals mainly caused by Trichophyton mentagrophytes (T. mentagrophytes), with a global prevalence of about 20%. Keratinocytes are the most abundant participants in skin immunity, and they also play a role in the first-line defence against T. mentagrophytes. However, no studies of keratinocyte responses against T. mentagrophytes infection based on the whole transcriptome have been reported.
OBJECTIVE: Here, we systematically analysed changes in keratinocytes infected with T. mentagrophytes using whole transcriptome sequencing technology.
METHODS: The phenotypic changes in keratinocytes after infection with 1 × 105 conidia/mL T. mentagrophytes were observed by light microscopy, scanning electron microscopy, transmission electron microscopy and terminal deoxynucleotidyl transferase dUTP nick end labeling. RNA-sequencing (RNA-seq), small RNA-seq technology and related bioinformatics methods were used to systematically analyse the whole transcriptome changes in keratinocytes upon T. mentagrophytes stimulation.
RESULTS: We found that T. mentagrophytes infection caused morphological changes, membrane damage, the formation of irregular organelles and keratinocyte apoptosis. A total of 204 differentially expressed (DE) circular RNAs (circRNAs), 868 DE long noncoding RNAs (lncRNAs), 2973 DE mRNAs and 209 DE micro RNAs (miRNAs) were identified between noninfected and T. mentagrophytes-infected keratinocytes. The expression level of selected RNAs was validated by quantitative real-time polymerase chain reaction (qRT-PCR). Functional enrichment analysis revealed that the parental genes of DE circRNAs were related to cell response, cell death and establishment of the skin barrier. Genes targeted by miRNA were involved in regulating the initiation of the immune response. Based on the expression level of circRNAs, lncRNAs, mRNAs and miRNAs, circRNA-miRNA-mRNA competing endogenous (ceRNA) networks comprised of 159 DE miRNAs, 141 DE circRNAs and 2307 DE mRNAs, and lncRNA-miRNA-mRNA ceRNA networks comprised of 790 DE lncRNAs, 190 DE miRNAs and 2663 DE mRNAs were constructed. The reliability of two selected ceRNA networks was verified using qRT-PCR. Further functional enrichment analysis revealed that the DE mRNAs interacting with circRNAs and lncRNAs in the ceRNA network mainly participated in fungal recognition, inflammation, the innate immune response and the death of keratinocytes.
CONCLUSIONS: Our findings might provide new evidence on the pathogenesis of T. mentagrophytes-induced dermatophytosis, which is essential for identifying new therapeutic targets for dermatophytosis treatment.