■肝纤维化(HF)的分子机制,与自噬密切相关,仍然不清楚。本研究旨在探讨HF的自噬特征。
■下载了基因表达谱(GSE6764、GSE49541和GSE84044),归一化,并合并。自噬相关的差异表达基因(ARDEGs)使用limmaR包和Wilcoxon秩和检验确定,然后通过GO进行分析。KEGG,GSEA和GSVA。免疫细胞的浸润,分析健康对照(HC)和HF的分子亚型和免疫类型。机器学习有两种方法,其中,获得了核心基因。构建肝纤维化体内外模型,验证核心基因和相应免疫细胞的表达。
■共鉴定出69个ARDEGs。系列功能聚类分析显示ARDEGs显著富集了自噬和免疫功能。活化的CD4T细胞,CD56bright自然杀伤细胞,CD56dim自然杀伤细胞,嗜酸性粒细胞,巨噬细胞,肥大细胞,中性粒细胞,和17型T辅助(Th17)细胞在HC和HF组之间的浸润差异显着。在ARDEGs中,确定了三个核心基因,ATG5、RB1CC1和PARK2。在三个核心基因的不同表达水平上观察到免疫细胞的浸润有相当大的变化,其中RB1CC1的表达与巨噬细胞浸润显著相关,Th17细胞,自然杀伤细胞和CD56dim自然杀伤细胞。在小鼠肝纤维化实验中,HF组ATG5、RB1CC1和PARK2水平高于HC组。与HC组相比,HF组F4/80、IL-17和CD56阳性面积较低,表明巨噬细胞表达减少,Th17细胞,自然杀伤细胞和CD56dim自然杀伤细胞。同时,发现敲低RB1CC1可以抑制肝星状细胞的活化并减轻肝纤维化。
■ATG5、RB1CC1和PARK2是有前途的自噬相关治疗HF的生物标志物。这是第一个在HF中鉴定RB1CC1的研究,这可能通过调节巨噬细胞促进肝纤维化的进展,Th17细胞,自然杀伤细胞和CD56dim自然杀伤细胞。
The molecular mechanisms of hepatic fibrosis (HF), closely related to autophagy, remain unclear. This study aimed to investigate autophagy characteristics in HF.
Gene expression profiles (GSE6764, GSE49541 and GSE84044) were downloaded, normalized, and merged. Autophagy-related differentially expressed genes (ARDEGs) were determined using the limma R package and the Wilcoxon rank sum test and then analyzed by GO, KEGG, GSEA and GSVA. The infiltration of immune cells, molecular subtypes and immune types of healthy control (HC) and HF were analyzed. Machine learning was carried out with two methods, by which, core genes were obtained. Models of liver fibrosis in vivo and in vitro were constructed to verify the expression of core genes and corresponding immune cells.
A total of 69 ARDEGs were identified. Series functional cluster analysis showed that ARDEGs were significantly enriched in autophagy and immunity. Activated CD4 T cells, CD56bright natural killer cells, CD56dim natural killer cells, eosinophils, macrophages, mast cells, neutrophils, and type 17 T helper (Th17) cells showed significant differences in infiltration between HC and HF groups. Among ARDEGs, three core genes were identified, that were ATG5, RB1CC1, and
PARK2. Considerable changes in the infiltration of immune cells were observed at different expression levels of the three core genes, among which the expression of RB1CC1 was significantly associated with the infiltration of macrophage, Th17 cell, natural killer cell and CD56dim natural killer cell. In the mouse liver fibrosis experiment, ATG5, RB1CC1, and
PARK2 were at higher levels in HF group than those in HC group. Compared with HC group, HF group showed low positive area in F4/80, IL-17 and CD56, indicating decreased expression of macrophage, Th17 cell, natural killer cell and CD56dim natural killer cell. Meanwhile, knocking down RB1CC1 was found to inhibit the activation of hepatic stellate cells and alleviate liver fibrosis.
ATG5, RB1CC1, and
PARK2 are promising autophagy-related therapeutic biomarkers for HF. This is the first study to identify RB1CC1 in HF, which may promote the progression of liver fibrosis by regulating macrophage, Th17 cell, natural killer cell and CD56dim natural killer cell.