Abstract: Objective To explore the expression of tumor necrosis factor receptor superfamily 21 (TNFRSF21) in hepatocellular carcinoma (HCC), related biological function and its relationships with prognosis and immune system. Methods The data sets of HCC cell lines were obtained from Gene Expression Omnibus (GEO) database to screen the common differentially expressed genes. The data of HCC patients were obtained from The Cancer Genome Atlas (TCGA) database, the expression of TNFRSF21 of HCC and paracancerous tissues was compared, and the relationship between TNFRSF21 and overall survival (OS), as well as clinicopathological characteristics was analyed. Moreover, expression level of TNFRSF21 in 67 patients with HCC from July 2009 to October 2010 were detected by real-time quantitative PCR (qPCR).The relationship of expression level of TNFRSF21 with OS of HCC patients was analyzed. The TNFRSF21 proteinprotein interaction network was constructed by STRING, and the biological functions and tumor-related pathways of TNFRSF21 were enriched by Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG). TIMER and TISIDB were used to analyze the correlation between TNFRSF21 and immune infiltrating cells in HCC. ResultsCommon differentially expressed genes TNFRSF21, CROT, APPBP2 and CPD in HepG2 and Huh7 cell lines with CSN5 knockdown, as well as HepG2 and Huh7 cell lines with COP1 knockdown were screened based on GEO database. Based on TCGA database, the median mRNA of TNFRSF21 was 3.125 in paracancerous tissues and 3.953 in 374 HCC tissues (P<0.001). TNFRSF21 mRNA in HCC tissues was 1.77±1.60, which was higher than 1.07±0.83 in paracancerous tissues (P=0.002). Survival analysis showed that only TNFRSF21 gene expression level was correlated with OS (P=0.030). In 67 HCC patients, the median OS in the high-expression group was unreached, superior to 35.0 months in the low-expression group (P=0.039). According to TCGA database, 374 HCC patients were divided into high and low expression groups based on the median TNFRSF21 expression of 2.678.It was found that TNFRSF21 expression level was significantly correlated with serum AFP level (P<0.001) and vascular invasion (P=0.029) of HCC patients. According to STRING and GO&KEGG enrichment analysis, TNFRSF21 may be involved in NF-κB and JNK signaling pathways through APP and TARDD, thus regulating the occurrence and development of tumors. TIMER analysis showed that the high expression of TNFRSF21 was associated with B cells (r=0.299,P<0.001), CD8+T cells (r=0.242, P<0.001), CD4+T cells (r=0.417, P<0.001), macrophages (r=0.527, P<0.001), neutrophils (r=0.389, P<0.001) and dendritic cells(r=0.363, P<0.001). TISIDB analysis showed that the expression of TNFRSF21 was positively correlated with the abundance of activated dendritic cells (r=0.271, P<0.001), follicular helper T cells (r=0.333, P<0.001), mast cells (r=0.270, P<0.001) and CD4+ central memory T cells (r=0.393, P<0.001). Conclusion TNFRSF21 gene is up-regulated in HCC, and related to poor prognosis. TNFRSF21 may promote the occurrence and development of HCC by inducing endothelial cell necrosis and tumor microenvironment through tumor immune infiltrating cells. TNFRSF21 can be used as a molecular marker for prognosis in HCC.

Key words: Hepatocellular carcinoma；Tumor necrosis factor receptor superfamily 21, Gene expression；Tumor immune infiltrating cells；Prognosis