RBP7 Regulated by EBF1 Affects Th2 Cells and the Oocyte Meiosis Pathway in Bone Metastases of Bladder Urothelial Carcinoma

Abstract

Background: Bladder urothelial carcinoma (BLCA) is a malignancy with a high incidence worldwide. One-third of patients may experience aggressive progression later on, and 70% of patients who have undergone surgical intervention will still suffer from metastasis.

Materials and methods: RNA sequencing profiles of BLCA samples were obtained from The Cancer Genome Atlas (TCGA) database. Differential expression and univariate Cox regression analyses were performed to identify prognosis-related differentially expressed immune genes (DEIGs). Subsequently, a proportional hazards model of DEIGs was then constructed by univariate regression analysis. Differential expression and correlation analyses, CIBERSORT, Single Sample Gene Set Enrichment Analysis (ssGSEA), GSVA were conducted on transcription factors (TFs), immune cells/pathways and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. The regulation network was then constructed. Eventually, ATAC-seq, ChIP-seq, scRNA-seq, and multiple online databases were employed for further validation.

Results: A proportional hazards model of 31 DEIGs was constructed and risk score was calculated and proven to be a independent prognostic factor. Then 5 immune genes were characterized to be significantly correlated with bone metastasis, stage and TF expression simultaneously. 4 TFs were identified to be significantly correlated with prognosis and RBP7 expression. 5 immune cells/pathways were revealed to be significantly correlated with RBP7 expression. Only 1 KEGG pathway was identified to be significant in Gene Set Enrichment Analysis (GSEA) and Gene Set Variation Analysis (GSVA) analyses. The regulatory relationship was then constructed, in which the correlation between EBF1 and RBP7 (R = 0.677, p < 0.001), Th2 cells and RBP7 (R = 0.23, p < 0.001), the oocyte meiosis pathway and RBP7 (R = 0.14, p = 0.042) were the most statistically significant. The results were further confirmed by Assay for Transposase Accessible Chromatin with high-throughput sequencing (ATAC-seq), Chromatin Immunoprecipitation sequencing (ChIP-seq), single-cell RNA sequencing (scRNA-seq), and multiple online databases validation.

Conclusions: This study revealed that the EBF1-RBP7 regulatory relationship had potential importance in the bone metastasis in BLCA through Th2 cells and the oocyte meiosis pathway.

Keywords: biomarkers; bladder urothelial carcinoma; bone metastasis; immune genes; retinol binding protein 7.