TOP2A drives T-cell infiltration and immune remodeling in cyclophosphamide-induced cystitis: a single-cell sequencing study with potential implications for interstitial cystitis.

Abstract

OBJECTIVE: To explore the potential mechanisms of interstitial cystitis (IC), we employed a cyclophosphamide (CYP)-induced cystitis rat model, a well-established tool for studying IC-like bladder inflammation and dysfunction. This study aimed to investigate the role of rhythmic genes and immune microenvironment remodeling in this model, focusing on TOP2A and its impact on T-cell infiltration. METHODS: CYP-induced cystitis rat models were established using cyclophosphamide. Single-cell RNA sequencing was performed on bladder tissues to analyze cellular heterogeneity. Differentially expressed genes (DEGs) and weighted gene co-expression network analysis (WGCNA) identified rhythmic and immune-related gene clusters. TOP2A was validated via RT-PCR, Western blot, and immunohistochemistry (IHC). Statistical analyses assessed correlations between TOP2A, CD4&#x2009;+&#x2009;T cells, and CD8&#x2009;+&#x2009;T cells. RESULTS: Single-cell sequencing revealed elevated T-cell infiltration in a CYP-induced cystitis rat model. TOP2A was the sole overlapping gene between rhythmic and immune clusters and showed significant upregulation in IC tissues (P&#x2009;<&#x2009;0.05). IHC confirmed increased TOP2A, CD4&#x2009;+&#x2009;T, and CD8&#x2009;+&#x2009;T cell levels, with strong positive correlations (r&#x2009;=&#x2009;0.89 and 0.64, respectively). Functional enrichment linked TOP2A to oxidative phosphorylation and ribosomal pathways. CONCLUSIONS: Our findings demonstrate that TOP2A drives immune dysregulation in CYP-induced cystitis by modulating T-cell infiltration. As T-cell infiltration is a hallmark of human IC, our findings in this CYP-induced model suggest that TOP2A may represent a novel therapeutic target worthy of further investigation in human IC tissues.