ScRNA-seq and bulk RNA-seq identified NUPR1 as novel biomarkers related to CD4 + T cells infiltration for abdominal aortic aneurysm.

Background: Developing a molecular signature associated with CD4 + T cell infiltration is essential for identifying biomarkers in abdominal aortic aneurysms (AAA). Establishing such a signature is vital for improving diagnostic accuracy and therapeutic strategies for AAA. This study focuses on CD4 + T cells, which are pivotal in the immune microenvironment of AAA, to pinpoint key targets.

Methods and results: We identified CD4 + T cell-related biomarkers in AAA using bulk and single-cell RNA sequencing data from the GEO database. We employed CIBERSORT to assess immune cell infiltration and applied weighted gene co-expression network analysis and differential expression analyses to pinpoint key genes. A nomogram and related score were developed based on these genes. Single-cell RNA sequencing further analyzed their expression across cell types, and KEGG/GO analyses were conducted for candidate genes. Four genes (NUPR1, CCL4L2, CCL3L3, MMP9) were identified finally. Validation via qPCR and immunohistochemistry showed NUPR1 downregulation in aneurysms and an inverse relationship with CD4 + T cells infiltration. Immunofluorescence results indicated NUPR1 was mainly expressed in the cytoplasm of vascular smooth muscle cells (VSMCs). After VSMCs-specific overexpression of NUPR1 via adeno-associated virus, the AAA diameter decreased, while treatment with the NUPR1 nuclear translocation inhibitor ZZW-115 hydrochloride had no effect on AAA size. Overexpression of NUPR1 in VSMCs suppresses the migration of CD4 + T cells.

Conclusion: The four-gene signature accurately predicts CD4 + T cell infiltration in AAA patients and may serve as a clinical index. NUPR1 could be a therapeutic target for the interaction between CD4 + T cells and AAA.