Exercise attenuates blood-testis barrier dysfunction in type 2 diabetic mice with restoration of Setd1b/H3K4me3-associated claudin transcription

Type 2 diabetes mellitus (T2D) is a prevalent metabolic disease that causes multisystem complications, including male reproductive dysfunction. However, the molecular basis of diabetes-induced spermatogenic impairment remains incompletely understood. Although exercise is widely recognized as protective for male reproduction, the mechanisms by which it alleviates blood-testis barrier (BTB) injury remain unclear. Here, we established a T2D mouse model using a high-fat diet combined with streptozotocin (STZ) and applied treadmill exercise intervention to investigate the mechanisms underlying exercise-mediated BTB protection. T2D disrupted testicular architecture, as evidenced by disorganized seminiferous tubules, irregular tubular contours, indistinct seminiferous epithelial layers, and loosely arranged germ cells, while biotin tracer staining demonstrated increased BTB permeability. Proteomic profiling revealed a clear separation between control and T2D testes, with differentially expressed proteins enriched in chromatin organization, chromatin binding, euchromatin, and transcriptional regulation. Among these, the histone methyltransferase Setd1b was aberrantly expressed, suggesting that epigenetic dysregulation contributes to BTB injury. RNA-seq further identified marked transcriptional alterations in BTB-associated Claudin family members. After 6 weeks of treadmill exercise, testicular histopathology was alleviated and BTB leakage was reduced. Exercise also increased DNase I–TUNEL signals, indicating enhanced chromatin accessibility, and restored H3K4me3 and Setd1b expression. In parallel, exercise upregulated Cldn3, Cldn5, Cldn9, and Cldn11 at both the protein and mRNA levels and increased H3K4me3 enrichment at their promoter regions. These findings reveal a critical role for Setd1b/H3K4me3-associated Claudin regulation in exercise-mediated protection against diabetes-associated BTB injury and further underscore exercise as an important non-pharmacological strategy for counteracting male reproductive dysfunction caused by metabolic disorders.