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Testicular injury is the primary pathogenesis of diabetes‐induced male infertility. Dioscorea zingiberensis (DZ), a traditional Chinese medicine (TCM) including saponins, flavonoids and cellulose, is used to treat diseases in the reproductive system. But the protective effects of DZ on diabetes‐induced testicular injury remain poorly understood. In this study, the therapeutic effects of chronic oral DZ treatment on testis impairment in a diabetic mouse model were explored by assessing sperm morphology, blood–testes barrier (BTB) integrity and testicular histological examination. Our results showed that DZ significantly reversed BTB disruption, testicular tissue injury and abnormal sperm morphology in diabetic mice. Interestingly, diabetes‐induced disruption of the BTB was associated with a decrease in the tight junction (TJ) protein zonula occludens‐1 (ZO‐1). Dioscorea zingiberensis effectively increased ZO‐1 expression in testis tissue to restore the integrity of the BTB. Moreover, DZ treatment significantly reduced hyperglycaemia‐induced increases in malondialdehyde (MDA) and 8‐hydroxy‐2′‐deoxyguanosine (8‐OHdG) levels. Further mechanistic studies revealed that DZ substantially enhanced the expression of Nrf2, NOQ1 and HO‐1, which indicated that DZ exerts potential antioxidant effects against testicular tissue damage via the activation of Nrf2. In conclusion, the protective effects of DZ rely on repairing the integrity of the BTB and on reducing oxidative stress damage by mediating ZO‐1 and Nrf2. The study contributes to discovering the DZ possible mechanism of action.

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