Protective Role of Folic Acid and Vitamin B12 Against Chronic Ivermectin‐Induced Reproductive Damage: Oxidative Stress, Immunohistochemical Changes (Caspase‐3, Bax, BCL2, and PCNA Expressions) and Mitochondrial 16S rRNA Analysis

ABSTRACT

This study aims to investigate the protective effects of folic acid (FA) and vitamin B12 (Vit.B12) against high‐dose effects of IVM on reproductive hormones, semen quality, oxidant/antioxidant profile in testicular tissue, histopathological changes, and immunohistochemical investigation (Caspase‐3, Bax, BCL2, and PCNA) in the testes of male rats. Twenty eight rats were divided into four equal groups: Control, Ivermectin, Ivermectin + Folic acid, and Ivermectin + Vit.B12. IVM was administered at 5 mg/kg/day, folic acid at 20 mg/kg, and vitamin B12 at 50 µg, with the respective routes and 30‐day treatment period specified. Testosterone and semen parameters were evaluated. Testicular specimens were examined for oxidant/antioxidant markers, histological changes, and immunohistochemical expressions. Mitochondrial 16S‐rRNA sequences were analyzed and deposited in GenBank under accession numbers PP320440.1–PP320443.1. Results showed that IVM significantly reduced testosterone level, sperm count, and antioxidants, while increasing oxidative stress. Seminiferous tubules damage was observed in the IVM group. Immunohistochemical analysis showed strong caspase‐3 and Bax expressions in the IVM group. BCL2 and PCNA showed altered expression patterns. 16S‐rRNA sequence analysis revealed limited sequence variation among the experimental groups. Co‐treatment with folic acid or Vit.B12 significantly improved all parameters affected by IVM. These findings indicate that FA and Vit. B12 may provide protective effects against IVM‐associated testicular alterations.