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This study investigates the toxic effects of Docetaxel (DTX) on rat testicular tissue and the blood-testis barrier (BTB), as well as the protective effects of mesenchymal stem cells (MSCs) and extracellular vesicles (EVs) against DTX-induced damage. A total of 60 Wistar Albino male rats (8–10 weeks old) were used, divided into six groups: Control, Docetaxel (5 mg/kg weekly for 4 weeks), MSCs (1 × 106 cells via tail vein), EVs (30 µg via tail vein), DTX + MSC, and DTX + EVs. The rats were sacrificed under anesthesia, and testicular tissues were fixed in 10% formaldehyde. Paraffin blocks were prepared, and 5 μm sections were examined histopathologically using Hematoxylin & Eosin and PAS-H staining. Protein levels of occludin, laminin α-2, and β1 integrin were measured by western blot, while ZO-1, mTOR, MMP-2, and MMP-9 expressions were assessed via immunohistochemistry. Malondialdehyde, Superoxide dismutase, Catalase, and testosterone levels were measured by ELISA. Results indicated that DTX caused disruption of the seminiferous epithelium. Significant improvements were noted in the DTX + MSC and DTX + EVs groups compared to the DTX group. Occludin, laminin α-2, β1 integrin, and ZO-1 levels were decreased, while MMP-2 and MMP-9 levels increased in the DTX group. Treatment groups showed levels approaching those of the control group. Malondialdehyde was higher, while Superoxide dismutase, Catalase, and testosterone were lower in the DTX group. These findings suggest that DTX disrupts testicular BTB and adversely affects reproductive health, while MSCs and EVs offer protective effects.