Vitamin C alleviates intestinal damage induced by 17α-methyltestosterone in Carassius auratus

17α-Methyltestosterone (MT), an environmental endocrine-disrupting chemical (EDC), is widely present in aquatic environments, posing potential threats to the health of aquatic organisms. This study aimed to explore the protective effects of Vitamin C (VC) against MT-induced intestinal injury in Carassius auratus and evaluate the optimal VC dosage. C. auratus were exposed to 50 ng/L MT with 0, 25, 50, and 150 mg/kg VC for 7, 14, and 21 d. Intestinal pathological changes were assessed using paraffin sections, digestive enzyme activity was measured, RT-qPCR was used to analyze the expression of genes related to the intestinal barrier and inflammation, and 16S rDNA sequencing was conducted to evaluate the intestinal flora. MT exposure caused villus damage, vacuolization, and free lymphocytes. Additionally, it upregulated TNF-α and Claudin-12 expression and downregulated IL-10, Occludin, and Muc2 expression, exacerbating intestinal inflammation, damaging barrier function, and reducing digestive enzyme activity. VC at 25 and 50 mg/kg significantly alleviated MT-induced damage by restoring villus length and mitigating the downregulation of anti-inflammatory factors and tight junction protein-related genes while inhibiting TNF-α mRNA expression. 16S rDNA sequencing revealed that MT disrupted the intestinal flora and increased the abundance of harmful bacteria such as Acinetobacter, whereas VC promoted Lactobacillus production and enhanced digestive enzyme activity. We hypothesize that MT exposure promotes the growth of harmful bacteria in the intestines, leading to inflammation and damage to the mucosal barrier. 25 and 50 mg/kg VC can mitigate MT-induced intestinal injury by regulating the intestinal microbiota and have potential protective effects.