Crocin Modulates AKT/mTORC1/ULK1 Cascade to Attenuate Ferritinophagy-Induced Colonic Oxidative Injury in Ulcerative Colitis: Network Pharmacology, Molecular Docking, and Experimental Validation.

Crocin (CRO) is an antioxidant carotenoid extracted from the Crocus sativus plant. CRO impacts many biological pathways related to inflammation, but its activity as an antioxidant and anti-inflammatory agent against ferritinophagy-associated oxidative damage and cell death in colonic mucosa is little understood. Therefore, this study predicted CRO-driven protective targets and processes against ulcerative colitis (UC) using network pharmacology, docking, and experimental analysis. Network analysis and molecular docking were used to analyze CRO and UC targets and pathways. To test the pharmacological efficacy of CRO in vivo, 24 rats were separated into four groups: Cont, UC, mesalazine (100 mg/kg), and CRO (40 mg/kg). Rats received oral treatments for 8 days before UC induction. A single intrarectal injection of 2 mL acetic acid (AA) caused UC in rats. Estimated DAI and colonic protein content. Oxidant/antioxidant status and inflammatory markers ULK1, FTH-1, Beclin-1, GPX4, NCOA4, and SLC7A11 were also evaluated. Additionally, western blot was used to measure mTOR, AKT, and LC3B protein. Both histopathological and immunohistochemical studies were done. The network showed 228 CRO-UC targets, including CASP3 in the top 10 targets. AKT2 and MAP1LC3B were 56th and 63rd, respectively. The top 30 markedly enhanced KEGG pathways included "Pathways in cancer" and "Kaposi sarcoma-associated herpesvirus infection". Docking confirmed CRO's significant binding affinity for ULK1, FRIH, GPX4, and SLC7A11. In vivo, CRO pre-treatment diminished ULK1, autophagic proteins (Beclin-1 and LC3B-II/LC3B-I), and ferritinophagy-related protein (NCOA4), while elevating Akt/mTORC1, FTH-1, SLC7A11, and GPX4, thereby mitigating iron overload, ROS, and ferroptosis. Changing AKT/mTORC1/ULK1 pathway in UC pathogeneses improved colonic morphological and macroscopic abnormalities in CRO-treated rats. This study suggests that CRO may reduce ferritinophagy-induced colonic oxidative damage in rats via modulating AKT/mTORC1/ULK1. Ferritinophagy as an attractive therapeutic target in different experimental and clinical conditions deserves further investigation.