Dynamics of viability and apoptotic regulation in chicken ileal explant culture

The development of a reliable ex vivo intestinal model is essential for studying gut physiology and host-microbe interactions under controlled conditions. This study evaluated the temporal dynamics of viability, morphology, and apoptotic regulation in chicken ileal explants cultured ex vivo. Ileal tissues from 21-d-old broiler chickens were incubated for 0, 4, and 8 h and analyzed using metabolic assays (MTS, CCK-8), histological staining (hematoxylin and eosin (H&E), Periodic Acid-Schiff (PAS)), and quantitative PCR for apoptosis-related genes (BCL2L11, cytochrome c, caspase-3, caspase-8, and caspase-9). After 4 h, explants retained structural integrity with well-developed villi and active mucus secretion. Metabolic assays confirmed preserved cell viability, while transcriptional analysis revealed moderate upregulation of cytochrome c and downregulation of BCL2L11, suggesting transient adaptation to culture stress. After 8 h, villi appeared shortened with epithelial exfoliation and reduced PAS reactivity, indicating progressive tissue deterioration. Correspondingly, BCL2L11 and cytochrome c were significantly upregulated together with caspase-8, reflecting activation of intrinsic and extrinsic apoptotic pathways. Metabolic activity declined markedly, confirming reduced tissue vitality. These findings demonstrate that chicken ileal explants with a diameter of 4 mm remain viable and functionally stable up to approximately 4 h of ex vivo culture, after which apoptotic processes intensify, compromising tissue integrity. The results define the practical viability window for short-term intestinal explant studies and provide methodological guidance for future investigations on host-pathogen or probiotic interactions in poultry gut models.