Ying-Huang Oral Liquid Ameliorates Oxidative Stress in the Liver of Heat-Stressed Broilers via KEAP1-NRF2 and PINK1/Parkin Pathway.

Heat stress (HS) impairs liver function and negatively affects the growth performance of poultry. This study evaluates the protective effects and underlying mechanisms of Yinhuang Oral Liquid (YOL) in alleviating liver injury in heat-stressed broilers, highlighting its potential application in mitigating HS-related liver damage. Broilers were randomly allocated into six groups: Control (23°C ± 2°C), Model (HS, 35°C ± 2°C for 8 h/day for 7 consecutive days), Positive (HS + Vitamin C, 60 mg/kg), YOL-H (HS + YOL, 10 g/kg), YOL-M (HS + YOL, 5 g/kg), and YOL-L (HS + YOL, 2.5 g/kg). Vitamin C and YOL were administered intragastrically 1 h after HS exposure for 7 days. HS impaired growth performance, as evidenced by decreased final body weight, average daily feed intake (ADFI), and average daily gain (ADG), along with an increased feed conversion ratio (FCR). YOL supplementation increased ADG compared with the model group. Histopathological analysis showed severe inflammatory cell infiltration around the hepatic central vein under heat stress, which was markedly attenuated by vitamin C and YOL. YOL activated the hepatic NRF2-KEAP1 signaling pathway by increasing the p-P62/P62 ratio, increasing NRF2 and its downstream antioxidant proteins HO-1 and NQO1, and downregulating KEAP1 expression. Network pharmacology analysis identified Sirt1 as a crucial bridging target linking YOL to heat stress. Molecular docking further demonstrated that the top four compounds-baicalin, wogonoside, oroxylin A-7-O-glucuronide, and acacetin-exhibited high binding affinity with the Sirt1 receptor. YOL significantly upregulated Sirt1, GPX4, and SLC7A11 protein levels and restored mitochondrial cristae integrity while promoting autolysosome formation. It also restored HS-induced suppression of PINK1 and Parkin. In conclusion, YOL effectively mitigates HS-induced oxidative stress in broilers by activating Sirt1, which coordinately enhances the NRF2-KEAP1 and PINK1/Parkin signaling pathways.