Protective Effects And The Potential Mechanisms Of Viola Yedoensis Makino On Liver Injury Induced By Lipopolysaccharide In Broiler

Background: With the rapid development of intensive and large-scale chicken farming,high-density breeding environment leads to the breeding of bacteria and the rapid spread of diseases.Liver,an important organ of the chickens which has the function of metabolism and detox,occurs inflammatory response and oxidative stress when the chickens are infected by external pathogens,which resulting in the decline of disease resistance and even death.Viola Yedoensis Makino(Vy),a traditional Chinese herbal medicine,possesses immunomodulatory,antioxidant and anti-inflammatory activities,which implies that Vy may have promising clinical applications in the chicken farming as an immunomodulator and antioxidant.Nevertheless,no studies have investigated the protective effects of Vy on liver damage in broiler chickens.Objective: The model of liver damage in broiler chickens was established by administrating lipopolysaccharide(LPS)intraperitoneally,and Vy was supplemented in broilers’ diet.The effects of Vy on the liver damage as well as the potential mechanisms was explored in broiler chickens,which was expected to lay a scientific and theoretical basis for Vy as a feed additive.Methods: A total of 50 1-day-old AA male broilers were divided into 5 groups.The control and the LPS groups were fed the basal diet,and the other three groups were fed the diet supplemented with 0.5%,1.5%,and 4.5% of Vy.Liver injury was established by intraperitoneal injection of LPS(1 mg/kg.BW)on the day 17,day 19 and day 21,and the control group was injected with an equal dose of normal saline(1 mg/kg.BW).The blood and the livers were collected after slaughtering of broilers on the day 21.The morphological changes of the liver were assessed by H&E staining,the ultrastructure was estimated by electron transmission microscope,and the percentage of cell apoptosis was detected by TUNEL assay,to investigate the protective effect of Vy on liver injury challenged with LPS.Moreover,the levels of inflammatory cytokines and inflammatory mediators as well as the nuclear factor kappa B(NF-κB)/nucleotide-binding oligomerization domain-like receptor protein 3(NOD-like receptor protein 3,NLRP3)signaling pathways-related m RNA expression were detected by RT-PCR,which to explore the anti-inflammatory effect and the regulatory mechanism of Vy on LPS-evoked liver injury.simultaneously,the redox system was assayed by kits,and the m RNA expression of nuclear factor E2-related factor 2(Nrf2)/mitogenactivated protein kinase(MAPK)signaling pathways were detected by RT-PCR,which to investigate the antioxidative effect and regulatory mechanism of Vy on LPS-triggered liver injury.Results:(1)Supplementation with Vy ameliorated the reduction in growth performance,liver index,and the level of TC,and inhibited hyperemia,inflammatory infiltration and the cell apoptosis in liver of broilers challenged with LPS,implying Vy had a protective effect on LPSinduced liver injury.(2)Supplementation with Vy down-regulated the m RNA expression of IL-1β,IL-18,IL-8 and TNF-α in liver of broiler induced by LPS,and 4.5%Vy down-regulated the m RNA expressions of My D88,NF-κB and NLRP3,suggesting that Vy alleviated LPSinduced liver inflammation by regulating NF-κB/NLRP3 signaling pathway.(3)Supplementation with Vy significantly decreased the LPS-induced increase of MDA,and inhibited the activities of CAT,SOD and GSH-Px compensatory.Moreover,Vy enhanced the m RNA expression of Nrf2 and HO-1,and inhibited the m RNA expression of MAPK9.Furthermore,4.5%Vy down-regulated the m RNA expression of p38,JNK and ERK,suggesting that Vy alleviated LPS-induced oxidative stress by activating Nrf2 and inhibiting MAPK signaling pathway.Conclusion: Vy supplementation alleviates inflammation and oxidative stress via regulating NF-κB/NLRP3 and MAPK/Nrf2 signaling pathways,thus providing preventive protection against LPS-induced liver injury in broilers.