Protective Effect Of Purple Tomato Anthocyanin On Autophagy Of LMH Cells Induced By Chromium(Ⅵ)

In animal husbandry,chromium is often used as a feed additive to improve the carcass quality of livestock and poultry,improve feed utilization and enhance anti-stress ability.However,improper application of chromium can also cause chronic poisoning in animal organism.Many studies have shown that the liver is one of the main target organs of chromium,and excess chromium can induce autophagy and endoplasmic reticulum(ER)stress in a variety of animal models.Anthocyanin are natural flavonoids that have antioxidant,scavenging free radicals,anti-cancer and anti-tumor effects.In order to study the toxic effects of Cr(Ⅵ)on LMH cells and the regulation of Purple tomato anthocyanin(PTA)on autophagy and ER stress induced by Cr(Ⅵ)in LMH cells,the protective effect of PTA was studied from two aspects of ER stress and autophagy.In this experiment,we established a model of LMH cells poisoning and then used PTA and Cr(Ⅵ)were treated separately or co-incubated for 20 h.Cr(Ⅵ)-induced LMH cells injury,autophagy and anthocyanin protection was examined by cell morphology observation and cell viability.The relationship between ER stress and autophagy in this study was verified by the addition of ER stress inducer(Tg)and inhibitor(4-PBA).The changes of ER stress proteins(GRP78/Bip and PERK),autophagy-related proteins(LC3,P62,Beclin1 and mTOR)and COX-2 expression by Western Blot and ELISA,were used to verify that effects of Cr(Ⅵ)and PTA on ER stress and autophagy in LMH cells.The result demonstrated that Cr(Ⅵ)induced cell shedding and decreased the cell size and density,reducd cell viability,while PTA could reduce the damage of Cr(Ⅵ)to cells.When 3-MA inhibited autophagy,it could significantly improve the cell viability reduction induced by Cr(Ⅵ),and PTA played a similar role as 3-MA.When an ER stress inducer(Tg)or inhibitor(4-PBA)is added,the decrease in cell activity caused by Cr(Ⅵ)could be correspondingly aggravated or alleviated.Cr(Ⅵ)could markedly induce autophagy-related proteins(Beclin1 and LC3 II/I),ER stress-related proteins(GRP78/Bip and PERK)andCOX-2 expression,inhibit the autophagy pathway protein mTOR and the degradation of autophagy-related protein P62.PTA could relieve Cr(Ⅵ)-induced the changes of above proteins.In conclusion,our study suggested that Cr(Ⅵ)can induce excessive autophagy in LMH cells,while PTA could ameliorate Cr(Ⅵ)-induced autophagy and ER stress,which provides a theoretical basis for the screening of Cr(Ⅵ)-induced heavy metal poisoning protectants.