Effects Of Dietary Supplementations Of Zn From Different Sources On Antioxidant Capacity Of Weaned Piglets

Dietary Zn supplementation can improve antioxidant capacity in weaned piglets.Yet there is certain disputation about the bioavailbility of Zn from organic and inorganic sources.HMZn is a kind of organic Zn with productive prospect,as it is chemically stable and provides both Zn and methionine.In present study,we systematically compared theregulation of HMZn and ZnSO₄ supplementation on diquat challenged piglets and Cd challenged IPEC-J2 cells,and furtherly explored the underlying mechanism.The main research contents and results are listed as follow:1.Regulation of HMZn and ZnSO₄ on antioxidant capacity of diquat challenged weaned pigletsA total of 32 30-day-old“D×L×Y”hybrid weaned piglets（9.41±0.11kg）were selected for single factor trial and randomly divided into the following 4 treatments:blank control group（basal diet+diquat challenge+80 mg/kg Zn as ZnSO₄）,diquat challenged control group（basal diet+80 mg/kg Zn as ZnSO₄）,HMZn group（basal diet+diquat challenge+200 mg/kg Zn as HMZn）,ZnSO₄ group（basal diet+diquat challenge+200 mg/kg Zn as ZnSO₄）.The piglets were treated with diquat by intraperitoneal injection on day 14^th.The results showed that there was no significant difference on growth performance,serum amino acid contents between the different diet treatments during the pre-starter period,while HMZn upregulated serum SOD1、GPx activity and T-AOC.During the starter period,diquat challenge resulted decreased ADG,increased F/G and diarrhea rate in diquat control group;serum Ser content decreased;liver,kidney and spleen index remained uneffected;Zn,Fe contents of liver,kidney and longissimus muscle remained uneffected;Zn T1 m RNA expression decreased;serum GPx activity and antioxidant capacity decreased;m RNA expression of antioxidant related genes（e.g.Nrf2,SOD1）in intestine,liver,kidney decreased;jejunal villus height decreased;m RNA expression of intestinal tight junction related genes decreased;intestinal NF-κB and m TOR were activated（P<0.05）.Compared with the diquat challenged control group,both HMZn and ZnSO₄supplementation of 200 mg/kg failed to show notable improving effect on growth performance;while they both improved serum GPx activity,upregulated duodenal Nrf2 m RNA expression;upregulated hepatic,renal and duodenal SOD1 m RNA expression;upregulated hepatic GPx m RNA expression;downregulated duodenal and jejunal IL-1βm RNA expression（P<0.05）.In addition,HMZn downregulated the diarrhea rate;upregulated hepatic Zn and renal Cu content;upregulated serum T-AOC;upregulated duodenal ZO-1,international occludin m RNA expression;up-regulated hepatic Nrf2,jejunal CAT m RNA expression;down-regulated renal,ileal IL-1βm RNA expression;down-regulated intestinal AMPK,m TOR,NF-κB phosphorylation levels（P<0.05）.In general,HMZn showed notable advantage over ZnSO₄ in improving antioxidant capacity,preventing diarrhea,increasing Zn accumulation,maintaining intestinal barrier,alleviating the diquat induced inflammation response in weaned piglets exposed to diquat challenge.2.Effects of mixed HMZn and ZnSO₄ on diquat challenged weaned pigletsA total of 32 30-day-old“D×L×Y”（9.39±0.13 kg）hybrid weaned piglets were selected for single factor trial and randomly divided into the following 4 treatments:blank control group（basal diet+diquat challenge+80 mg/kg Zn as ZnSO₄）,diquat challenged control group（basal diet+80 mg/kg Zn as ZnSO₄）,HMZn+ZnSO₄ group（basal diet+diquat challenge+120 mg/kg Zn as HMZn+80 mg/kg Zn as ZnSO₄）,ZnSO₄ group（basal diet+diquat challenge+200 mg/kg Zn as ZnSO₄+545.3 mg/kg HM）.The piglets were treated with diquat by intraperitoneal injection on day 14^th.The results showed that there was no significant difference on growth performance,serum amino acid contents and antioxidant capacity between the different diet treatments during the pre-starter period.During the starter period,compared with diquat control group,both mixed Zn and ZnSO₄ supplementation upregulated serum GPx activity and hepatic GPx m RNA expression;upregulated hepatic and renal SOD1m RNA expression;upregulated jejunal and ileal CAT m RNA expression;upregulated duodenal Zn T1,ileal ZO-1 m RNA expression increased;down-regulated duodenal and jejunum IL-1βm RNA expression decreased（P<0.05）;In addition,the mixed Zn also up-regulated hepatic and duodenal MT-1 m RNA expression,jejunal occludin m RNA expression;down-regulated renal IL-1βm RNA expression（P<0.05）.In general,200 mg/kg supplementation of both mixed Zn and ZnSO₄ exerted certain effect on antioxidant capacity of diquat challenged piglets,wihle the mixed Zn was slightly advanced.3.Effects of HMZn and ZnSO₄ on Cd challenged IPEC-J2 cellsIPEC-J2 cells were challenged with various concentrations of Cd Cl₂（10～200μmol/L measured by Cd）,and 30μmol/L was proved to be the optimized concentration to establish oxidative stress in IPEC-J2 cells according to the resulted cell viability.6 groups were designed by Cd challenge and Zn supplementations:blank Con,Cd Con,Cd+60μmol/L MHZn,Cd+90μmol/L MHZn,Cd+60μmol/L ZnSO₄,Cd+90μmol/L ZnSO₄.Cell viability,expression and phosphorylation of NF-κB related proteins were determined after 24 h of exposure to the designed treatments.The results showed that,compared with Con group,phosphorylation of NF-Κb,IKKα/β,IκBα,PI3K and expression of TRAF6 were significantly upregulated in Cd Con group,indicating that NF-κB signal pathway was activated by Cd challenge;compared with Cd Con group,both HMZn and ZnSO₄ significantly alleviated the Cd induced alteration in phosphorylation levels of IKKα/β,IκBα,PI3K and expression levels of TRAF6,while ZnSO₄ especially downregulated phosphorylation of JNK（P<0.05）.In conclusion,60μmol/L,90μmol/L of both HMZn and ZnSO₄ were effective in alleviating the Cd induced alteration in phosphorylation and expression levels of NF-κB signal pathway related proteins,and60μmol/L ZnSO₄ turned out to be as effective as 90μmol/L HMZn in alleviating the Cd induced stress.Conclusion:HMZn was more efficient than ZnSO₄ in promoting Zn accumulation and antioxidant capacity,protecting intestinal barrier and maintaining its function by downregulating imflammatory factor genetic expression and AMPK and NF-κB phosphorylation in diquat challenged piglets.In general,HMZn was more effective than ZnSO₄ in alleviating diquat induced oxidative stress in weaned piglets,and the combination of HMZn and ZnSO₄showed no advantage.