Role Of Key Phase-Ⅰ Metabolizing Enzymes In AFB1 Biotransformation And Curcumin Interventions In Liver Of Arbor Acres Broiler

Aflatoxin B1(AFB1)is the most well-studied,abundant and toxic form among all aflatoxins.AFB1 is often ingested by broilers through contaminated feed causing toxicity,depress immunity and leads to severe economic losses.It has been demonstrated that the main target organ of AFB1 is liver.Its metabolism mainly occurs in liver producing toxic metabolites and deposited in broilers edible meat which in turn ingested by human causing public health problems.However,the metabolism of AFB1 in inducing sub-chronic liver damage in Arbor Acres broilers is still unknown.Curcumin is a diferuloylmethane,extracted from the rhizome of ginger family.Curcumin acts as an anti-tumor,anti-oxidant and antiinflammatory in many diseases including toxicity.Recent studies described that curcumin plays a hepatoprotective role in fatty liver syndrome,hepatitis,liver cirrhosis,and liver cancer etc.However,the effect of curcumin in protecting broilers from AFB1-induced liver damage via cytochrome p450 enzymes especially CYP2A6 still remain unclear.AFB1 exerts toxic and carcinogenic effects after its bioactivation by cytochrome P450 enzymes.Previous studies reported that human hepatic CYP2A6 enzyme responsible for catalyzing several carcinogenic substances.In the present study,Arbor Acres(AA)broiler liver CYP2A6 gene partial sequence is characterized,and further scrutinized its role in AFB1 biotransformation.In addition,the inhibitory effects of curcumin on CYP2A6 enzyme is examined.Broilers were placed randomly into six groups and provided different experimental diets for 28 days.Experimental groups including control group(fed normal diet),curcumin control group(450 mg/kg feed),low dose curcumin group(150 mg),medium dose curcumin group(300 mg),high dose curcumin group(450 mg)and AFB1-fed group(5 mg/kg feed).Different analyses were carried out on liver samples after the end of treatment period.Histopathological examination revealed toxicity signs in liver in AFB1-fed group relative to control and curcumin control group.Intriguingly,curcumin-supplementation partially prevented AFB1-induced toxicity in broiler liver.The abnormal morphology of liver cells ameliorated with the medium and high dose curcumin compared to low-dose curcumin group.AFB1 causes significant(p<0.05)increase in liver weight and marked(p<0.05)decrease in body weights.While,curcumin supplementation(dose-dependently)in feed significantly(p<0.05)alleviated the rise in liver weight and decline in body weight.It has been noted that both protein and mRNA expression level increased in AFB1-alone group.Similarly,a significant(p<0.05)increase has been noted in CYP2A6 enzyme activity in AFB1-treated group,showing that AFB1 biotransformation was actively carried out by CYP2A6 enzyme.However,the supplementation of curcumin in diet inhibited CYP2A6 at both protein and mRNA expression levels in AFB1 group.Intriguingly,the inhibitory effect of high dose curcumin group(450 mg/kg curcumin)is more obvious.The current research for the first time identified and confirmed that CYP2A6 enzyme plays a crucial role in bioactivation of AFB1 in broiler liver(in-vivo),and in addition provided evidence about the hepatopreventive effects of curcumin by the subsequent inhibition of CYP2A6 expression at both molecular and enzyme activity level.Most importantly,the current research contributed to identify a novel CYP enzyme which is crucial for the bioactivation of AFB1,and therefore,could provide a mechanistic base for further studies to prevent the harmful effects of AFB1.Furthermore,the inhibitory effect of curcumin is examined on CYP1A1,CYP3A4 and CYP1A2.Compared to control group,the mRNA expression level of the above three CYP enzymes were significantly(p<0.05)decreased in high dose treated group.Importantly,the trends of inhibition of all the CYP isozymes are similar.CYP1A1,CYP1A2 and CYP3A4 expression level increased significantly in AFB1-fed group relative to control group.Therefore,all the four CYP enzymes(CYP1A1,CYP1A2,CYP3A4 and CYP2A6)were actively involved in AFB1 bioactivation,and responsible for AFB1 harmful metabolites in Arbor Acres broiler liver.Our data showed that curcumin could prevent AFB1 bioactivation via inhibiting CYP isozymes and provide scientific basis for the use of curcumin in broiler feed.