The Contamination Of Heavy Metals Of Feedstuff In Anhui Province And The Cytotoxicity Induced By As?Hg?Cr?Pb To HepG2

This study surveyed the content of heavy metals in different feed in Anhui province,using a tetrazolium salt(MTT)assay and isobologram analysis the effect to HepG2 cells.This study was aimed to study the effect of the different joint action of heavy metals and provide theoretical basis for the different joint action of heavy metals of damage to animal.According to the cytotoxicity we choose two more significant impact on the HepG2 cells,arsenic and lead,to study the mechanism of them,and provide a reference of heavy metals for them cytotoxicity.1.This study surveyed the content of heavy metals in different pig,chick and duck farm in Anhui province and detected by ICP-MS.The results showed that the different type of heavy metals in the feed,such as arsenic(As),mercury(Hg),chromium(Cr),lead(Pb)and cadmium(Cd).The concentration of As,Hg,Cr,Pb and Cd are.0.15,0.038,1.19,2.66 mg/kg in early feed of piglets and 0.14,0.043,3.14,2.0 mg/kg in early feed of chickens,0.27,0.033,3.43,3.76 mg/kg in the feed of fish.Among the results,the As,Hg,Cr,Pb were 0.11-0.23 mg/kg,0.039-0.053 mg/kg,1.01-2.56 mg/kg,0.29-4.31 mg/kg in the all kinds of specimen.The results showed that As,Hg,Cr,Pb are widespread in all kinds of specimen in a low concentration than national standard.2.This study investigated the cytotoxic effects of different combinations of arsenic(As),chromium(Cr),mercury(Hg)and lead(Pb)on human liver HepG2 cells,using a tetrazolium salt(MTT)assay and isobologram analysis.The cell viability was decreased in a concentration-dependent manner from 95 to 36%,75 to 42%,76 to 22%,94 to 12%after exposure to As(2.65 to 4.65 ?g/ml),Hg(2.65 to 4.65 ?g/ml),Cr(40 to 70 ?g/ml),and Pb(160 to 280 ?g/ml),respectively,with the IC50 4.20,4.06,56.22,228.02 ?g/ml,respectively.The cytotoxicity of different heavy metal combinations follow an order of As+Cr+Pb>As+Cr+Hg+Pb>Cr+Hg+Pb>As+Cr+Pb>Cr+Pb>As+Hg+Pb>As+Hg+Cr>As+Pb= Hg+Pb>As+Cr>Hg+Cr>As+Hg.Isobologram analysis indicated an antagonistic effect for As+Hg,As+Cr,As+Pb,Hg+Cr,Hg+Pb,As+Cr+Hg and As+Hg+Pb respectively;a synergistic effect for Cr+Pb,Hg+Cr+Pb,respectively;both synergistic effect and additive effect for As+Cr+Pb;both synergistic effect and antagonistic effect for As+Hg+Cr+Pb.3.This study aimed to evaluate the cytotoxic effects and of single and combined effects of arsenic(As),lead(Pb)and their joint action on human liver carcinoma cells(HepG2).The mechanism of oxidative damage on HepG2 was also investigated.The HepG2 cells were cultured,adding 2.65,3.15,3.65,4.15,4.65 ?g/mL of As;160,190,220,250,280 ?g/ml of Pb;the joint action of As+Pb at the concentration of ratiosl:55,based on the IC50.The cell viability was assayed by MTT method after 24,48 and 72 h of exposure.The quantitative analysis of lactate dehydrogenase(LDH),reactive oxygen species(ROS),glutathione(GSH)and malondialdehyde(MDA)was carried out to represent oxidative damage.After 24,48 and 72 h exposure of As,the IC50 values were 4.20,4.03 and 3.74?g/mL,respectively;After 24,48 and 72 h exposure of Pb,the IC50 values were 228.01,205.46 and 203.40 ?g/mL,respectively;After 24,48 and 72 h exposure of the joint action of As+Pb,the IC50 values were 3.10?3.17?3.18 ?g/mL,respectively.The HepG2 exposured in As,Pb and As+Pb with different concentration and time resulted in significant lower levels of cell viability and inhibition of cell growth(P<0.05).The As,Pb as well as the joint action of As+Pb caused significant higher levels of LDH,ROS and MDA(P<0.05),but the GSH level showed significant lower level(P<0.05).In conclusion,the joint action of As+Pb exerted a stronger cytotoxicity than As,Pb,indicating an antagonistic effect.The cytotoxicity followed an order of:As+Pb>As>Pb.The reactive oxygen and metabolites from oxidative stress may be an important factor for cytotoxicity.