Sequence Analysis Of The Mitochondrial DNA D-Loop Region In Hepatocellular Carcinoma Exposeing To Hepatitis B Virus And Aflatoxin B₁ In Guangxi

Objective:To find out the characteristic molecular biology marks in the mitochondrial DNA(mtDNA)D-Loop region in hepatocellular carcinoma(HCC)exprosing to hepatitis B virus(HBV)and aflatoxin B1(AFB1)in Guangxi and the correlation between the variation of D-Loop region sequence and HCC pathogenesis.Methods:32 tumors from the patients for radical resection of HCC in the department of hepatobiliary surgery of the First Affiliated Hospital of Guangxi Medical University were included for study,32 matching non-HCC liver tissues and the peripheral blood leukocyte from those patients were categorized into self control Ⅰ and self control Ⅱ.Enzyme-linked immuno sorbent assay(ELISA)was used to detecte the levels of hepatitis B virus surface antigen(HBsAg)for defining the HBV exprosure.Polymerase chain reaction(PCR)and direct sequencing were applied to study the mutations in codon 249 of p53 for delimiting the AFB1 exprosure.According to situation of HBV and AFB1 exprosure,32 tumors were categorized into the subgroup A,B,C and D.The normal control included 8 normal liver tissues from the donors for liver transplantation or the patients underwent liver trauma without any liver disease.We detected and analysed the D-Loop region sequences of all samples by PCR and direct sequencing.Results:There were 71.9%patients exprosed to HBV and 48.9%patients exprosed to AFB1 with HCC.The alteration of mtDNA D-Loop region in tumors(7.6%),non-HCC liver tissues(7.0%)and peripheral blood leukocyte(7.6%)were remarkable higher then that in nomal control(2.7%,P<0.01).The sequences of tumors,non-HCC liver tissues and peripheral blood leukocyte from the same patient were nearly in accordant.The match scores were from 92.8%to 100.0%.In the 98 variational positions,81.7%were reported polymorphisms,then 5 mutations and 18 new polymorphisms had been found.All of the samples have the polymorphisms:NT73A>G and NT263A>G.NT150C>T,NT310T>CTC(NT309Cins and NT311Cins),NT489T>C,NT16093T>C,NT16223C>T,NT16234C>T,NT16362T>C and NT16519T>C occur in study group,self control I,self control II and nomal control with a high frequency and unconspicuous difference.The polymorphisms of NT249A-del,NT523A-del,NT524C-del and NT16129G>A can been significative observed in the HCC patients greater than 40%,but not in nomal control.Conclusions:1.The exposure to HBV and AFB1 were the primary dangerous factors for high incidence of HCC in Guangxi.The most important task for HCC was prevention of HBV and AFB1 exposure.2.The alteration of NT73A>G,NT150C>T,NT263A>G,NT3110T>CTC(NT309Cins and NT311Cins),NT489T>C,NT16093T>C,NT16223C>T,NT16234C>T,NT16362T>C,NT16519T>C may be the polymorphism of specific haploid groups in Guangxi.3.In this study we had not found any precise and characteristic molecular biology marks or mutaions of nucleotide sequences in the mtDNA D-Loop region of HCC exprosing to HBV and AFB1 in Guangxi,but we suggested that the polymorphism of NT249A-del,NT523A-del,NT524C-del and NT16129G>A may associate with the HCC pathogenesis.