| Part I:The study on the relationship between the content of Mitochondrial DNA and 10398 mutation with clinical pathology in patients with cervical cancerObjective Mitochondrial DNA(mtDNA)mutations have been studied in many solid tumors,including changes in mtDNA content and mtDNA locus mutation.Previous studies have shown that mtDNA mutations were closely related with the occurrence,development and prognosis of solid tumor.This study aimed to explore the relationship between mtDNA mutations and pathologic grade,clinical stage,and prognosis of patients with cervical cancer(CC)in han Chinese women.Methods A total of 122 paraffin-embedded tissue samples were collected from patients pathologically diagnosed with CC since last 10 years in Yichang oncology hospital and Zhongnan Hospital of Wuhan University.The content of mtDNA was measured by quantitative real-time polymerase chain reaction assay;A10398G polymorphism was determined by PCR.Results In this part,we have recruited 122 patients.Aging from 22 to 77,the median age was45 years old,mtDNA contentvaried a lot,ranging from 14.27 to 21769.19,the average was 1758.18±3335.72.It was found that differences in mtDNA contentin patients with different stages and different pathological types was not statistically significant(P=0.213,0.361respectively),however,the difference in different age groups was statistically significant.,where the content was higher in elderly age group(>45 years)(P=0.019).And it also found that 10398 allele in CC had nothing to do with age,stages and pathological types(P=0.271,0.244,0.3respectively).As for the prognosis,we divided the patients into two subgroups of high and low mtDNA copy number,according to their mtDNA content above or below the average of mtDNA copy number in all 122 patients.The Kaplan-Meier survival curve demonstrated that the overall survival(OS)rate of patients with higher mtDNA content was significantly lower than that of lower mtDNA content(5-year OS rate was 51.3%vs.76.4%,p = 0.005;8-year OS rate was 34.5%vs.65.9%,p=0.002).But there were no statistically significant differences between 10398A allele and 10398G allele in prognosis of CC.When patients were divided into four subgroups,according to the mutation of 10398(A or G)and mtDNA content(low or high),the Kaplan-Meier survival curve illustrated that the OS of the patients with high mtDNA copy number plus 10398 A was significantly lower than that of others(5-year OS rate was 40%P=0.013;8-year OS rate was 18.2%P=0.009)Analysis of single factor COX regression and Multiariable COX regression showed that age,pathological type,and 10398 locus had no effect on prognosis of patients with CC(P>0.05),while the content of mtDNA and clinical stages had a significant effect on prognosis.Multivariate Cox analysis indicated thatthe risk of mortality in subgroup with higher mtDNA content significantly increased compared to the lower one(total 5 years survival of HR =2.461,95%Cl ＝1.190 5.090,P = 0.015;8 years total survival of HR = 2.470,95%CI =1.221 4.995,P = 0.012).moreover,the stage had a greater influence on prognosis,and patients of late stge had a significantly higher risk for mortality(total 5 years survival HR = 2.853,95%CI=1.434 5.677,P = 0.003;8 years total survival of HR = 3.225,95%Cl=1.5916.537,P = 0.001).Conclusions This study demonstrated that the mutation of mitochondria DNA 10398 locus does little influence on the prognosis of CC,while changes of mtDNA content have a significant effect on the prognosis of CC,which indicated that mtDNA can be expected as an independent prognostic factor for patients with CC,and can also be used as a molecular marker to predict prognosis of CC.Part Ⅱ:The relationship between the infection of HPV(HPV16 and HPV18)and the clinicopathology features of patients with cervical cancerObjective CC is one of the most common malignant tumors in female reproductive tract.A great deal of studies showed that high-risk type human papillomavirus(HR-HPV)infection is a necessary condition for the development of CC.This studywas to explore the effect of HPV16 and HPV18 infection status on the development of CC,and to figure out the relationship between HPV infection status and the mutation of mtDNA,thus to find out their influences on the prognosis of CC,combining with the results of part I of this study.Methods A total of 122 paraffin-embedded tissue samples were collected from patients pathologically diagnosed with CC since last 10 years in Yichang oncology hospital and Zhongnan Hospital of Wuhan University.The qualitative detection of HPV 16 and HPV18was performed by general PCR,then quantitative real-time PCR was used(Real-time PCR)for the determination of the content of positive cases.Results Among 122 cases of CC included in this study,there were 51 cases of HPV 18 positive,and 38 cases of HPV 16 positive,10 cases of both positive,and 79 cases of single positive in all.The positive rate of HPV 18,HPV 16,both positive and signal positive were 31.15%、41.80%、8.20%and 64.75%.The results indicated that the positive rate of HPV 18 was significantly higher than that of HPV 16(P=0.000),but the content of positive cases was higher in HPV 16 than that in HPV 18(P=0.013).There are two age groups with higher HPV positive infection rate,those were in the age of 30-39 and 40-49 groups,their positive rates were 76.92%and 71.15%respectively,and with the growth of age,their positive rate drops;And the positive rate of HPV 16 infection peaks in two age groups which are the same to that of above-mentioned HPV infection,and their positive rates were 50.00%and 34.62%respectively,then decreasing with age;While the two peaks of the positiverate of HPV 18 appeared in the age of 40-49 group and the group of people over 60 years old,their positive infection rate were 48.08%and 50%respectively.It also found that mtDNA content had positive correlation with HPV18 content(correlation coefficient r = 0.456,P = 0.022)in young group(<= 45).HPV 16 and total HPV infection positive rates are higher in 10398G,while the HPV18 positive rate is higher in the locus of 10398 A,but there were no statistically significant difference(P>0.05).Survival analysis indicated that patients with negative infection of HPV have a higher survival rate,moreover,patients of positive infection with a lower virus copy number also has a higher survival rate,but the differences were not significant(P>0.05).Single and Multiariable COX regression analysis both found that the status of HPV infection has a significant effect on the prognosis of patients with CC,and the content of mtDNA and stages are two independent factors for the prognosis of CC.Conclusions In Chinese han women,there are two age peaks of patients with HPV infection(HPV16 infection peaks were in the age of 30-39 and 40-49 groups;HPV18 appeared in the age of 40-49 group and the group of people over 60 years old).and the status of HPV infection are closely related with mtDNA mutation,although the status of HPV infection and 10398 locus mutation may have little effect on the prognosis of patients with CC,mtDNA content can be used as a molecular indicators for prognosis of CC. |
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