The Effects Of Aflatoxin,Fumonisin And Nitrite Exposure And Oxidative Damage On Esophageal Precancerous Lesions:A Case-control Study

Objective:（1）To explore the risk factors and protective factors of esophageal precancerous lesions（EPL）in Huai’an District,Jiangsu Province,a high-incidence area of esophageal cancer.（2）To determine the levels of aflatoxin B₁（AFB₁）,fumonisin（FB）and nitrite exposures in EPL cases and normal controls and analyze the differences between the two groups,and to explore the relationship between the exposure of AFB₁,FB and nitrite and EPL.（3）To test the levels of oxidation and anti-oxidation in EPL cases and controls,and to explore the relationship between oxidative stress,oxidative damage and EPL.In general,this study aimed to provide scientific evidence for primary prevention and etiology of esophageal cancer.Methods:（1）The population was from the project of early diagnosis and treatment of esophageal cancer in Huai’an District,Huai’an City,Jiangsu Province from January 2015 to June 2017.100 patients with EPL and 100 controls were matched by region,age,and sex.Unified questionnaire was used to collect data,including demographic characteristics,dietary habits,smoking history,drinking history,disease history,family history of cancer,food frequency,et al.Logistic regression analysis was used to analyze the results of the survey.（2）Duplicate portions of three-consecutive-day diets were collected from 100 cases and 100controls.Contents of AFB₁,FB and nitrite in diet were determined,and then the differences in the exposure levels of AFB₁,FB and nitrite between EPL cases and normal controls were analyzed.AFB₁-albumin adducts（AFB₁-Alb）,sphinganine（Sa）,sphingosine（So）and nitrite content in blood samples were determined,and differences in the exposure levels of AFB₁-Alb,Sa,So,Sa/So and nitrite between EPL cases and normal controls were also analyzed.Pearson correlation test was performed on dietary and blood indicators.（3）Levels of vitamin C,total superoxide dismutase（T-SOD）,glutathione peroxidase（GSH-PX）,catalase（CAT）,malondialdehyde（MDA）and 8-hydroxy-2-deoxyguanosine（8-OHdG）in blood samples of EPL cases and controls were determined,and oxidative stress and oxidative damage in the study subjects were analyzed as well.Results:（1）Multivariate Logistic regression analysis showed that drinking aclcohol[odds ratio（OR）=4.086,95%confidence interval（95%CI）:1.605-10.402],eating pickled foods（OR=2.963,95%CI:1.561-5.625）and eating corn（OR=2.468,95%CI:1.183-5.152）were risk factors for the development of EPL,eating soy products（OR=0.326,95%CI:0.159-0.666）was a protect factor for EPL.（2）AFB₁ was detectable in 93%of 100 food samples collected from patients with EPL and 98%of 100 food samples collected from controls.There was no significant difference in dietary AFB₁ levels between two groups（P>0.05）.Comparing this level with AFB₁ levels in previous studies,it was found that there was a high level of AFB₁exposure in both EPL cases and controls.There was a significant difference in serum AFB₁-Alb levels between two groups（P<0.05）.According to the median level of serum AFB₁-Alb,the population was divided into AFB₁ high exposure group and AFB₁ low exposure group.High levels of serum AFB₁-Alb significantly increased the risk of EPL（OR=11.232,95%CI:5.810-21.714）.A significant positive correlation was found between dietary AFB₁ levels and serum AFB₁-Alb（r=0.272,P<0.01）.（3）FB was detectable in 58%of 100 food samples collected from patients with ELP and 52%of 100 food samples collected from controls.There was no significant difference in dietary FB levels between two groups（P>0.05）.Compared with FB levels in previous studies,it was found that there was not a high level of FB exposure in patients with EPL and controls.The Joint FAO/WHO Expert Committee on Food Additives（JECFA）set a provisional maximum tolerable daily intake（PMTDI）of 2μg/kg body weight（b.w.）for fumonisin B₁（FB₁）.66 and 72 persons in the case group and control group respectively within this standard.There was no statistical difference between two groups in dietary FB exceeding standard（P>0.05）.There were no significant differences in serum Sa,So and Sa/So between two groups（P>0.05）.There was no significant correlation between dietary FB levels and serum Sa,So and Sa/So（r=0.078,P=0.306;r=-0.023,P=0.745;r=0.106,P=0.160）.（4）Nitrite was detectable in 100%of all 200 food samples collected from 200 subjects.Subjects with lesions had higher dietary exposure to nitrite than that in controls（P<0.05）.An increased risk of EPL was observed for cases or controls in the highest intake quartile of nitrite（highest vs.lowest quartile OR=2.256,95%CI:1.012-5.026）.There is a dose-effect relationship between nitrite intake and EPL risk.JECFA set an acceptable daily intake（ADI）of 0-0.07mg/kg b.w.for nitrite.When 0.07mg/kg b.w.for nitrite was used as the maximum limit,76 and 59 persons in the case group and control group were found respectively to exceed the standard.Subjects in the case group had a higher rate of exceeding the standard than those in the control group（P<0.05）.There was no significant difference in plasma nitrite levels between two groups（P>0.05）.There was no significant correlation between dietary nitrite and plasma nitrite（r=-0.019,P=0.793）.（5）The levels of serum GSH-PX,T-SOD and CAT in patients with EPL were higher than those in normal controls（P<0.05）.Serum vitamin C level in patients with EPL was lower than that in controls（P<0.05）.Serum MDA level in patients with EPL was higher than that in controls（P<0.05）.There was no significant difference in serum 8-OHdG levels between case group and control group（P>0.05）.Conclusion:（1）The questionnaire survey of patients with EPL and normal controls in Huai’an area found that drinking aclcohol,consumptions of corn and pickled foods were risk factors for EPL,whereas eating soy products was a protect factor.The region should strengthen the prevention and control of mycotoxins in major food crops.Additionally,the intake of alcohol and pickled foods should be controlled,and bad eating habits should be changed to prevent the occurrence of esophageal cancer.（2）The high incidence of esophageal cancer in residents of Huai’an,Jiangsu Province may be related to the high exposure of aflatoxin and nitrite in the diet.No strong association between fumonisin and esophageal cancer was found.There was no significant correlation or weak correlation between dietary and blood exposure indicators in two groups,so we need to futher explore the causes or find more effective internal exposure indicators to reflect the external exposure.（3）Dietary nitrite and blood AFB₁-Alb levels in patients with EPL were significant higher than normal controls.The oxidative stress state was also stronger than that in controls and oxidative damage was more serious.Aflatoxin and nitrite may play an important role in the development of EPL through oxidative damage.Patients with EPL may consider regulating the levels of oxidative and anti-oxidant levels by increasing the intake of exogenous antioxidants to reduce the risk of oxidative damage to the body.