| Lead is widespread in the environment and harmful for human.With the development of modern industry,lead is extensively used.Occupational lead poisoning is becoming one of the common occupational diseases,so prevention of the occurrence is particularly important.The studies on lead genetic toxicity have now been finalized: Study shows that lead has a clear genetic toxicity,high concentrations have a direct mutagenesis and the role of performance for the fracturing(clastogenesis),low concentrations can interfere with DNA damage repair and the synergistic mutagenesis .On the results there are not only animal experiments.Detection DNA damage of peripheral blood cell in occupational lead exposure worker with comet assay has been reported in the literature.Traditional method that detects DNA damage is single-cell gel electrophoresis assay(comet assay).Single cell gel electrophoresis assay is the final outcome that a cell response to toxic substances on the genetic,not the early,sensitive reflection of the genetic.toxicity substances on DNA of the cell.The operation of large samples in the crowd,there are some limitations.So it needs to find a more objective,simple and effective method to detect large samples in the crowd means.γH2AX is a hot research of stress responses in the DNA damage cell,after DNA double-strand breaks occurred,the histone H2A family member,H2AX,serine 139 residue phosphorylate rapidly,formingγH2AX,at fracture sites collecting damage repair-related proteins cluster to form the focus.This process can happen in minutes.γH2AX focus formation in fluorescence can be clearly discerned under the microscope. Subsequently,γH2AX at DNA double-strand break damage play a role in the repair process.Research proves that the number ofγH2AX focus formation and the number of DSBs was on one-to-one relationship.γH2AX focus formation and disappearance of this process with the formation and repair of DSBs must have relevance,flow cytometer(FCM) assay is considered a sensitive to detection of a valid indicator of DSBs.It is a new technology that detects DNA damage repair,cell cycle regulation.It is a very useful research tool with rapid,objective advantages.This topic is to detect DNA damage in occupational workers exposed to lead. Exposure group come from workers of a battery factory.As the lead pollution are primarily caused through lead dust,lead smoke,and affect water,soil,food near the factory,increasing the lead content.Besides workers non-exposure to lead of the battery factory are few.It is little significant to select control group in the factory.This topic is to select island crowd as control group outside factory,to survey DNA damage of peripheral blood lymphocytes in island crowd.At the same time,to analyze related factors of DNA damage in island crowd.Objective1.To study whether lead exposure can introduce DNA damage with flow cytometer (FCM) assay.To explore genetic toxicity of occupational lead exposure by the comparison of DNA damage in different exposure levels,different labor types, different labor ages.2.To study DNA damage in the island crowd with flow cytometer assay,through analysis of different gender,age,smoking,drinking and other related factors,to explore DNA damage in the island crowd with above mentioned related factors.Methods1.Exposure group come from 41 cases of a battery factory workers.Control group come from 41 cases of the island crowd non-occupational exposure to harmful factors. To investigate object's age,sex,smoking,drinking and the general epidemiological data.To obtain blood lead,urine lead,d-ALA and other biological information from medical reports of 41 workers exposed to lead.At the same time to investigate the battery factory environment,testing the air lead levels. 2.Each object collects blood about 3ml with heparin tube,preserved by mobile refrigerator to laboratory.Blood:human lymphocyte separation medium = 1:1,the horizontal centrifuge:2000r~*30min,lymphocytes are separated.Samples can be preserved at -20℃after being fixed with 2ml 75%ice ethanol,divided into two pipes. When samples are detected,the centrifuge:2000r~* 10min,to suck out the supernatant, then samples are closed 2h with 100μl goat serum working solution at room temperature.After being closed,with PBS washed once,samples were incubated 2h with FITC labeled anti-γH2AX antibody(1:500) at room temperature away from light. With PBS washed once,samples are made into suspension with 400μlPBS.Then samples are tested by the flow cytometer.3.To analyze DNA damage with age,gender,smoking,drinking,blood lead,urine lead,d-ALA between exposure group and control group.To explore genetic toxicity of lead and the relation between DNA damage and above mentioned related factors in the crowd non-occupational exposure to harmful factors.Results1.Test of air lead concentration in battery factoryThe test results of air lead concentration in battery factory showed that only in the welding group(high concentration group of lead exposure) the air lead concentration exceed the national occupational exposure limits,in bagging group(also named packing group),weighting group,sealing group,big and small boxing group,fitting group,electric group(middle and low concentration group of lead exposure),the air lead concentration are below the national occupational exposure limits.2.Detection of occupational lead exposure workers(1)DNA damage rate and geometric mean fluorescence intensity in occupational group was significantly higher than that in control group(p＜0.05).(2)There was no statistical significance of the difference on DNA damage rate and geometric mean fluorescence intensity between different age groups(p＞0.05).(3)There was no statistical significance with multiple linear regression analysis between DNA damage rate and blood lead,urine lead,d-ALA(p＞0.05).There was also no statistical significance with multiple linear regression analysis between geometric mean fluorescence intensity and blood lead,urine lead,d-ALA(p＞0.05).(4) DNA damage rate and blood lead,d-ALA of high concentration group were positive correlation with partial correlation analysis,geometric mean fluorescence intensity and blood lead of high concentration group was positive correlation with partial correlation analysis,and the correlation coefficient r had statistical significance(p＜0.05).3.Survey of DNA damage rate in island crowd(1)The difference of geometric mean fluorescence intensity between different gender groups in the island crowd was significant(p＜0.05).(2)The difference of geometric mean fluorescence intensity in different age groups in the island crowd was significant(p＜0.05).(3)The difference of geometric mean fluorescence intensity in different drinking history groups in the island crowd was significant(p＜0.05).(4)The difference of DNA damage and geometric mean fluorescence intensity in different smoking history groups in the island crowd was not significant(p＞0.05).(5) To analyze DNA damage,geometric mean fluorescence intensity and gender,age, smoking,drinking with partial correlation analysis,the results showed that DNA damage,geometric mean fluorescence intensity and drinking history in the island crowd were positive correlation with partial correlation analysis,and the correlation coefficient r had statistical significance(p＜0.05).Conclusions1.Detection of occupational lead exposure workersOccupational lead exposure can cause DNA damage.DNA damage rate and geometric mean fluorescence intensity in occupational group was significantly higher than that in control group(p＜0.05).But there was no statistical significance of the difference on DNA damage rate and geometric mean fluorescence intensity between different age groups.To analyze DNA damage rate,geometric mean fluorescence intensity and blood lead,urine lead,d-ALA with multiple linear regression analysis,the results showed that there was no statistical significance(p＞0.05).DNA damage rate and blood lead,d-ALA of high concentration group were positive correlation with partial correlation analysis,geometric mean fluorescence intensity and blood lead of high concentration group was positive correlation with partial correlation analysis,and the correlation coefficient r had statistical significance(p＜0.05).2.Survey of DNA damage rate in island crowdγH2AX flow cytometer assay is a sensitive,objective and effective method of peripheral blood lymphocytes DNA damage.The differences of geometric mean fluorescence intensity in different age groups,different drinking history groups, different smoking history groups were significant,but not to DNA damage rate.To analyze DNA damage,geometric mean fluorescence intensity and gender,age, smoking history,drinking history with partial correlation analysis,the results showed that DNA damage,geometric mean fluorescence intensity and drinking history in the island crowd were positive correlation with partial correlation analysis,and the correlation coefficient r had statistical significance(p＜0.05).
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