| Objective:The purpose of this study was to explore a pre-treatment method for the detection of small amounts of dioxins in the serum using a bioassay based on the CBG2.8D cell line,and to complete the detection of serum samples for preliminary understanding of its suitability;There was a significant difference in the expression levels of three apolipoproteins in the serum of people exposed to dioxin-like compounds compounds,and the relationship between exposure levels and protein expression was discussed.Methods:1.The sera of rats exposed to different concentrations of TCDD were used to investigate the pretreatment method,and multiple experiments were conducted to establish a stable and effective biological detection method suitable for serum samples(CBG2.8D).After finding a suitable test procedure,a serum sample from a group of seven chlorinated workers in Tianjin was selected as the experimental subject.The test was performed using the CBG2.8D bioassay method,and the results were compared with the obtained high-resolution gas chromatograms.High resolution mass spectrometer(HRGC-HRMS)analysis results were compared.2.Using Western Blot technology to verify the three apolipoproteins with significant differences in proteomics data and discuss the trend of expression changes.Results:1.The experiment found that for 0.5 mL of high dioxin concentration rat serum,the organic solvent extraction method of n-hexane and isopropanol was used in the purification and concentration of the acidic silica gel column to carry out pretreatment and the CBG2.8D bioassay was more appropriate.Bioassay detection results of human serum samples differ significantly from high resolution results.2.Western Blot's validation results are consistent with the results of serum proteomics studies.The expression level of ApoCI protein in the T1 group was 66% higher than that in the C1 group,the expression level in the T2 group was 22% higher than that in the C1 group,and the expression level in the T3 group was 82% higher than that in the C1 group,and the difference between the groups was statistically significant.(P<0.05);The expression level of ApoCIII protein in T1 group was 45% lower than that in C1 group,the expression level in T2 group was 62% lower than that in C1 group,and the expression level in T3 group was 5% lower than that in C1 group.All had statistical significance(P<0.05);the expression level of ApoCIV protein in the T1 group was 14.57 times that in the C1 group,the expression level in the T2 group was 11.0 times that in the C1 group,and the expression level in the T3 group was 14.84% in the C1 group.Times,the differences between the groups were statistically significant(P<0.05);The ApoCI,ApoCIII,and ApoCIV all showed lower exposure and higher exposure in the three exposed groups than in the medium exposure group.In the exposed and clean control groups,the expression of ApoCI and ApoCIV in the exposure group was higher than the control group.In the group,the expression level of ApoCIII was lower in the exposure group than in the control group.Conclusion:1.For a small amount of high dioxins rat serum samples,a suitable pretreatment method for CBG2.8D bioassay is the organic solvent extraction method of n-hexane and isopropanol combined with the purification and concentration method of an acidic silica gel column.Its test results are similar to those of high-resolution detection methods.For serum samples from chlorinated plant workers,the detection results of the CBG2.8D bioassay method and the HRGC-HRMS high resolution method showed significant differences and could not be considered to be from the same population.Speculation reason:(1)Certain non-dioxin-like compounds in serum also activate the aryl hydrocarbon receptor pathway to respond,because the detection principle of CBG2.8D bioassay depends on the activation of AhR pathway,so the result is directly reflected in the response degree of AhR pathway.,While the rat's culture and exposure to the poisonous environment is basically completely controllable,but the living environment of the chlorinated plant workers is more complex,the exposure factors are not completely controllable,and the serum may contain other substances that may activate the AhR signaling pathway,leading to The results of high-resolution detection of total TEQ and HRGC-HRMS in the serum derived from the CBG2.8D bioassay method are quite different.(2)Due to the low human serum sample size(0.5 mL)used in this study,repeated experiments may not be repeated multiple times,which may lead to a relatively large bias,leading to no correlation in the test results.If the number of test samples is further increased,the test results may improve.2.Different levels of serum dioxin concentrations affect the expression levels of ApoCI,ApoCIII,and ApoCIV.The expression of these three apolipoproteins is associated with the occurrence of various lipid metabolism-related diseases,and it can be speculated that serum dioxins The concentration of the compound affects the incidence of certain lipid metabolism related diseases. |
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