The Toxic Effects Of 4-nonylphenol On Male Reproductive System Were Determined By Biospectroscopy Techniques Analysis

ObjectiveIn this study,4-nonylphenol(NP)was used as the test substance,SD male rats and testicular Sertoli cells(SCs)as the research object,bio-spectroscopy was applied to study effect of NP on rat reproductive toxicity.Methods1.The animal model was constructed by SD male rats exposed to NP every other day with a series of concentrations(25,50,100 mg/kg)10 times in total,corn oil exposure as a control.Then,testicular tissue was extracted and testicular cells were sorted for primary culture,followed with detection for biochemical changes on rat testicular tissue and related cells by Raman spectroscopy and ATR-FTIR,respectively.2.Testicular SCs were exposed to serial concentrations(2.5,5,10,and 20?M)of NP for 24 hours,DMSO as a control.Cells were assayed with ATR-FTIR after pre-treated.3.Principal component analysis and linear discriminant analysis(PCA-LDA)were used to perform variable analysis on the acquired maps.Results1.ATR-FTIR was used to detect the testicular cells of rats of different ages,and the cluster separation between NP-exposed and control groups was obvious.Except for the lipid area of the 25mg/kg NP exposure group in 50-day-old group(P>0.05),the difference between the other exposed groups and the control group was statistically significant(P<0.001),and showed a dose-effect relationship;Clustering vector analysis showed that the biomarkers changed in different NP exposures groups were different and had significant differences compared with the control group(P<0.001);There were significant differences in lipid to protein ratios,peptide aggregation levels,Amide I to Amide II ratios,and phosphate to carbohydrate ratios between different concentrations of NP exposures groups(P<0.001).2.ATR-FTIR analysis of testicular SCs cultured in vitro with different concentrations of NP exposed,NP exposure group and control group were isolated,and the effect of NP exposure group and control group was significantly different(P<0.001);Clustering vector analysis showed that the biomarkers changed in each group were different and had significant differences compared with the control group(P<0.001);The lipid-to-protein ratio,peptide aggregation level,Amide I and Amide II ratio,and phosphate-to-carbohydrate ratio between different NP exposures groups(P<0.001).3.Raman analysis of testicular tissue in rats of different ages observed far from the control group,and the difference between the different NP exposure groups and the control group was statistically significant(P<0.001);Clustering vector analysis showed that the biomarkers changed in each group were different and had significant differences compared with the control group;The lipid-protein ratio and the level of unsaturated fat in different NP exposured groups were significantly different from those of the control group(P<0.001).Conclusion1.Through in vitro and in vivo experiments,it was found that the effects of different concentrations of NP on testicular tissue and cells are different associated with dose-effect relationship;at the same time,the effects on different day-old rats are also different.2.By detecting the characteristic changes of the cell's biomolecules,it was found that NP exposure can interfere with cell metabolism such as lipid metabolism and glucose metabolism in testicular SCs and Leydig cells,affecting the testicular cell proliferation activity,and also induce changes in the secondary structure of the protein.