Multiple generation distinct toxicant exposures induce epigenetic transgenerational inheritance of enhanced pathology and obesity

Abstract

Three successive multiple generations of rats were exposed to different toxicants and then bred to the transgenerational F5 generation to assess the impacts of multiple generation different exposures. The current study examines the actions of the agricultural fungicide vinclozolin on the F0 generation, followed by jet fuel hydrocarbon mixture exposure of the F1 generation, and then pesticide dichlorodiphenyltrichloroethane on the F2 generation gestating females. The subsequent F3 and F4 generations and F5 transgenerational generation were obtained and F1-F5 generations examined for male sperm epigenetic alterations and pathology in males and females. Significant impacts on the male sperm differential DNA methylation regions were observed. The F3-F5 generations were similar in ∼50% of the DNA methylation regions. The pathology of each generation was assessed in the testis, ovary, kidney, and prostate, as well as the presence of obesity and tumors. The pathology used a newly developed Deep Learning, artificial intelligence-based histopathology analysis. Observations demonstrated compounded disease impacts in obesity and metabolic parameters, but other pathologies plateaued with smaller increases at the F5 transgenerational generation. Observations demonstrate that multiple generational exposures, which occur in human populations, appear to increase epigenetic impacts and disease susceptibility.

Keywords: DDT; Epigenetic; inheritance; jet fuel; obesity; pathology; review; sperm; testis; transgenerational; vinclozolin.

Figure 1:

DMR identification. The number of DMRs found using different P-value cutoff thresholds. The All-Window column shows all DMRs. The Multiple Window column shows the number of DMRs containing at least two nearby significant windows (1 kb each). The number of DMRs with the number of significant windows (1 kb per window) at a P-value threshold of P < 1e-04 for DMR is bolded. (A) Experimental design; (B) F1 generation 1e-04; (C) F2 generation 1e-04; (D) F3 generation 1e-04; (E) F4 generation 1e-04; (F) F5 generation 1e-04

Figure 2:

DMR chromosomal locations. (A) F1 generation 1e-04; (B) F2 generation 1e-04; (C) F3 generation 1e-04; (D) F4 generation 1e-04; (E) F5 generation 1e-04

Figure 3:

DMR CpG density. (A) F1 CpG density; (B) F2 CpG density; (C) F3 CpG density; (D) F4 CpG density; (E) F5 CpG density

Figure 4:

DMR length. (A) F1 DMR length; (B) F2 DMR length; (C) F3 DMR length; (D) F4 DMR length; (E) F5 DMR length

Figure 5:

DMR overlaps. (A) DMR overlap P < 10-4 with P < 0.05 in other generations in male sperm; (B) DMR overlap P < 10-4 male sperm; and (C) extended overlap for F5 generation (D) extended overlap DMRs for F1 generation, and (E) DMR single exposure overlaps

Figure 6:

DMR gene association pathway analysis. (A) Number of DMR-associated genes in each gene category. (B) Gene pathway analysis. KEGG pathway identifier numbers and names are presented. The number of DMR-associated genes that fall into each pathway is indicated in parentheses ()

Figure 7:

pathology. (A) Testis disease; (B) prostate disease; (C) prostate disease; (D) male kidney disease; (E) ovary disease; (F) testis pathologies DL results; (G) prostate pathologies DL results; (H) male kidney individual pathologies DL results; (I) female kidney pathologies DL results; and (J) ovary pathologies DL results. Statistical differences were determined by Fisher’s exact tests. F3, F4, and F5 are compared to F3 Control. * P < 0.05. ** P < 0.01

Figure 8:

DL individual tissue sub-pathologies. (A) DL testis individual pathologies; (B) DL prostate individual pathologies; (C) DL male kidney individual pathologies; (D) DL female kidney individual pathologies; and (E) ovary individual pathologies. Statistical differences were determined by Fisher’s exact tests. * =P < 0.05. *** =P < 0.001

Figure 9:

Multigen study other pathologies. (A) Male tumors; (B) female tumors; (C) multiple disease (male); (d) multiple disease (female); (E) DL pathology for multiple disease (male); and (F) DL pathology for multiple disease (female). Statistical differences were determined by Fisher’s exact tests. F3, F4, and F5 are compared to F3 control. * P < 0.05. *** P < 0.001

Figure 10:

Obesity parameters. (A) Average weight in multigen and control females; (B) Average weight in multigen and control males; (C) Average BMI in multigen and control females; (D) Average BMI in multigen and control males; (E) Average adipocyte area in multigen and control females; (F) Average adipocyte area in multigen and control males; (G) Obesity disease in females; and (H) Obesity disease in males. Statistical differences determined by Student’s t-test for A–F. Statistical differences were determined by Fisher’s exact tests for G and H. F1 compared to F1 Control. F3, F4, and F5 are compared to F3 Control. * P < 0.05. ** P < 0.01. *** P < 0.001