Gender-Dependent Mechanisms of Alpha-Tocopherol Protection from Benzo[a]pyrene Exposure in Rats

Polycyclic aromatic hydrocarbons (PAHs), including benzo[a]pyrene (BP), are environmental pollutants linked to increased disease susceptibilities. Alpha-Tocopherol (aT) supplementation decreases BP-DNA adducts in smokers, particularly women; but the mechanism is unknown. To test the hypothesis that aT protection from BP exposure is gender-dependent, male and female rats received 7 daily subcutaneous (SQ) injections of aT (100 mg aT/ kg body wt) or vehicle, followed by a single ip injection of BP (20 mg/kg, spiked with 3H-BP) on day 9. Urine and bile were collected pre- and post-BP. Plasma and tissues were collected 5 or 24 hr post-BP. aT supplementation increased aT levels in females greater than males. Compared to vehicle, aT supplementation increased total urinary and biliary excretion of BP and/or BP metabolites more than 2.5-fold in females, but decreased total BP and/or BP metabolite excretion in males (p<0.05). SQ aT prevented BP-induced increases in urine 8-isoprostanes (males) and decreased tissue malondialdehyde levels in a tissue- and gender-dependent manner. These data are the first to suggest that aT protection from BP exposure is gender-dependent and occurs by both antioxidant and non-antioxidant mechanisms. Further elucidation of the mechanism(s) of aT protection against PAHs may lead to the development of novel protective strategies for occupational PAH exposures.Thesis poster fair presentation.Keywords: Xenobiotic, Polycyclic aromatic hydrocarbon, Benzo[a]pyrene, Alph-tocopherol, Vitamin E, Gende

The A31P missense mutation in cardiac myosin binding protein C alters protein structure but does not cause haploinsufficiency

Mutations in MYBPC3, the gene encoding cardiac myosin binding protein C (cMyBP-C), are a major cause of hypertrophic cardiomyopathy (HCM). While most mutations encode premature stop codons, missense mutations causing single amino acid substitutions are also common. Here we investigated effects of a single proline for alanine substitution at amino acid 31 (A31P) in the C0 domain of cMyBP-C, which was identified as a natural cause of HCM in cats. Results using recombinant proteins showed that the mutation disrupted C0 structure, altered sensitivity to trypsin digestion, and reduced recognition by an antibody that preferentially recognizes N-terminal domains of cMyBP-C. Western blots detecting A31P cMyBP-C in myocardium of cats heterozygous for the mutation showed a reduced amount of A31P mutant protein relative to wild-type cMyBP-C, but the total amount of cMyBP-C was not different in myocardium from cats with or without the A31P mutation indicating altered rates of synthesis/degradation of A31P cMyBP-C. Also, the mutant A31P cMyBP-C was properly localized in cardiac sarcomeres. These results indicate that reduced protein expression (haploinsufficiency) cannot account for effects of the A31P cMyBP-C mutation and instead suggest that the A31P mutation causes HCM through a poison polypeptide mechanism that disrupts cMyBP-C or myocyte function