Effects of cigarette smoke borne reactive nitrogen species on salivary alpha-amylase activity and protein modifications.

Cigarette smoke (CS) is associated with a variety of human pathologies including cardiovascular disease and cancer. Oral squamous cell carcinoma (OSCC) is the most common malignancy of the head and neck. The major inducer of OSCC is exposure to tobacco. Recent studies demonstrated that oxidative and nitrosative stress contributes to the development of oral carcinogenesis through DNA damage. All salivary reactive nitrogen species (RNS) analyzed from OSCC patients are significantly higher in comparison with healthy subjects. Our findings show that CS and external RNS addition induced reduction in alpha-amylase activity and produced some excited carbonyl formation, but to a much less extant than CS. The addition of epigallocatechine-3-gallate (EGCG) to saliva produced no protective effect against damage to alpha-amylase activity. Our proposed mechanism for the decrease in alpha-amylase activity is the formation of adducts at SH groups of the alpha-amylase active site. In this case, EGCG was unable to counteract this phenomenon, as it does not reduce the concentration of disulfides, and does not alter the amount of protein-SH moieties. However, EGCG did reduce the levels of excited carbonyl formation. Our results indicate that although RNS are abundant in CS, a significant decrease in amylase activity is due to other components in CS, probably aldehydes, reacting with the thiol group of proteins by the Michael addition reaction.