Streptococcus mutans Binding to Collagen and Fibrinogen in Nicotine

Abstract

Introduction: Our overall goal is to find the mechanism for atherosclerosis. Smokers have increased incidence of atherosclerosis. Atherosclerosis occurs when there is a build up of plaque in the arteries. There is evidence that Streptococcus mutans help cause this blockage. We have already proven that S. mutans produces more biofilm in certain concentrations of nicotine. Also, we have found that nicotine upregulates S. mutans binding to proteins in certain concentrations; other labs have also demonstrated this. The intent of this study was to evaluate the binding of S. mutans to both collagen type I and fibrinogen, which are both proteins that are already present on the surface of endothelial cells lining arteries. Methods: S. mutans UA159 was cultured in 0.00-4.00 mg/mL nicotine. The cells were killed in formaldehyde and then coated with biotin. The proteins studied were plated (1 ug/ml) on 96-well microtiter plates. In order to block the empty spaces that the protein did not bind to, 1% BSA in sodium bicarbonate buffer was added to the plate. Each nicotine dilution of S. mutans was added to the plate and the amount of binding was assessed. Extra-avidin HRP and OPD were added to the plate and the intensity was measured at an absorbance of 490 nm using a spectrophotometer. Results: The intensity was directly related to the number of cells bound to the proteins. There was a significant increase in S. mutans binding when compared to the baseline for both collagen type I and fibrinogen. The binding was highest when S. mutans were cultured in 2 and 4 mg/mL nicotine. Conclusions: The data collected suggests that collagen type I and fibrinogen contribute to the mechanism of atherosclerosis. When S. mutans are cultured in moderately high concentrations of nicotine, more binding of the bacteria to these proteins occurs.