Douglas Follmer

The Ohio State University, Columbus, Ohio, United States

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Publications (3)10.18 Total impact

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    ABSTRACT: The endogenous vasodilator nitric oxide (NO) is metabolized in tissues in an O2 -dependent manner. In skeletal and cardiac muscle, high concentrations of myoglobin (Mb) function as a potent NO scavenger. However, Mb concentration is very low in vascular smooth muscle where low concentrations of cytoglobin (Cygb) may play a major role in metabolizing NO. Questions remain regarding how low concentrations of Cygb and Mb differ in their NO metabolism and the basis for their different cellular roles and functions. In this study, electrode techniques were applied to perform comparative measurements of the kinetics of NO consumption by Mb and Cygb. UV/Vis spectroscopic methods and computer simulations were performed to study the reaction of Mb and Cygb with ascorbate (Asc) and the underlying mechanism. It was observed that the initial rate of Cygb(3+) reduction by Asc was 415-fold greater than that of Mb(3+) . In the low [O2 ] range (0-50 μM), Cygb-mediated NO consumption rate is ~500 times more sensitive to changes in O2 concentration than that of Mb. The reduction of Cygb(3+) by Asc follows a reversible kinetic model while its reduction of Mb(3+) is irreversible. A reaction mechanism for Cygb(3+) reduction by Asc was proposed and the reaction equilibrium constants determined. Our results suggest that the rapid reduction of Cygb by cellular reductants enables Cygb to efficiently regulate NO metabolism in the vascular wall in an oxygen-dependent manner, while the slow rate of Mb reduction does not provide this oxygen dependence. This article is protected by copyright. All rights reserved.
    FEBS Journal 05/2013; 280(15). DOI:10.1111/febs.12352 · 3.99 Impact Factor
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    ABSTRACT: The endogenous vasodilator nitric oxide (NO) is metabolized in tissues in an O(2)-dependent manner. This regulates NO levels in the vascular wall; however, the underlying molecular basis of this O(2)-dependent NO consumption remains unclear. While cytoglobin (Cygb) was discovered a decade ago, its physiological function remains uncertain. Cygb is expressed in the vascular wall and can consume NO in an O(2)-dependent manner. Therefore, we characterize the process of the O(2)-dependent consumption of NO by Cygb in the presence of the cellular reductants and reducing systems ascorbate (Asc) and cytochrome P(450) reductase (CPR), measure rate constants of Cygb reduction by Asc and CPR, and propose a reaction mechanism and derive a related kinetic model for this O(2)-dependent NO consumption involving Cygb(Fe(3+)) as the main intermediate reduced back to ferrous Cygb by cellular reductants. This kinetic model expresses the relationship between the rate of O(2)-dependent consumption of NO by Cygb and rate constants of the molecular reactions involved. The predicted rate of O(2)-dependent consumption of NO by Cygb is consistent with experimental results supporting the validity of the kinetic model. Simulations based on this kinetic model suggest that the high efficiency of Cygb in regulating the NO consumption rate is due to the rapid reduction of Cygb by cellular reductants, which greatly increases the rate of consumption of NO at higher O(2) concentrations, and binding of NO to Cygb, which reduces the rate of consumption of NO at lower O(2) concentrations. Thus, the coexistence of Cygb with efficient reductants in tissues allows Cygb to function as an O(2)-dependent regulator of NO decay.
    Biochemistry 05/2012; 51(25):5072-82. DOI:10.1021/bi300291h · 3.01 Impact Factor
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    ABSTRACT: Carbon fiber microelectrodes and carbon fiber composite minielectrodes (CFM/CFCM) have been generally used for measurements of nitric oxide (NO) concentration in chemical and biological systems. The response time of a CFM/CFCM is usually from milliseconds to seconds depending on the electrode size, the thickness of coating layers on the electrode, and NO diffusion coefficients of the coating layers. As a result, the time course of recoded current changes (I-t curves) by the CFM/CFCM may be different from the actual time course of NO concentration changes (c-t curves) if the half-life of NO decay is close to or shorter than the response time of the electrode used. This adds complexity to the process for determining rate constants of NO decay kinetics from the recorded current curves (I-t curves). By computer simulations based on a mathematical model, an approximation method was developed for determining rate constants of NO decay from the recorded current curves. This method was first tested and valuated using a commercial CFCM in several simple reaction systems with known rate constants. The response time of the CFCM was measured as 4.7±0.7 s (n=5). The determined rate constants of NO volatilization and NO autoxidation in our measurement system at 37 °C are (1.9±0.1)×10(-3) s(-1) (n=4) and (2.0±0.3)×10(3) M(-1) s(-1) (n=7), which are close to the reported rate constants. The method was then applied to determine the rate of NO decay in blood samples from control and smoking exposed mice. It was observed that the NO decay rate in the smoking group is >20% higher than that in control group, and the increased NO decay rate in the smoking group was reversed by 10 μM diphenyleneiodonium chloride (DPI), an inhibitor of flavin enzymes such as leukocyte NADPH oxidase.
    Nitric Oxide 12/2010; 23(4):311-8. DOI:10.1016/j.niox.2010.09.002 · 3.18 Impact Factor