Superoxide, hydroxyl radical, and hydrogen peroxide effects on single-diaphragm fiber contractile apparatus

Reactive oxygen species contribute to diaphragm dysfunction in certain pathophysiological conditions (i.e., sepsis and fatigue). However, the precise alterations induced by reactive oxygen species or the specific species that are responsible for the derangements in skeletal muscle function are incompletely understood. In this study, we evaluated the effect of the superoxide anion radical (O₂/^-·), hydroxyl radical (·OH), and hydrogen peroxide (H₂O₂) on maximum calcium-activated force (F_max) and calcium sensitivity of the contractile apparatus in chemically skinned (Triton X-100) single rat diaphragm fibers. O₂/^-· was generated using the xanthine/xanthine oxidase system; ·OH was generated using 1 mM FeCl₂, 1 mM ascorbate, and 1 mM H₂O₂; and H₂O₂ was added directly to the bathing medium. Exposure to O₂/^-· or ·OH significantly decreased F_max by 14.5% (P < 0.05) and 43.9% (P < 0.005), respectively. ·OH had no effect on Ca²⁺ sensitivity. Neither 10 nor 1,000 μM H₂O₂ significantly altered F_max or Ca²⁺ sensitivity. We conclude that the diaphragm is susceptible to alterations induced by a direct effect of ·OH and O₂/^-·, but not H₂O₂, on the contractile proteins, which could, in part, be responsible for prolonged depression in contractility associated with respiratory muscle dysfunction in certain pathophysiological conditions.