Reactive Oxygen Species in Unstimulated Hemocytes of the Pacific Oyster Crassostrea gigas: A Mitochondrial Involvement

Abstract

The Pacific oyster Crassostrea gigas is a sessile bivalve mollusc whose homeostasis relies, at least partially, upon cells circulating in hemolymph and referred to as hemocytes. Oyster’s hemocytes have been reported to produce reactive oxygen species (ROS), even in absence of stimulation. Although ROS production in bivalve molluscs is mostly studied for its defence involvement, ROS may also be involved in cellular and tissue homeostasis. ROS sources have not yet been described in oyster hemocytes. The objective of the present work was to characterize the ROS sources in unstimulated hemocytes. We studied the effects of chemical inhibitors on the ROS production and the mitochondrial membrane potential (Δψm) of hemocytes. First, this work confirmed the specificity of JC-10 probe to measure Δψm in oyster hemocytes, without being affected by ΔpH, as reported in mammalian cells. Second, results show that ROS production in unstimulated hemocytes does not originate from cytoplasmic NADPH-oxidase, nitric oxide synthase or myeloperoxidase, but from mitochondria. In contrast to mammalian cells, incubation of hemocytes with rotenone (complex I inhibitor) had no effect on ROS production. Incubation with antimycin A (complex III inhibitor) resulted in a dose-dependent ROS production decrease while an over-production is usually reported in vertebrates. In hemocytes of C. gigas, the production of ROS seems similarly dependent on both Δψm and ΔpH. These findings point out differences between mammalian models and bivalve cells, which warrant further investigation about the fine characterization of the electron transfer chain and the respective involvement of mitochondrial complexes in ROS production in hemocytes of bivalve molluscs.

Antimicrobial Histones and DNA Traps in Invertebrate Immunity

“…7H), which results from NADPH-oxidase activity and/or mitochondrial respiration (32). Indeed, we observed a strong positive correlation between ROS production and ET formation (Figs.…”

Rapid mitochondrial adjustments in response to short-term hypoxia and re-oxygenation in the Pacific oysterCrassostrea gigas

“…However, even in very well studied plant models, it is difficult to quantify their relative contributions using these inhibitors (Njogu et al, 1980;Bingham and Stevenson, 1995). In preliminary trials, the use of SHAM to inhibit AOX in C. gigas was not conclusive, and might be due to a non-specific effect of SHAM on CCO (Tschischka et al, 2000), or to AOX in oysters that would be not sensitive to SHAM (Donaghy et al, 2012). In the present study, we blocked the respiratory chain at complex III with antimycin A to assess the potential contribution of AOX to mitochondrial respiration.…”

Low hydrogen peroxide production in mitochondria of the long‐lived Arctica islandica: underlying mechanisms for slow aging

“…AOX is a dimer located in the inner mitochondrial membrane that transfers electrons from reduced quinone (coenzyme Q) to oxygen, thus bypassing CIII, without participating in proton pumping (van Dongen et al, 2011). Although AOX are cyanide-and antimycin-insensitive, they are specifically inhibited by salicylhydroxamic acid (SHAM) (reviewed in Lenaz & Genova, 2010).…”

“…Their affinity for oxygen is much lower than CIV, and their recruitment requires a high degree of reduction of the Q-pool. Because of these last two characteristics, AOX is thought to represent a simple mechanism that protects the organism against conditions known to elicit high ROS production rates; this would be achieved at the expense of maintaining futile proton cycling (Abele et al, 2007;Donaghy et al, 2012). For instance, AOX mRNA levels have been found to be upregulated after 12 and 24 h of hypoxia and during reoxygenation (Sussarellu et al, 2013).…”