Oxidative stress and Ca²⁺toxicity are mechanisms of hypoxic–ischemic (HI) brain injury. This work investigates if partial inhibition of mitochondrial respiratory chain protects HI brain by limiting a generation of oxidative radicals during reperfusion. HI insult was produced in p10 mice treated with complex I (C-I) inhibitor, pyridaben, or vehicle. Administration of P significantly decreased the extent of HI injury. Mitochondria isolated from the ischemic hemisphere in pyridaben-treated animals showed reduced H₂O₂emission, less oxidative damage to the mitochondrial matrix, and increased tolerance to the Ca²⁺-triggered opening of the permeability transition pore. A protective effect of pyridaben administration was also observed when the reperfusion-driven oxidative stress was augmented by the exposure to 100% O₂which exacerbated brain injury only in vehicle-treated mice.In vitro, intact brain mitochondria dramatically increased H₂O₂emission in response to hyperoxia, resulting in substantial loss of Ca²⁺buffering capacity. However, in the presence of the C-I inhibitor, rotenone, or the antioxidant, catalase, these effects of hyperoxia were abolished.Our data suggest that the reperfusion-driven recovery of C-I-dependent mitochondrial respiration contributes not only to the cellular survival, but also causes oxidative damage to the mitochondria, potentiating a loss of Ca²⁺buffering capacity. This highlights a novel neuroprotective strategy against HI brain injury where the major therapeutic principle is a pharmacological attenuation, rather than an enhancement of mitochondrial oxidative metabolism during early reperfusion.

This study investigated whether mitochondrial dysfunction contributes to alveolar developmental arrest in a mouse model of bronchopulmonary dysplasia (BPD). To induce BPD, 3-day-old mice were exposed to 75% O2. Mice were studied at two time points of hyperoxia (72 h or 2 wk) and after 3 weeks of recovery in room air (RA). A separate cohort of mice was exposed to pyridaben, a complex-I (C-I) inhibitor, for 72 hours or 2 weeks. Alveolarization was quantified by radial alveolar count and mean linear intercept methods. Pulmonary mitochondrial function was defined by respiration rates, ATP-production rate, and C-I activity. At 72 hours, hyperoxic mice demonstrated significant inhibition of C-I activity, reduced respiration and ATP production rates, and significantly decreased radial alveolar count compared with controls. Exposure to pyridaben for 72 hours, as expected, caused significant inhibition of C-I and ADP-phosphorylating respiration. Similar to hyperoxic littermates, these pyridaben-exposed mice exhibited significantly delayed alveolarization compared with controls. At 2 weeks of exposure to hyperoxia or pyridaben, mitochondrial respiration was inhibited and associated with alveolar developmental arrest. However, after 3 weeks of recovery from hyperoxia or 2 weeks after 72 hours of exposure to pyridaben alveolarization significantly improved. In addition, there was marked normalization of C-I and mitochondrial respiration. The degree of hyperoxia-induced pulmonary simplification and recovery strongly (r2 = 0.76) correlated with C-I activity in lung mitochondria. Thus, the arrest of alveolar development induced by either hyperoxia or direct inhibition of mitochondrial oxidative phosphorylation indicates that bioenergetic failure to maintain normal alveolar development is one of the fundamental mechanisms responsible for BPD.

Mitochondrial dysfunction is the most fundamental mechanism of cell damage in cerebral hypoxia-ischemia and reperfusion. Mitochondrial respiratory chain (MRC) is increasingly recognized as a source for reactive oxygen species (ROS) in the postischemic tissue. Potentially, ROS originating in MRC can contribute to the reperfusion-driven oxidative stress, promoting mitochondrial membrane permeabilization. The loss of mitochondrial membranes integrity during reperfusion is considered as the major mechanism of secondary energy failure. This paper focuses on current data that support a pathogenic role of ROS originating from mitochondrial respiratory chain in the promotion of secondary energy failure and proposes potential therapeutic strategy against reperfusion-driven oxidative stress following hypoxia-ischemia-reperfusion injury of the developing brain.

Hypoxic–ischemic (HI) brain injury in infants is a leading cause of lifelong disability. We report a novel pathway mediating oxidative brain injury after hypoxia–ischemia in which C1q plays a central role. Neonatal mice incapable of classical or terminal complement activation because of C1q or C6 deficiency or pharmacologically inhibited assembly of membrane attack complex were subjected to hypoxia–ischemia. Only C1q^−/−mice exhibited neuroprotection coupled with attenuated oxidative brain injury. This was associated with reduced production of reactive oxygen species (ROS) in C1q^−/−brain mitochondria and preserved activity of the respiratory chain. Compared with C1q^+/+neurons, cortical C1q^−/−neurons exhibited resistance to oxygen–glucose deprivation. However, postischemic exposure to exogenous C1q increased both mitochondrial ROS production and mortality of C1q^−/−neurons. This C1q toxicity was abolished by coexposure to antioxidant Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid). Thus, the C1q component of complement, accelerating mitochondrial ROS emission, exacerbates oxidative injury in the developing HI brain. The terminal complement complex is activated in the HI neonatal brain but appeared to be nonpathogenic. These findings have important implications for design of the proper therapeutic interventions against HI neonatal brain injury by highlighting a pathogenic priority of C1q-mediated mitochondrial oxidative stress over the C1q deposition-triggered terminal complement activation.

Background and Purpose— This study was undertaken to determine whether the initial component of the classical complement (C) activation pathway contributes to hypoxic-ischemic brain injury in neonatal mice. Methods— Hypoxia-ischemia (HI) was produced in C1q ^−/− and wild-type (WT) neonatal mice. At 24 hours after HI, neonatal mouse reflex performance and cerebral infarct volume were assessed. Long-term outcomes were measured by water-maze performance and degree of cerebral atrophy at 7 to 8 weeks after HI. Activation of circulating neutrophils, and C1q, C3, and neutrophil deposition in brains were examined. Results— C1q ^−/− mice were significantly protected against HI (mean±SE infarct volume in C1q ^−/− mice=17.3±5.5% versus 53.6±6.8% in WT mice; P <0.0001) and exhibited significantly less neurofunctional deficit compared with WT mice. Immunostaining revealed significantly greater deposition of C3 (and C1q) as well as granulocytes in the infarcted brains in WT mice compared with C1q ^−/− animals. Activation of circulating leukocytes was significantly decreased in C1q ^−/− mice compared with WT mice, which correlated strongly ( r =0.7) with cerebral infarct volumes. Conclusions— Cerebral deposition of C1q and C3 after hypoxic-ischemic insult is associated with significantly greater neurologic damage in WT mice compared with C1q ^−/− mice, providing strong evidence that the classical C pathway contributes to the hypoxic-ischemic brain injury. Significantly decreased activation of circulating neutrophils associated with diminished local accumulation and attenuation of brain injury in C1q ^−/− mice suggests a potential cellular mechanism by which C1q mediates neurodegeneration in HI.

Establishing sustained reoxygenation/reperfusion ensures not only the recovery, but may initiate a reperfusion injury in which oxidative stress plays a major role. This study offers the mechanism and this mechanism-specific therapeutic strategy against excessive release of reactive oxygen species (ROS) associated with reperfusion-driven recovery of mitochondrial metabolism. In neonatal mice subjected to cerebral hypoxia-ischaemia (HI) and reperfusion, we examined conformational changes and activity of mitochondrial complex I with and without post-HI administration of S-nitrosating agent, MitoSNO. Assessment of mitochondrial ROS production, oxidative brain damage, neuropathological and neurofunctional outcomes were used to define neuroprotective strength of MitoSNO. A specificity of reperfusion-driven mitochondrial ROS production to conformational changes in complex I was examined in-vitro. HI deactivated complex I, changing its conformation from active form (A) into the catalytically dormant, de-active form (D). Reperfusion rapidly converted the D-form into the A-form and increased ROS generation. Administration of MitoSNO at the onset of reperfusion, decelerated D→A transition of complex I, attenuated oxidative stress, and significantly improved neurological recovery. In cultured neurons, after simulated ischaemia-reperfusion injury, MitoSNO significantly reduced ROS generation and neuronal mortality. In isolated mitochondria subjected to anoxia-reoxygenation, MitoSNO restricted ROS release during D→A transitions. Rapid D→A conformation in response to reperfusion reactivates complex I. This is essential not only for metabolic recovery, but also contributes to excessive release of mitochondrial ROS and reperfusion injury. We propose that the initiation of reperfusion should be followed by pharmacologically-controlled gradual reactivation of complex I.