Cell biology of ischemia/reperfusion injury.

Disorders characterized by ischemia/reperfusion (I/R), such as myocardial infarction, stroke, and peripheral vascular disease, continue to be among the most frequent causes of debilitating disease and death. Tissue injury and/or death occur as a result of the initial ischemic insult, which is determined primarily by the magnitude and duration of the interruption in the blood supply, and then subsequent damage induced by reperfusion. During prolonged ischemia, ATP levels and intracellular pH decrease as a result of anaerobic metabolism and lactate accumulation. As a consequence, ATPase-dependent ion transport mechanisms become dysfunctional, contributing to increased intracellular and mitochondrial calcium levels (calcium overload), cell swelling and rupture, and cell death by necrotic, necroptotic, apoptotic, and autophagic mechanisms. Although oxygen levels are restored upon reperfusion, a surge in the generation of reactive oxygen species occurs and proinflammatory neutrophils infiltrate ischemic tissues to exacerbate ischemic injury. The pathologic events induced by I/R orchestrate the opening of the mitochondrial permeability transition pore, which appears to represent a common end-effector of the pathologic events initiated by I/R. The aim of this treatise is to provide a comprehensive review of the mechanisms underlying the development of I/R injury, from which it should be apparent that a combination of molecular and cellular approaches targeting multiple pathologic processes to limit the extent of I/R injury must be adopted to enhance resistance to cell death and increase regenerative capacity in order to effect long-lasting repair of ischemic tissues.

Effects of sildenafil and tadalafil on ischemia/reperfusion injury in fetal rat brain.

OBJECTIVE: The aim of the present study was to evaluate the effects of phosphodiesterase type 5 (PDE5) inhibitory drugs, sildenafil and tadalafil, in ischemia/reperfusion (I/R)-induced oxidative injury in fetal rat brain. METHODS: Timed pregnant adult Wistar rats were randomly assigned to the following groups (n = 6 for each group): saline + none I/R (1), saline + I/R (2), sildenafil + none I/R (3); sildenafil + I/R (4), tadalafil + none I/R (5) and tadalafil + I/R (6). Fetal ischemia was induced by clamping the utero-ovarian artery bilaterally. Fetuses were delivered and 268 fetal rats were decapitated. Malondialdehyde (MDA) levels and, superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were assessed in fetal brain tissue homogenates by spectrophotometric methods. RESULTS: In saline + I/R group, MDA levels were increased and, SOD and GSH-Px activities were decreased significantly comparing with saline + none I/R group. Both tadalafil and sildenafil treatment decreased the MDA levels significantly in ischemia/reperfusion groups, whereas this effect was significantly more potent with tadalafil. SOD levels were significantly decreased in all groups after I/R. Tadalafil seems to be more effective than sildenafil by means of increasing GSH-Px activity significantly after I/R. CONCLUSION: Our results indicate some beneficial effects of PDE5 inhibitory drugs, especially tadalafil, on oxidative I/R injury in fetal rat brains.

Spatial heterogeneity in oligodendrocyte lineage maturation and not cerebral blood flow predicts fetal ovine periventricular white matter injury.

Although periventricular white matter injury (PWMI) is the leading cause of chronic neurological disability and cerebral palsy in survivors of premature birth, the cellular-molecular mechanisms by which ischemia-reperfusion contributes to the pathogenesis of PWMI are not well defined. To define pathophysiologic relationships among ischemia, acute cerebral white matter damage, and vulnerable target populations, we used a global cerebral ischemia-reperfusion model in the instrumented 0.65 gestation fetal sheep. We developed a novel method to make repeated measurements of cerebral blood flow using fluorescently labeled microspheres to resolve the spatial heterogeneity of flow in situ in three-dimensional space. Basal flow in the periventricular white matter (PVWM) was significantly lower than in the cerebral cortex. During global cerebral ischemia induced by carotid occlusion, flow to all regions was reduced by nearly 90%. Ischemia of 30 or 37 min duration generated selective graded injury to frontal and parietal PVWM, two regions of predilection for human PWMI. Injury was proportional to the duration of ischemia and increased markedly with 45 min of ischemia to extensively damage cortical and subcortical gray matter. Surprisingly, the distribution of PVWM damage was not uniform and not explained by heterogeneity in the degree of white matter ischemia. Rather, the extent of white matter damage coincided with the presence of a susceptible population of late oligodendrocyte progenitors. These data support that although ischemia is necessary to generate PWMI, the presence of susceptible populations of oligodendrocyte progenitors underlies regional predilection to injury.

Post-hypoxic hypoperfusion is associated with suppression of cerebral metabolism and increased tissue oxygenation in near-term fetal sheep.

Secondary cerebral hypoperfusion is common following perinatal hypoxia-ischaemia. However, it remains unclear whether this represents a true failure to provide sufficient oxygen and nutrients to tissues, or whether it is simply a consequence of reduced cerebral metabolic demand. We therefore examined the hypothesis that cerebral oxygenation would be reduced during hypoperfusion after severe asphyxia, and further, that the greater neural injury associated with blockade of the adenosine A(1) receptor during the insult would be associated with greater hypoperfusion and deoxygenation. Sixteen near-term fetal sheep received either vehicle or 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) for 1 h, followed by 10 min of severe asphyxia induced by complete occlusion of the umbilical cord. Infusions were discontinued at the end of the occlusion and data were analysed for the following 8 h. A transient, secondary fall in carotid artery blood flow and laser Doppler flow was seen from approximately 1-4 h after occlusion (P < 0.001), with no significant differences between vehicle and DPCPX. Changes in laser Doppler blood flow were highly correlated with carotid blood flow (r(2)= 0.81, P < 0.001). Cortical metabolism was suppressed, reaching a nadir 1 h after occlusion and then resolving. Cortical tissue P(O(2)) was significantly increased at 1, 2 and 3 h after occlusion compared to baseline, and inversely correlated with carotid blood flow (r(2)= 0.69, P < 0.001). In conclusion, contrary to our initial hypothesis, delayed posthypoxic hypoperfusion was associated with suppression of cerebral metabolism and increased tissue P(O(2)), and was not significantly affected by preceding adenosine A1 blockade. These data suggest that posthypoxic hypoperfusion is actively mediated and reflects suppressed cerebral metabolism.

Prenatal cocaine exposure increases heart susceptibility to ischaemia-reperfusion injury in adult male but not female rats.

The present study tested the hypothesis that prenatal cocaine exposure differentially regulates heart susceptibility to ischaemia-reperfusion (I/R) injury in adult offspring male and female rats. Pregnant rats were administered intraperitoneally either saline or cocaine (15 mg kg(-1)) twice daily from day 15 to day 21 of gestational age. There were no differences in maternal weight gain and birth weight between the two groups. Hearts were isolated from 2-month-old male and female offspring and were subjected to I/R (25 min/60 min) in a Langendorff preparation. Preischaemic values of left ventricular (LV) function were the same between the saline control and cocaine-treated hearts for both male and female rats. Prenatal cocaine exposure significantly increased I/R-induced myocardial apoptosis and infarct size, and significantly attenuated the postischaemic recovery of LV function in adult male offspring. In contrast, cocaine did not affect I/R-induced injury and postischaemic recovery of LV function in the female hearts. There was a significant decrease in PKCepsilon and phospho-PKCepsilon levels in LV in the male, but not female, offspring exposed to cocaine before birth. These results suggest that prenatal cocaine exposure causes a sex-specific increase in heart susceptibility to I/R injury in adult male offspring, and the decreased PKCepsilon gene expression in the male heart may play an important role.

Role of mitochondrial permeability transition in fetal brain damage in rats.

Recirculation after transient in utero ischemia has previously been found to be accompanied by delayed deterioration of cellular bioenergetic state and of mitochondrial function in the fetal rat brain. Our objective was to assess whether the delayed deterioration is a result of the activation of mitochondrial permeability transition which is observed ultrastructurally as mitochondrial swelling. The respiratory activities and ultrastructure of isolated mitochondria and the cellular bioenergetic state in fetal rat brain were examined at the end of 30 minutes of in utero ischemia and after 1, 2, 3 and 4 hours of recirculation. Cyclosporin A, a potent and virtually specific mitochondrial permeability transition blocker, or vehicle was administered 1 hour after recirculation. In the vehicle-treated animals, the transient ischemia was associated with a delayed deterioration of cellular bioenergetic state and mitochondrial activities at 4 hours of recirculation. The number of swollen mitochondria increased markedly after 4 hours of recirculation. The deterioration and the swelling were prevented by cyclosporin A. The present study indicates that cyclosporin A treatment improves recovery of fetal brain energy metabolism and inhibits the mitochondrial swelling after transient in utero ischemia. The results suggest that mitochondria and mitochondrial permeability transition may be involved in the development of ischemic brain damage in the immature rat.

Hemodynamic changes during complete umbilical cord occlusion in fetal sheep related to hippocampal neuronal damage.

OBJECTIVES: The purpose of our study was to examine the physiologic changes caused by 10 minutes of umbilical cord occlusion in fetal sheep and to determine the correlation between fetal acidemia or cerebral ischemia and hippocampal neuronal damage. STUDY DESIGN: Thirteen fetal sheep were instrumented and catheterized. Carotid artery blood flow (CaF), fetal mean arterial blood pressure (FMABP), pH, PCO (2), base excess, oxygen saturation (SatO(2)), and PO (2) were monitored throughout the occlusion study. Brain sections were examined for the hippocampal neuronal damage. RESULTS: Our data showed severe ischemia (CaF: 10 +/- 7 mL/min; FMABP: 29 +/- 8 mm Hg) and acidemia (pH: 7.0 +/- 0.05; base excess: -9.9 +/- 2.4 mEq/L) at the end of occlusion. The neuronal damage score had significant correlations with ischemia and also with reperfusion, but not with the acidemic or hypoxic parameters. CONCLUSION: We demonstrated that the degree of hippocampal damage was correlated with the degree of ischemia and reperfusion.