Reduction of neutrophil activity decreases early microvascular injury after subarachnoid haemorrhage

<p>Abstract</p> <p>Background</p> <p>Subarachnoid haemorrhage (SAH) elicits rapid pathological changes in the structure and function of parenchymal vessels (≤ 100 μm). The role of neutrophils in these changes has not been determined. This study investigates the role of neutrophils in early microvascular changes after SAH</p> <p>Method</p> <p>Rats were either untreated, treated with vinblastine or anti-polymorphonuclear (PMN) serum, which depletes neutrophils, or treated with pyrrolidine dithiocarbamate (PDTC), which limits neutrophil activity. SAH was induced by endovascular perforation. Neutrophil infiltration and the integrity of vascular endothelium and basement membrane were assessed immunohistochemically. Vascular collagenase activity was assessed by <it>in situ </it>zymography.</p> <p>Results</p> <p>Vinblastine and anti-PMN serum reduced post-SAH accumulation of neutrophils in cerebral vessels and in brain parenchyma. PDTC increased the neutrophil accumulation in cerebral vessels and decreased accumulation in brain parenchyma. In addition, each of the three agents decreased vascular collagenase activity and post-SAH loss of vascular endothelial and basement membrane immunostaining.</p> <p>Conclusions</p> <p>Our results implicate neutrophils in early microvascular injury after SAH and indicate that treatments which reduce neutrophil activity can be beneficial in limiting microvascular injury and increasing survival after SAH.</p

Metamorphosis of Subarachnoid Hemorrhage Research: from Delayed Vasospasm to Early Brain Injury

Delayed vasospasm that develops 3–7 days after aneurysmal subarachnoid hemorrhage (SAH) has traditionally been considered the most important determinant of delayed ischemic injury and poor outcome. Consequently, most therapies against delayed ischemic injury are directed towards reducing the incidence of vasospasm. The clinical trials based on this strategy, however, have so far claimed limited success; the incidence of vasospasm is reduced without reduction in delayed ischemic injury or improvement in the long-term outcome. This fact has shifted research interest to the early brain injury (first 72 h) evoked by SAH. In recent years, several pathological mechanisms that activate within minutes after the initial bleed and lead to early brain injury are identified. In addition, it is found that many of these mechanisms evolve with time and participate in the pathogenesis of delayed ischemic injury and poor outcome. Therefore, a therapy or therapies focused on these early mechanisms may not only prevent the early brain injury but may also help reduce the intensity of later developing neurological complications. This manuscript reviews the pathological mechanisms of early brain injury after SAH and summarizes the status of current therapies

Aneurysmal Subarachnoid Hemorrhage Models: Do They Need a Fix?

The discovery of tissue plasminogen activator to treat acute stroke is a success story of research on preventing brain injury following transient cerebral ischemia (TGI). That this discovery depended upon development of embolic animal model reiterates that proper stroke modeling is the key to develop new treatments. In contrast to TGI, despite extensive research, prevention or treatment of brain injury following aneurysmal subarachnoid hemorrhage (aSAH) has not been achieved. A lack of adequate aSAH disease model may have contributed to this failure. TGI is an important component of aSAH and shares mechanism of injury with it. We hypothesized that modifying aSAH model using experience acquired from TGI modeling may facilitate development of treatment for aSAH and its complications. This review focuses on similarities and dissimilarities between TGI and aSAH, discusses the existing TGI and aSAH animal models, and presents a modified aSAH model which effectively mimics the disease and has a potential of becoming a better resource for studying the brain injury mechanisms and developing a treatment

Gender influences the initial impact of subarachnoid hemorrhage: an experimental investigation.

Aneurysmal subarachnoid hemorrhage (SAH) carries high early patient mortality. More women than men suffer from SAH and the average age of female SAH survivors is greater than that of male survivors; however, the overall mortality and neurological outcomes are not better in males despite their younger age. This pattern suggests the possibility of gender differences in the severity of initial impact and/or in subsequent pathophysiology. We explored gender differences in survival and pathophysiology following subarachnoid hemorrhage induced in age-matched male and female rats by endovascular puncture. Intracranial pressure (ICP), cerebral blood flow (CBF), blood pressure (BP) and cerebral perfusion pressure (CPP) were recorded at and after induction of SAH. Animals were sacrificed 3 hours after lesion and studied for subarachnoid hematoma size, vascular pathology (collagen and endothelium immunostaining), inflammation (platelet and neutrophil immunostaining), and cell death (TUNEL assay). In a second cohort, 24-hour survival was determined. Subarachnoid hematoma, post-hemorrhage ICP peak, BP elevation, reduction in CPP, intraluminal platelet aggregation and neutrophil accumulation, loss of vascular collagen, and neuronal and non-neuronal cell death were greater in male than in female rats. Hematoma size did not correlate with the number of apoptotic cells, platelet aggregates or neutrophil. The ICP peak correlated with hematoma size and with number of apoptotic cells but not with platelet aggregates and neutrophil number. This suggests that the intensity of ICP rise at SAH influences the severity of apoptosis but not of inflammation. Mortality was markedly greater in males than females. Our data demonstrate that in rats gender influences the initial impact of SAH causing greater bleed and early injury in males as compared to females

Vascular pathology.

<p>Cerebral vessels of animals sacrificed 3 hours after SAH. Panel A: representative images of ICA from a single male and a single female rat. Panel B: average vessel sizes. Note that the internal circumference of ICA in SAH males is smaller compared to females. Panel C: representative images showing brain vessels stained for platelets, RECA-1 (an endothelium marker), and collagen-IV (a basal lamina marker); note the greater numbers of RECA-1 and collagen IV stained vessels containing platelet aggregates (arrows) in males. Panel D: average area fractions of RECA-1 and collagen-IV positive vascular profiles of SAH animals as percent changes over sham-operated cohorts. The reduction in the area fraction of RECA-1 is similar in males and females but that of collagen -IV is different. Data are mean ± sem from 5 animals per gender. * significantly difference than females (p <0.01).</p

Cell death.

<p>Activated caspase-3 immunoreactivity and TUNEL staining 3h after SAH. Panels A-B: 4-color fluorescence staining for TUNEL, NeuN, collagen-IV, and DAPI. Panel A: typical micrographs from male and female SAH animals. Each channel is shown as a separate image. Small arrows: TUNEL-positive neurons; large arrowheads: TUNEL-positive vascular cells. Note the greater frequency of TUNEL-positive neurons in male as compared to female. Panel B: average numbers of TUNEL-only, TUNEL+NeuN, and TUNEL+collagen-IV profiles. TUNEL-only and TUNEL-NeuN profiles are significantly greater in males than in females. Panel C: average numbers of profiles positive for activated caspase-3 (Cas) only, Cas+NeuN, and Cas+collagen-IV. All three indexes are significantly greater in male animals. Panel D: Fluoro-Jade B-positive cells (arrows) in representative SAH male and female brain sections. Panel E: Average numbers of Fluoro-Jade B-positive cells in SAH animals. Data are mean ± sem from 5 animals per gender. * significantly gender difference (p <0.05). </p

Cerebral inflammation.

<p>Luminal platelet aggregates and neutrophil accumulation in animals sacrificed 3 hours after SAH. Panel A: representative images of neutrophil staining. Note the greater number of neutrophils in male as compared to female brain. Scale bar = 500 μm. Panels B, C: average numbers of neutrophils and vascular platelet aggregates per whole brain section and per image field, respectively. Both parameters are greater in male as compared to female brains. Data are mean ± sem from 5 animals per gender. * significantly difference than females (p <0.05).</p

Correlation analysis.

<p>The association of ICP peak values with numbers of platelet aggregates (A), neutrophils (B), and apoptotic cells (C). The ICP peak significantly correlated with the number of apoptotic cells but not with the numbers of platelet aggregates or neutrophils. Each point is the mean from a single animal.</p