The Integrin Pathway Partially Mediates Stretch-Induced Deficits in Primary Rat Microglia.

Stretch-injured microglia display significantly altered morphology, function and inflammatory-associated gene expression when cultured on a synthetic fibronectin substrate. However, the mechanism by which stretch induces these changes is unknown. Integrins, such as α5ß1, mediate microglial attachment to fibronectin via the RGD binding peptide; following integrin ligation the integrin-associated signaling enzyme, focal adhesion kinase (FAK), autophosphorylates tyrosine residue 397 and mediates multiple downstream cellular processes. We therefore hypothesize that blocking the RGD binding/integrin pathway with a commercially available RGD peptide will mimic the stretch-induced morphological alterations and functional deficits in microglia. Further, we hypothesize that upregulation of stretch-inhibited downstream integrin signaling will reverse these effects. Using primary rat microglia, we tested the effects of RGD binding peptide and a FAK activator on cellular function and structure and response to stretch-injury. Similar to injured cells, RGD peptide administration significantly decreases media nitric oxide (NO) levels and iNOS expression and induced morphological alterations and migratory deficits. While stretch-injury and RGD peptide administration decreased phosphorylation of the tyrosine 397 residue on FAK, 20 nM of ZINC 40099027, an activator specific to the tyrosine 397 residue, rescued the stretch-induced decrease in FAK phosphorylation and ameliorated the injury-induced decrease in media NO levels, iNOS expression and inflammatory associated gene expression. Additionally, treatment alleviated morphological changes observed after stretch-injury and restored normal migratory behavior to control levels. Taken together, these data suggest that the integrin/FAK pathway partially mediates the stretch-injured phenotype in microglia, and may serve as a pathway to modulate microglial responses.

Selective vulnerability of the cerebral vasculature to blast injury in a rat model of mild traumatic brain injury.

BACKGROUND: Blast-related traumatic brain injury (TBI) is a common cause of injury in the military operations in Iraq and Afghanistan. How the primary blast wave affects the brain is not well understood. The aim of the present study was to examine whether blast exposure affects the cerebral vasculature in a rodent model. We analyzed the brains of rats exposed to single or multiple (three) 74.5 kPa blast exposures, conditions that mimic a mild TBI. Rats were sacrificed 24 hours or between 6 and 10 months after exposure. Blast-induced cerebral vascular pathology was examined by a combination of light microscopy, immunohistochemistry, and electron microscopy. RESULTS: We describe a selective vascular pathology that is present acutely at 24 hours after injury. The vascular pathology is found at the margins of focal shear-related injuries that, as we previously showed, typically follow the patterns of penetrating cortical vessels. However, changes in the microvasculature extend beyond the margins of such lesions. Electron microscopy revealed that microvascular pathology is found in regions of the brain with an otherwise normal neuropil. This initial injury leads to chronic changes in the microvasculature that are still evident many months after the initial blast exposure. CONCLUSIONS: These studies suggest that vascular pathology may be a central mechanism in the induction of chronic blast-related injury.

Select small nucleolar RNAs in blood components as novel biomarkers for improved identification of comorbid traumatic brain injury and post-traumatic stress disorder in veterans of the conflicts in Afghanistan and Iraq.

BACKGROUND: The present study was designed to validate the ability of our recently identified set of small noncoding RNA candidate mild traumatic brain injury (mTBI) biomarkers to diagnose mTBI in the presence or absence of post-traumatic stress disorder (PTSD) comorbidity. Using qPCR, we explored the regulation of the candidate biomarkers in peripheral blood mononuclear cells (PBMC) from 58 veterans. RESULTS: We confirmed that 4 small nucleolar RNAs (snoRNAs), ACA48, U35, U55, and U83A, are significantly down-regulated in PBMC from veterans with mTBI and PTSD compared to non-TBI, control subjects with PTSD only. We found that the snoRNA biomarkers are able to dissect subjects with comorbid mTBI and PTSD from PTSD subjects without mTBI with 100% sensitivity, 81% accuracy, and 72% specificity. No significant differential expression of snoRNA biomarkers was found in mTBI subjects without comorbid PTSD. However, we found significantly lower U55 contents in subjects with PTSD. We explored the regulation of ACA48 in rodent models of PTSD or blast-induced mTBI to gather proof-of-concept evidence that would connect the regulation of the biomarkers and the development of mTBI or PTSD. We found no change in the regulation of ACA48 in the mTBI rat model. We did, however, find significant down-regulation of ACA48 in the PTSD mouse model 24 hours following psychological trauma exposure. This may reflect a short-term response to trauma exposure, since we found no change in the regulation of ACA48 in veteran PTSD subjects 3.6 years post-deployment. CONCLUSIONS: Additional application of the 4 snoRNA biomarker to current diagnostic criteria may provide an objective biomarker pattern to help identify veterans with comorbid mTBI and PTSD. Our observations suggest that biological interactions between TBI and PTSD may contribute to the clinical features of veterans with comorbid mTBI and PTSD. Future investigations on mTBI mechanisms or TBI biomarkers should consider their interactions with PTSD.

Blast overpressure induces shear-related injuries in the brain of rats exposed to a mild traumatic brain injury.

BACKGROUND: Blast-related traumatic brain injury (TBI) has been a significant cause of injury in the military operations of Iraq and Afghanistan, affecting as many as 10-20% of returning veterans. However, how blast waves affect the brain is poorly understood. To understand their effects, we analyzed the brains of rats exposed to single or multiple (three) 74.5 kPa blast exposures, conditions that mimic a mild TBI. RESULTS: Rats were sacrificed 24 hours or between 4 and 10 months after exposure. Intraventricular hemorrhages were commonly observed after 24 hrs. A screen for neuropathology did not reveal any generalized histopathology. However, focal lesions resembling rips or tears in the tissue were found in many brains. These lesions disrupted cortical organization resulting in some cases in unusual tissue realignments. The lesions frequently appeared to follow the lines of penetrating cortical vessels and microhemorrhages were found within some but not most acute lesions. CONCLUSIONS: These lesions likely represent a type of shear injury that is unique to blast trauma. The observation that lesions often appeared to follow penetrating cortical vessels suggests a vascular mechanism of injury and that blood vessels may represent the fault lines along which the most damaging effect of the blast pressure is transmitted.

Blast traumatic brain injury in the rat using a blast overpressure model.

Traumatic brain injury (TBI) is a serious health concern for civilians and military populations, and blast-induced TBI (bTBI) has become an increasing problem for military personnel over the past 10 years. To understand the biological and psychological effects of blast-induced injuries and to examine potential interventions that may help to prevent, attenuate, and treat effects of bTBI, it is valuable to conduct controlled animal experiments. This unit discusses available paradigms to model traumatic brain injury in animals, with an emphasis on the relevance of these various models to study blast-induced traumatic brain injury (bTBI). This paper describes the detailed methods of a blast overpressure (BOP) paradigm that has been used to conduct experiments with rats to model blast exposure. This particular paradigm models the pressure wave created by explosions, including improvised explosive devices (IEDs).

Acute blast injury reduces brain abeta in two rodent species.

Blast-induced traumatic brain injury (TBI) has been a major cause of morbidity and mortality in the conflicts in Iraq and Afghanistan. How the primary blast wave affects the brain is not well understood. In particular, it is unclear whether blast injures the brain through mechanisms similar to those found in non-blast closed impact injuries (nbTBI). The ß-amyloid (Aß) peptide associated with the development of Alzheimer's disease is elevated acutely following TBI in humans as well as in experimental animal models of nbTBI. We examined levels of brain Aß following experimental blast injury using enzyme-linked immunosorbent assays for Aß 40 and 42. In both rat and mouse models of blast injury, rather than being increased, endogenous rodent brain Aß levels were decreased acutely following injury. Levels of the amyloid precursor protein (APP) were increased following blast exposure although there was no evidence of axonal pathology based on APP immunohistochemical staining. Unlike the findings in nbTBI animal models, levels of the ß-secretase, ß-site APP cleaving enzyme 1, and the γ-secretase component presenilin-1 were unchanged following blast exposure. These studies have implications for understanding the nature of blast injury to the brain. They also suggest that strategies aimed at lowering Aß production may not be effective for treating acute blast injury to the brain.

Blast exposure induces post-traumatic stress disorder-related traits in a rat model of mild traumatic brain injury.

Blast related traumatic brain injury (TBI) has been a major cause of injury in the wars in Iraq and Afghanistan. A striking feature of the mild TBI (mTBI) cases has been the prominent association with post-traumatic stress disorder (PTSD). However, because of the overlapping symptoms, distinction between the two disorders has been difficult. We studied a rat model of mTBI in which adult male rats were exposed to repetitive blast injury while under anesthesia. Blast exposure induced a variety of PTSD-related behavioral traits that were present many months after the blast exposure, including increased anxiety, enhanced contextual fear conditioning, and an altered response in a predator scent assay. We also found elevation in the amygdala of the protein stathmin 1, which is known to influence the generation of fear responses. Because the blast overpressure injuries occurred while animals were under general anesthesia, our results suggest that a blast-related mTBI exposure can, in the absence of any psychological stressor, induce PTSD-related traits that are chronic and persistent. These studies have implications for understanding the relationship of PTSD to mTBI in the population of veterans returning from the wars in Iraq and Afghanistan.