Ihab Halaweish

Assessment of coagulopathy, endothelial injury, and inflammation after traumatic brain injury and hemorrhage in a porcine model

BACKGROUND Traumatic brain injury (TBI) and hemorrhagic shock (HS) can be associated with coagulopathy and inflammation, but the mechanisms are poorly understood. We hypothesized that a combination of TBI and HS would disturb coagulation, damage the endothelium, and activate inflammatory and complement systems. METHODS A total of 33 swine were allocated to either TBI + HS (n = 27, TBI and volume-controlled 40% blood loss) or controls (n = 6, anesthesia and instrumentation). TBI + HS animals were left hypotensive (mean arterial pressure, 30–35 mm Hg) for 2 hours. Blood samples were drawn at baseline, 3 minutes and 15 minutes after injury, as well as following 2 hours of hypotension. Markers of coagulation, anticoagulation, endothelial activation/glycocalyx shedding, inflammation, complement, and sympathoadrenal function were measured. RESULTS The TBI + HS group demonstrated an immediate (3 minutes after injury) activation of coagulation (prothrombin fragment 1 + 2, 289 ng/mL vs. 232 ng/mL, p = 0.03) and complement (C5a, 2.83 ng/mL vs. 2.05 ng/mL, p = 0.05). Shedding of the endothelial glycocalyx (syndecan 1) was evident 15 minutes after injury (851.0 ng/ml vs. 715.5 ng/ml, p = 0.03) while inflammation (tumor necrosis factor &agr; [TNF-&agr;], 81.1 pg/mL vs. 50.8 pg/mL, p = 0.03) and activation of the protein C system (activated protein C, 56.7 ng/mL vs. 26.1 ng/mL, p = 0.01) were evident following the 2-hour hypotension phase. CONCLUSION The combination of TBI and shock results in an immediate activation of coagulation and complement systems with subsequent endothelial shedding, protein C activation, and inflammation.

Hirschsprung-associated enterocolitis: pathogenesis, treatment and prevention

Hirschsprung-associated enterocolitis (HAEC) is a common and sometimes life-threatening complication of Hirschsprung disease (HD). Presenting either before or after definitive surgery for HD, HAEC may manifest clinically as abdominal distension and explosive diarrhea, along with emesis, fever, lethargy, and even shock. The pathogenesis of HAEC, the subject of ongoing research, likely involves a complex interplay between a dysfunctional enteric nervous system, abnormal mucin production, insufficient immunoglobulin secretion, and unbalanced intestinal microflora. Early recognition of HAEC and preventative practices, such as rectal washouts following a pull-through, can lead to improved outcomes. Treatment strategies for acute HAEC include timely resuscitation, colonic decompression, and antibiotics. Recurrent or persistent HAEC requires evaluation for mechanical obstruction or residual aganglionosis, and may require surgical treatment with posterior myotomy/myectomy or redo pull-through. This chapter describes the incidence, pathogenesis, treatment, and preventative strategies in management of HAEC.

Inhibition of Histone Deacetylase 6 Improves Long-term Survival in a Lethal Septic Model

BACKGROUND We recently demonstrated that suberoylanilide hydroxamic acid, a broad-spectrum histone deacetylase (HDAC) inhibitor that inhibits HDACs 1, 2, 3, and 6, improves survival in a mouse model of cecal ligation and puncture (CLP)–induced lethal sepsis. The current study was undertaken to determine the effect of selective inhibition of HDAC isoform on survival, key cytokine production, organ injury, bacteria clearance, and cell apoptosis. METHODS In Experiment 1, C57BL/6J mice were subjected to CLP and, 1 hour later, given intraperitoneal injections of (1) Tubastatin A (inhibitor of HDAC6) dissolved in dimethyl sulfoxide (DMSO), (2) MS-275 (inhibitor of HDACs 1, 2, and 3) in DMSO, and (3) DMSO only. Survival was monitored for 10 days. In Experiment 2, 1 hour after CLP, animals were treated with DMSO vehicle or Tubastatin A. Sham-operated animals served as control. Peritoneal fluid and blood samples were collected for measurement of cytokines at 24 hours or 48 hours. Blood at 48 hours was also used to determine bacteria load. Liver was harvested to evaluate acute liver injury. In Experiment 3, Primary splenocytes were used to assess cytokine responses and phagocytosis. Macrophages were cultured and harvested 3 hours and 6 hours after lipopolysaccharide stimulation in the absence or presence of Tubastatin A to analyze cell apoptosis. RESULTS Animals treated with Tubastatin A, but not MS-275, displayed a significant improvement in survival. Moreover, Tubastatin A significantly inhibited cytokine production in peritoneal fluid and plasma as well as in supernatant from splenocytes stimulated with lipopolysaccharide. Tubastatin A significantly attenuated acute liver injury, increased blood bacteria clearance and splenocyte phagocytosis, and decreased macrophage apoptosis. CONCLUSION HDAC6 inhibition significantly improves survival, reduces “cytokine storm,” attenuates acute livery injury, increases bacteria clearance and immune cell phagocytosis, and inhibits macrophage apoptosis in a lethal mouse CLP model.

Changing Demographics of the American Population

Since 1950, the United States has been in the midst of a profound demographic change: the rapid aging of the population. The baby boom generation began turning 65 in 2011 and is now driving growth at the older ages of the population. This article highlights geriatric demographic changes and illustrates how these and future trends will have wide ranging implications for the US health care system.

Creating a “Pro-survival Phenotype” Through Histone Deacetylase Inhibition: Past, Present, and Future

ABSTRACT Traumatic injuries and their sequelae represent a major source of mortality in the United States and globally. Initial treatment for shock, traumatic brain injury, and polytrauma is limited to resuscitation fluids to replace lost volume. To date, there are no treatments with inherent prosurvival properties. Our laboratory has investigated the use of histone deacetylase inhibitors (HDACIs) as pharmacological agents to improve survival. This class of drugs acts through posttranslational protein modifications and is a direct regulator of chromatin structure and function, as well as the function of numerous cytoplasmic proteins. In models of hemorrhagic shock and polytrauma, administration of HDACIs offers a significant survival advantage, even in the absence of fluid resuscitation. Positive results have also been shown in two-hit models of hemorrhage and sepsis and in hemorrhagic shock combined with traumatic brain injury. Accumulating data generated by our group and others continue to support the use of HDACIs for the creation of a prosurvival phenotype. With further research and clinical trials, HDACIs have the potential to be an integral tool in the treatment of trauma, especially in the prehospital phase.

Creating a Prosurvival Phenotype through a Histone Deacetylase Inhibitor in a Lethal Two-Hit Model

ABSTRACT Objectives: Hemorrhagic shock (HS) can initiate an exaggerated systemic inflammatory response and multiple organ failure, especially if followed by a subsequent inflammatory insult (“second hit”). We have recently shown that histone deacetylase inhibitors can improve survival in rodent models of HS or septic shock, individually. In the present study, we examined whether valproic acid (VPA), a histone deacetylase inhibitor, could prolong survival in a rodent “two-hit” model: HS followed by septic shock from cecal ligation and puncture (CLP). Methods: Male Sprague-Dawley rats (250–300 g) were subjected to sublethal HS (40% blood loss) and then randomly divided into two groups (n = 7/group): VPA and control. The VPA group was treated intraperitoneally with VPA (300 mg/kg in normal saline [NS], volume = 750 &mgr;L/kg). The control group was injected with 750 &mgr;L/kg NS. After 24 h, all rats received CLP followed immediately by injection of the same dose of VPA (VPA group) or NS (vehicle group). Survival was monitored for 10 days. In a parallel study, serum and peritoneal irrigation fluid from VPA- or vehicle-treated rats were collected 3, 6, and 24 h after CLP, and enzyme-linked immunosorbent assay was performed to analyze myeloperoxidase activity and determine tumor necrosis factor &agr; and interleukin 6 concentrations. Hematoxylin-eosin staining of lungs at 24-h time point was performed to investigate the grade of acute lung injury. Results: Rats treated with VPA (300 mg/kg) showed significantly higher survival rates (85.7%) compared with the control (14.3%). Moreover, VPA significantly suppressed myeloperoxidase activity (marker of neutrophil-mediated oxidative damage) and inhibited levels of proinflammatory cytokine tumor necrosis factor &agr; and interleukin 6 in the serum and peritoneal cavity. Meanwhile, the severity of acute lung injury was significantly reduced in VPA-treated animals. Conclusions: We have demonstrated that VPA treatment improves survival and attenuates inflammation in a rodent two-hit model.

Addition of low-dose valproic acid to saline resuscitation provides neuroprotection and improves long-term outcomes in a large animal model of combined traumatic brain injury and hemorrhagic shock.

BACKGROUND Combined traumatic brain injury (TBI) and hemorrhagic shock (HS) is highly lethal. In a nonsurvival model of TBI + HS, addition of high-dose valproic acid (VPA) (300 mg/kg) to hetastarch reduced brain lesion size and associated swelling 6 hours after injury; whether this would have translated into better neurologic outcomes remains unknown. It is also unclear whether lower doses of VPA would be neuroprotective. We hypothesized that addition of low-dose VPA to normal saline (NS) resuscitation would result in improved long-term neurologic recovery and decreased brain lesion size. METHODS TBI was created in anesthetized swine (40–43 kg) by controlled cortical impact, and volume-controlled hemorrhage (40% volume) was induced concurrently. After 2 hours of shock, animals were randomized (n = 5 per group) to NS (3× shed blood) or NS + VPA (150 mg/kg). Six hours after resuscitation, packed red blood cells were transfused, and animals were recovered. Peripheral blood mononuclear cells were analyzed for acetylated histone-H3 at lysine-9. A Neurological Severity Score (NSS) was assessed daily for 30 days. Brain magnetic resonance imaging was performed on Days 3 and 10. Cognitive performance was assessed by training animals to retrieve food from color-coded boxes. RESULTS There was a significant increase in histone acetylation in the NS + VPA–treated animals compared with NS treatment. The NS + VPA group demonstrated significantly decreased neurologic impairment and faster speed of recovery as well as smaller brain lesion size compared with the NS group. Although the final cognitive function scores were similar between the groups, the VPA-treated animals reached the goal significantly faster than the NS controls. CONCLUSION In this long-term survival model of TBI + HS, addition of low-dose VPA to saline resuscitation resulted in attenuated neurologic impairment, faster neurologic recovery, smaller brain lesion size, and a quicker normalization of cognitive functions.

Valproic acid for the treatment of hemorrhagic shock: a dose-optimization study

BACKGROUND Valproic acid (VPA) has been shown to improve survival in animal models of hemorrhagic shock at a dose of 300 mg/kg. Our aim was to identify the ideal dose through dose-escalation, split-dosing, and dose de-escalation regimens. MATERIALS AND METHODS Rats were subjected to sublethal 40% hemorrhage and treated with vehicle or VPA (dose of 300, 400, or 450 mg/kg) after 30 min of shock. Acetylated histones and activated proteins from the PI3K-Akt-GSK-3β survival pathway at different time points were quantified by Western blot analysis. In a similar model, a VPA dose of 200 mg/kg followed 2 h later by another dose of 100 mg/kg was administered. Finally, animals were subjected to a lethal 50% hemorrhage and VPA was administered in a dose de-escalation manner (starting at dose of 300 mg/kg) until a significant drop in percent survival was observed. RESULTS Larger doses of VPA resulted in greater acetylation of histone 3 and increased activation of PI3K pathway proteins. Dose-dependent differences were significant in histone acetylation but not in the activation of the survival pathway proteins. Split-dose administration of VPA resulted in similar results to a single full dose. Survival was as follows: 87.5% with 300 and 250 mg/kg of VPA, 50% with 200 mg/kg of VPA, and 14% with vehicle-treated animals. CONCLUSIONS Although higher doses of VPA result in greater histone acetylation and activation of prosurvival protein signaling, doses as low as 250 mg/kg of VPA confer the same survival advantage in lethal hemorrhagic shock. Also, VPA can be given in a split-dose fashion without a reduction in its cytoprotective effectiveness.

Early Resuscitation with Fresh Frozen Plasma for Traumatic Brain Injury Combined with Hemorrhagic Shock Improves Neurologic Recovery

BACKGROUND We have shown that early administration of fresh frozen plasma (FFP) reduces the size of brain lesions 6 hours after injury in a large animal model of traumatic brain injury (TBI) and hemorrhagic shock (HS). To examine long-term outcomes, we hypothesized that early treatment with FFP would result in faster neurologic recovery and better long-term outcomes in a combined TBI and HS model. STUDY DESIGN Anesthetized Yorkshire swine underwent combined TBI and volume-controlled hemorrhage (40% blood volume). After 2 hours of shock, animals were randomized (n = 5/group) to normal saline (3× shed blood) or FFP (1× shed blood) treatment. A neurologic severity score was assessed for 30 days. Magnetic resonance imaging of the brain was performed at days 3, 10, and 24. Cognitive function was tested by training animals to retrieve food from color-coded boxes. RESULTS Neurologic impairment was lower and speed of recovery was considerably faster in the FFP-treated animals. There was a trend toward a smaller lesion size in FFP-treated animal at days 3 and 10, but this did not reach statistical significance. Both groups reached baseline performance on the cognitive testing; however, FFP-treated animals were able to participate, on average, 8 days earlier due to quicker recovery. CONCLUSIONS This is the first study to demonstrate the beneficial effects of FFP treatment in a long-term survival model of combined TBI and HS. Our data show that early treatment with FFP substantially attenuates the degree of neurologic impairment, improves the rate of recovery, and preserves the cognitive functions.

Roller and centrifugal pumps: A retrospective comparison of bleeding complications in extracorporeal membrane oxygenation

Centrifugal pumps are increasingly used for extracorporeal membrane oxygenation (ECMO) rather than roller pumps. However, shear forces induced by these types of continuousflow pumps are associated with acquired von Willebrand factor deficiency and bleeding complications. This study was undertaken to compare adverse bleeding complications with the use of centrifugal and roller pumps in patients on prolonged ECMO support. The records of all adult ECMO patients from June 2002 to 2013 were retrospectively reviewed using the University of Michigan Health System database and the Extracorporeal Life Support Organization registry, focusing on patients supported for at least 5 days. Ninety-five ECMO patients met criteria for inclusion (48 roller vs. 47 centrifugal pump). Indications included pulmonary (79%), cardiac (15%), and extracorporeal cardiopulmonary resuscitation (6%), without significant difference between the two groups. Despite lower heparin anticoagulation (10.9 vs. 13.7 IU/kg/hr) with centrifugal pumps, there was a higher incidence of nonsurgical bleeding (gastrointestinal, pulmonary, and neurological) in centrifugal pump patients (26.1 vs. 9.0 events/1,000 patient-days, p = 0.024). In conclusion, in our historical comparison, despite reduced anticoagulation, ECMO support using centrifugal pumps was associated with a higher incidence of nonsurgical bleeding. The mechanisms behind this are multifactorial and require further investigation