Experimental Brain Death Research Articles

115.1: Immunologic activation in an experimental brain death liver donor model.

115.1: Immunologic activation in an experimental brain death liver donor model.

337.4: Tacrolimus for Prevention of Right and Left Ventriculo-Arterial Coupling Changes in Experimental Brain Death

337.4: Tacrolimus for Prevention of Right and Left Ventriculo-Arterial Coupling Changes in Experimental Brain Death

Tacrolimus for Prevention of Right and Left Ventriculo-Arterial Coupling Changes in Experimental Brain Death

<h3>Purpose</h3> Right ventricular (RV) dysfunction remains the leading cause of early mortality after cardiac transplantation. Tacrolimus (FK506) is an immune suppressor that could preserve heart function via its anti-inflammation effect in animal models. We sought to determine whether Tacrolimus might prevent brain death-induced RV dysfunction, and biological changes in myocarditis caused by BD acting on inflammation and apoptosis. <h3>Methods</h3> After randomisation to placebo (n=9) or to Tacrolimus (n=8; 0.05 mg.kg⁻¹.day⁻¹), seventeen pigs were assigned to a brain death procedure. One, three, five and seven hours after Cushing reflex, the animals underwent hemodynamic evaluation. After euthanasia of the animals, myocardial tissue was sampled. This was repeated in a control group (n=7). <h3>Results</h3> Seven hours after Cushing reflex, brain death had resulted in increased pulmonary artery pressure (29±2 versus 19±1 mmHg) and in a 33%-decreased RV ratio of end-systolic to pulmonary arterial elastances (Ees/Ea), while left ventriculo (LV)-arterial coupling did not change. This was prevented by Tacrolimus. Brain death-induced RV dysfunction was associated with increased RV expression of interleukin(IL)-6/IL-10, IL-1β, receptors for IL-1 and IL-6, β-3 adrenergic receptor, Toll-like receptor (TLR)-4 and neutrophil infiltration, while β-1 adrenergic receptor, TLR-2 and NLR family pyrin-domain-containing-3 expressions decreased. RV but not LV apoptosis was confirmed by TUNEL staining. Tacrolimus pre-treatment prevented RV-arterial uncoupling and changes in RV expression of IL-1 R, IL-6 R, RV apoptosis, as well as in neutrophil infiltration. <h3>Conclusion</h3> Brain death-induced isolated RV dysfunction is associated with RV activation of inflammation and apoptosis, partly limited by Tacrolimus.

Immunomodulatory effects of thalidomide in an experimental brain death liver donor model

Brain death is characterized by a generalized inflammatory response that results in multiorgan damage. This process is mainly mediated through cytokines, which amplify graft immunogenicity. We investigated the immunological response in a brain death liver donor model and analysed the effects of thalidomide, a drug with powerful immunomodulatory properties. Brain death was induced in male Lewis rats. We studied three groups: Control (sham-operated rats in which trepanation was performed without inserting the balloon catheter), BD (rats subjected to brain death by increasing intracranial pressure) and BD + Thalid (BD rats receiving thalidomide after brain death). After 6 h, serum levels of AST, ALT, LDH, and ALP as well as systemic and hepatic levels of TNF-α, IL1-β, IL-6, and IL-10 were analysed. We also determined the mRNA expression of MHC Class I and Class II, NF-κB, and macrophage infiltration. NF-κB was also examined by electrophoretic mobility shift assay. Thalidomide treatment significantly reduced serum levels of hepatic enzymes and TNF-α, IL-1-β, and IL-6. These cytokines were evaluated at either the mRNA expression or protein level in liver tissue. In addition, thalidomide administration resulted in a significant reduction in macrophages, MHC Class I and Class II, and NF-κB activation. This study reveals that thalidomide significantly inhibited the immunologic response and graft immunogenicity, possibly through suppression of NF-κB activation.

Cardiac MicroRNA Expression Profile After Experimental Brain Death Is Associated With Myocardial Dysfunction and Can Be Modulated by Hypertonic Saline.

Brain death (BD) is associated with systemic inflammatory compromise, which might affect the quality of the transplanted organs. This study investigated the expression profile of cardiac microRNAs (miRNAs) after BD, and their relationship with the observed decline in myocardial function and with the changes induced by hypertonic saline solution (HSS) treatment. Wistar rats were assigned to sham-operation (SHAM) or submitted to BD with and without the administration of HSS. Cardiac function was assessed for 6 h with left ventricular (LV) pressure-volume analysis. We screened 641 rodent miRNAs to identify differentially expressed miRNAs in the heart, and computational and functional analyses were performed to compare the differentially expressed miRNAs and find their putative targets and their related enriched canonical pathways. An enhanced expression in canonical pathways related to inflammation and myocardial apoptosis was observed in BD induced group, with 2 miRNAs, miR-30a-3p, and miR-467f, correlating with the level of LV dysfunction observed after BD. Conversely, HSS treated after BD and SHAM groups showed similar enriched pathways related to the maintenance of heart homeostasis regulation, in agreement with the observation that both groups did not have significant changes in LV function. These findings highlight the potential of miRNAs as biomarkers for assessing damage in BD donor hearts and to monitor the changes induced by therapeutic measures like HSS, opening a perspective to improve graft quality and to better understand the pathophysiology of BD. The possible relation of BD-induced miRNA's on early and late cardiac allograft function must be investigated.

Sumoylation of IkB attenuates NF-kB-induced nitrosative stress at rostral ventrolateral medulla and cardiovascular depression in experimental brain death

BackgroundSmall ubiquitin-related modifier (SUMO) is a group of proteins that participates in post-translational modifications. One known SUMO target is the transcription factor nuclear factor-kB (NF-kB) that plays a pivotal role in many disease processes; sumoylation inactivates NF-kB by conjugation with inhibitors of NF-kB (IkB). Our laboratory demonstrated previously that transcriptional upregulation of nitric oxide synthase II (NOS II) by NF-kB, leading to nitrosative stress by the formation of peroxynitrite in the rostral ventrolateral medulla (RVLM), underpins the defunct brain stem cardiovascular regulation that precedes brain death. Based on an experimental endotoxemia model, this study evaluated the hypothesis that sumoylation plays a pro-life role in brain death by interacting with the NF-kB/NOS II/peroxynitrite signaling pathway in the RVLM.ResultsIn Sprague–Dawley rats, intravenous administration of Escherichia coli lipopolysaccharide (LPS; 10 mg kg−1) elicited an augmentation of SUMO-1 and ubiquitin-conjugase 9 (Ubc9) mRNA or protein levels, alongside SUMO-1-conjugated proteins in the RVLM. Immunoneutralization of SUMO-1 or Ubc9 in the RVLM significantly potentiated the already diminished sumoylation of IkBα and intensified NF-kB activation and NOS II/peroxynitrite expression in this brain stem substrate, together with exacerbated fatality, cardiovascular depression and reduction of an experimental index of a life-and-death signal detected from arterial pressure that disappears in comatose patients signifying failure of brain stem cardiovascular regulation before brain death.ConclusionWe conclude that sumoylation of IkB in the RVLM ameliorates the defunct brain stem cardiovascular regulation that underpins brain death in our experimental endotoxemia modal by reducing nitrosative stress via inhibition of IkB degradation that diminishes the induction of the NF-kB/NOS II/peroxynitrite signaling cascade.

Roles of inflammation and apoptosis in experimental brain death–induced right ventricular failure

Right ventricular (RV) dysfunction remains the leading cause of early death after cardiac transplantation. Methylprednisolone is used to improve graft quality; however, evidence for that remains empirical. We sought to determine whether methylprednisolone, acting on inflammation and apoptosis, might prevent brain death-induced RV dysfunction. After randomization to placebo (n = 11) or to methylprednisolone (n = 8; 15 mg/kg), 19 pigs were assigned to a brain-death procedure. The animals underwent hemodynamic evaluation at 1 and 5 hours after Cushing reflex (i.e., hypertension and bradycardia). The animals euthanized, and myocardial tissue was sampled. This was repeated in a control group (n = 8). At 5 hours after the Cushing reflex, brain death resulted in increased pulmonary artery pressure (27 ± 2 vs 18 ± 1 mm Hg) and in a 30% decreased ratio of end-systolic to pulmonary arterial elastances (Ees/Ea). Cardiac output and right atrial pressure did not change. This was prevented by methylprednisolone. Brain death-induced RV dysfunction was associated with increased RV expression of heme oxygenase-1, interleukin (IL)-6, IL-10, IL-1β, tumor necrosis factor (TNF)-α, IL-1 receptor-like (ST)-2, signal transducer and activator of transcription-3, intercellular adhesion molecules-1 and -2, vascular cell adhesion molecule-1, and neutrophil infiltration, whereas IL-33 expression decreased. RV apoptosis was confirmed by terminal deoxynucleotide transferase-mediated deoxy uridine triphosphate nick-end labeling staining. Methylprednisolone pre-treatment prevented RV-arterial uncoupling and decreased RV expression of TNF-α, IL-1 receptor-like-2, intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and neutrophil infiltration. RV Ees/Ea was inversely correlated to RV TNF-α and IL-6 expression. Brain death-induced RV dysfunction is associated with RV activation of inflammation and apoptosis and is partly limited by methylprednisolone.

Triphasic response of glomerular filtration rate after experimental brain death in pigs

Background Clinical outcome after transplantation of kidneys derived from deceased brain death (DBD) donors is inferior compared to living donor kidneys. In this observational model, the effect of experimental brain death on renal function was evaluated. Furthermore, we tested the effect of DBD transplantation on inflammation and renal metabolism in donors and recipients. Methods Eight Danish landrace pigs served as DBD donors. After four hours of brain death, kidneys were removed and stored for 19 hours at 4°C in Custodiol®. Next, the donor kidneys were transplanted into eight recipients. Glomerular filtration rate (GFR) was determined as urinary clearance of 51Cr-EDTA. Results Immediately following brain death, GFR and urine output were reduced while during the second hour hyperfiltration was observed. Subsequently, GFR and urine output decreased again. No systemicor renal inflammation was observed in the donors, while in recipients only renal inflammatory markers were increased ten hours post-reperfusion. Furthermore, mitochondrial dysfunction tended to develop after reperfusion. Conclusions In conclusion, brain death caused a triphasic response of GFR. No systemic inflammation in brain death donors was observed probably due to active control of hemodynamics by fluid administration. These new insights in the effects of brain death may inspire development of new strategies to improve outcome of DBD kidney transplantation.

Donor Management with N-Octanoyl-Dopamine Improves Renal Function and Reduces Inflammation in An Experimental Brain Death Transplantation Model

We have previously demonstrated protective effects of dopamine (DA) in renal transplantation. Recently, we have developed a non-hemodynamic dopamine derivative, i.e. N-Octanoyl-Dopamine (NOD) that is more effective than DA to protect Human-Umbilical-Vein-Endothelial-Cells (HUVEC) against cold preservation injury. In the present study we tested if NOD donor treatment has anti-inflammatory effects in a brain death animal model. Brain death (BD) was induced in Fisher rats. The kidneys were harvested after 6h and processed for further analysis. Allogeneic renal transplantations were performed to address the influence of NOD donor treatment on renal function and renal inflammation. The kidneys were harvested after 7 days. NOD treatment in brain dead donors neither influenced the hemodynamic status of the animals nor induced tachycardia. Expression of adhesion molecules was significantly reduced in donor kidneys treated with NOD (NOD vs. Control; ICAM-1, 3.4±1 vs. 5.6±2; VCAM-1, 1.6±0.9 vs. 2.9±0.8; P< 0.05). Early renal function in recipients revealed a significantly better renal function in recipients from NOD treated donors (NOD vs. Control: 0.85±0.43 vs. 1.53±0.87, day1; P< 0.05). Banff-classification revealed significantly less vasculitis (P< 0.0001) and tubulitis (P< 0.05) in NOD treated grafts 1 week after transplantation. Our data demonstrate that NOD has potent reno-protective effects. NOD donor treatment not only mitigates deterioration in renal function but also reduces renal inflammation in recipients. Hence, NOD donor treatment might be a new approach in donor management to improve graft outcome.

Amelioration of renal damage by administration of anti-thymocyte globulin to potential donors in a brain death rat model

Brain death (BD), a non-immunological factor of renal injury, triggers an inflammatory process causing pathological signs of cell death in the kidney, such as necrosis and apoptosis. Kidneys from brain dead donors show lower success rates than kidneys from living donors and one strategy to improve transplantation outcome is to precondition the donors. For the first time, anti-rat thymoglobulin (rATG) was administered in an experimental brain death animal model to evaluate if it could ameliorate histopathological damage and improve organ function. Animals were divided into three groups: V (n=5) ventilated for 2h; BD (n=5) brain death and ventilated for 2h; and BD+rATG (n=5) brain death, ventilated for 2h, rATG was administered during brain death (10mg/kg). We observed lower creatinine levels in treatment groups (means): V, 0·88±0·22 mg/dl; BD, 1·37±0·07 mg/dl; and BD+rATG, 0·64±0·02 mg/dl (BD versus BD+rATG, P<0·001). In the BD group there appeared to be a marked increase of ATN, whereas ATN was decreased significantly in the rATG group (V, 2·25±0·5 versus BD, 4·75±0·5, P<0·01; BD+rATG, 2·75±0·5 versus BD 4·75±0·5 P<0·01). Gene expression was evaluated with reverse transcription-polymerase chain reaction; tumour necrosis factor (TNF)-α, interleukin (IL)-6, C3, CD86 showed no significant difference between groups. Increased IL-10 and decreased CCL2 in BD+rATG compared to BD (both cases P<0·01). Myeloperoxidase was increased significantly after the brain death setting (V: 32±7·5 versus BD: 129±18). Findings suggest that rATG administered to potential donors may ameliorate renal damage caused by BD. These findings could contribute in the search for specific cytoprotective interventions to improve the quality and viability of transplanted organs.

Engagement of ubiquitination and de-ubiquitination at rostral ventrolateral medulla in experimental brain death

BackgroundWhereas brain death is a vitally important clinical phenomenon, our contemporary understanding on its underlying cellular mechanisms remains elusive. This study evaluated whether the ubiquitin-proteasome system (UPS) in the rostral ventrolateral medulla (RVLM), a neural substrate that our laboratory identified previously to be intimately related to brain death, is engaged in this fatal process.MethodsWe performed proteomics, Western Blot, real-time PCR, ELISA and pharmacological experiments in conjunction with a clinically relevant experimental endotoxemia model of brain death based on intravenous administration of Escherichia coli lipopolysaccharide in adult male Sprague–Dawley rats.ResultsProteomics, Western blot and enzyme activity analyses demonstrated that polyubiquitination was preserved and de-ubiquitination by ubiquitin C-terminal hydrolase isozyme-L1 (UCH-L1) was sustained, alongside increased monoubiquitin availability or proteasome activity in RVLM over the course of experimental endotoxemia. However, real-time PCR revealed no significant alteration in proteasome subunit alpha type-1, ubiquitin or UCH-L1 at mRNA level. Functionally, whereas microinjection into the bilateral RVLM of proteasome inhibitors (lactacystin or proteasome inhibitor II) potentiated survival, an inhibitor of ubiquitin-recycling (ubiquitin aldehyde) or an UCH-L1 inhibitor exacerbated mortality.ConclusionsWe proposed previously that the progression towards brain death entails a tug-of-war between pro-death and pro-life programs in RVLM. It is conceivable that ubiquitination or de-ubiquitination in RVLM participate in brain death by regulating the degradation of the proteins involved in those programs.

A Double-Edged Sword Role for Ubiquitin-Proteasome System in Brain Stem Cardiovascular Regulation During Experimental Brain Death

BackgroundBrain stem cardiovascular regulatory dysfunction during brain death is underpinned by an upregulation of nitric oxide synthase II (NOS II) in rostral ventrolateral medulla (RVLM), the origin of a life-and-death signal detected from blood pressure of comatose patients that disappears before brain death ensues. Furthermore, the ubiquitin-proteasome system (UPS) may be involved in the synthesis and degradation of NOS II. We assessed the hypothesis that the UPS participates in brain stem cardiovascular regulation during brain death by engaging in both synthesis and degradation of NOS II in RVLM.Methodology/Principal FindingsIn a clinically relevant experimental model of brain death using Sprague-Dawley rats, pretreatment by microinjection into the bilateral RVLM of proteasome inhibitors (lactacystin or proteasome inhibitor II) antagonized the hypotension and reduction in the life-and-death signal elicited by intravenous administration of Escherichia coli lipopolysaccharide (LPS). On the other hand, pretreatment with an inhibitor of ubiquitin-recycling (ubiquitin aldehyde) or ubiquitin C-terminal hydrolase isozyme L1 (UCH-L1) potentiated the elicited hypotension and blunted the prevalence of the life-and-death signal. Real-time polymerase chain reaction, Western blot, electrophoresis mobility shift assay, chromatin immunoprecipitation and co-immunoprecipitation experiments further showed that the proteasome inhibitors antagonized the augmented nuclear presence of NF-κB or binding between NF-κB and nos II promoter and blunted the reduced cytosolic presence of phosphorylated IκB. The already impeded NOS II protein expression by proteasome inhibitor II was further reduced after gene-knockdown of NF-κB in RVLM. In animals pretreated with UCH-L1 inhibitor and died before significant increase in nos II mRNA occurred, NOS II protein expression in RVLM was considerably elevated.Conclusions/SignificanceWe conclude that UPS participates in the defunct and maintained brain stem cardiovascular regulation during experimental brain death by engaging in both synthesis and degradation of NOS II at RVLM. Our results provide information on new therapeutic initiatives against this fatal eventuality.

Sumoylation of Hypoxia-Inducible Factor-1α Ameliorates Failure of Brain Stem Cardiovascular Regulation in Experimental Brain Death

BackgroundOne aspect of brain death is cardiovascular deregulation because asystole invariably occurs shortly after its diagnosis. A suitable neural substrate for mechanistic delineation of this aspect of brain death resides in the rostral ventrolateral medulla (RVLM). RVLM is the origin of a life-and-death signal that our laboratory detected from blood pressure of comatose patients that disappears before brain death ensues. At the same time, transcriptional upregulation of heme oxygenase-1 in RVLM by hypoxia-inducible factor-1α (HIF-1α) plays a pro-life role in experimental brain death, and HIF-1α is subject to sumoylation activated by transient cerebral ischemia. It follows that sumoylation of HIF-1α in RVLM in response to hypoxia may play a modulatory role on brain stem cardiovascular regulation during experimental brain death.Methodology/Principal FindingsA clinically relevant animal model that employed mevinphos as the experimental insult in Sprague-Dawley rat was used. Biochemical changes in RVLM during distinct phenotypes in systemic arterial pressure spectrum that reflect maintained or defunct brain stem cardiovascular regulation were studied. Western blot analysis, EMSA, ELISA, confocal microscopy and immunoprecipitation demonstrated that drastic tissue hypoxia, elevated levels of proteins conjugated by small ubiquitin-related modifier-1 (SUMO-1), Ubc9 (the only known conjugating enzyme for the sumoylation pathway) or HIF-1α, augmented sumoylation of HIF-1α, nucleus-bound translocation and enhanced transcriptional activity of HIF-1α in RVLM neurons took place preferentially during the pro-life phase of experimental brain death. Furthermore, loss-of-function manipulations by immunoneutralization of SUMO-1, Ubc9 or HIF-1α in RVLM blunted the upregulated nitric oxide synthase I/protein kinase G signaling cascade, which sustains the brain stem cardiovascular regulatory machinery during the pro-life phase.Conclusions/SignificanceWe conclude that sumoylation of HIF-1α in RVLM ameliorates brain stem cardiovascular regulatory failure during experimental brain death via upregulation of nitric oxide synthase I/protein kinase G signaling. This information should offer new therapeutic initiatives against this fatal eventuality.

Standardized experimental brain death model for studies of intracranial dynamics, organ preservation, and organ transplantation in the pig*

Brain death impairs organ function and outcome after transplantation. There is a need for a brain death model to allow studies of organ viability and preservation. For neurointensive care research, it is also of interest to have a relevant brain death model for studies of intracranial dynamics and evaluation of cerebral monitoring devices. Therefore, the objective was to develop a standardized clinically relevant brain death model. Six pigs of both sexes (10-12 wks old; mean weight, 24.5±1.4 kg) were included. Mean arterial blood pressure, heart rate, intracranial pressure, intracranial compliance, cerebral perfusion pressure, and brain tissue oxygenation (BtiPo2) were recorded during stepwise elevation of intracranial pressure by inflation of an epidural balloon catheter with saline (1 mL/20 mins). Brain death criteria were decided to be reached when cerebral perfusion pressure was <0 mm Hg for 60 mins and at least 10 mL saline was inflated epidurally. BtiPo2 and arterial injections of microspheres were used for confirmation of brain death. A gradual volume-dependent elevation of intracranial pressure was observed. After 10 mL of balloon infusion, mean intracranial pressure was 89.8±9.7 (sd) mm Hg. Intracranial compliance decreased from 0.137±0.069 mL/mm Hg to 0.007±0.001 mL/mm Hg. The mean arterial pressure decreased and the heart rate increased when the intracranial volume was increased to between 5 and 6 mL. All animals showed cerebral perfusion pressure≤0 after 7 to 10 mL of infusion. In all animals, the criteria for brain death with negative cerebral perfusion pressure and BtiPo2 ∼0 mm Hg were achieved. Only a negligible amount of microspheres were found in the cerebrum, confirming brain death. The kidneys showed small foci of acute tubular necrosis. The standardized brain death model designed in pigs simulates the clinical development of brain death in humans with a classic pressure-volume response and systemic cardiovascular reactions. Brain death was convincingly confirmed.

Improvement of Donor Myocardial Function after Treatment of Autonomic Storm During Brain Death

In experimental brain death models, autonomic storm (AS) triggers severe myocardial dysfunction, which can be attenuated by pharmacologic treatment. The aim of this study was to determine the incidence of AS in a cohort of human organ donors and to evaluate the potential interest of AS treatment on myocardial function, cardiac harvesting and transplantation. The cohort consisted of 152 patients. Among them, 46 patients were initially considered as potential cardiac donors (main criteria: age < 60 years, no history of cardiac disease). AS diagnosis included increased systolic arterial pressure > 200 mm Hg associated with tachycardia >140 beats/min. Heart acceptance criteria were associated creatine kinase (CK), troponin Ic, and left ventricle ejection fraction (LVEF) estimated by echocardiography and visual inspection. AS was observed in 29 patients (63%). Hypertension was treated in 12 patients (esmolol n = 6, urapidil n = 5, nicardipine). Cardiac harvesting was performed in 28 donors (61%). LVEFs were significantly higher after AS treatment (no AS: 55.4 +/- 13.4%, untreated AS: 49.0 +/- 18.8%, treated AS: 63.9+ +/- 10.3%, P = 0.049). AS treatment was found to be independently associated with LVEF in > 50% of the cases (P = 0.034). Treatment of AS or the lack of AS were associated with an increased probability of successful cardiac transplantation (OR = 8.8; 95% CI 2.1-38.3, P = 0.002). Treatment of hypertension during AS may attenuate brain death-induced myocardial dysfunction and increase the number of available cardiac grafts.

High Energy Compound Stability During Experimental Brain Death

The aim of this study was to examine the effect of sudden brain death (BD) on myocardial function and high energy phosphate (HEP) stores. BD was induced by cerebral vessel ligation in six swine (BD group) that were compared to six control swine. At the end of the BD period (3 hours), harvested hearts were stored at 4°C. Myocardial tissue HEP were assessed by: (i) 31P-NMR spectroscopy of left ventricle for phosphocreatine (PCr), adenosine triphosphate (ATP), inorganic phosphate (Pi) and intracellular pH (pHi), and by (ii) HPLC for ATP, ADP, and AMP levels in left ventricle biopsies. Brain death resulted in a instantaneous major increase in catecholamines (>50-fold, P < .001) and paradoxically a significant progressive decrease in the regional contractility of the left ventricle. After cardioplegia, no significant differences on HEP compounds (ATP/Pi, PCr/Pi, ATP, energetic index) or in pHi were observed between BD and control groups. These data suggest that early heart injury occuring during BD does not seem to be an ischemic phenomenon.

Alteration in the Transcriptional Profile of Livers from Brain-dead Organ Donors

There is evidence that brain death causes changes in peripheral organs. Marked inflammation is found in organs collected during experimental brain death and clinical studies indicate that, despite genetic mismatch, organs obtained from living donors show improved survival over those from brain-dead donors. The aim of the present clinical research was to explore changes in the transcriptional profile of livers from brain-dead organ donors. Using the cDNA macroarray technique, we compared gene expression in liver biopsies from 21 brain-dead organ donors and in normal liver tissue obtained during resection of benign focal lesions. Analysis of gene expression showed significant differences in the mRNA levels of 117 genes. There was reduced expression of 93 genes whereas expression of 24 genes was enhanced. Downregulated pathways included transcripts related to morphogenesis, blood coagulation, complement cascade, amine metabolism, lipid metabolism, nucleic acid metabolism, biodegradation of xenobiotics, signal transduction, and transcription. Conversely, there was induction of genes related to acute phase response, damage-related response, electron transport, and energy metabolism. The present research demonstrates major changes in the transcriptional profile of livers from brain-dead organ donors. The presence of both down- and upregulated gene families suggests that the alteration in transcriptional profile is not a consequence of death-associated organ failure, but rather, an active change in regulatory mechanisms.

Stable Myocardial Function and Endocrine Dysfunction During Experimental Brain Death

The origin of cardiac impairment during brain death (BD) is controversial. Using a pig experimental model we sought to assess hormonal changes during the first stage of brain death and how these changes contribute to hemodynamic alteration and myocardial dysfunction. Twenty-two pigs were randomized into 2 groups: a control (C) group and a BD group. BD was induced by sub-dural inflation of a balloon catheter. Micromanometers and ultrasonic flow probes were placed on the myocardium to measure cardiovascular parameters. Blood samples and hemodynamic parameters were analyzed before and after induction of BD. A biphasic release of catecholamines was observed, with an initial peak occurring 1 minute after BD induction, followed by a second peak at 60 minutes. Similarly, a biphasic evolution of dP/dt(max) and systolic blood pressure (SBP) was observed at BD, in parallel with catecholamine evolution. In the BD group, both cortisol and aldosterone decreased progressively over time. Circulating triiodothyronine (T3), levothyroxine (T4), prolactin and melatonin concentrations were similar to those of the control group. The difference in arteriovenous (AV) lactate level in arterial and coronary sinus blood was not significantly different between the 2 groups, suggesting an absence of myocardial ischemia. Furthermore, myocardial contractility was not altered during the 3 hours of BD. During the initial period after induction of brain death, cerebral and thyroid hormones remained stable while cortico- and medullo-surrenal hormones varied significantly. We suggest that suprarenal gland impairment is among the first events occurring during brain death. Paradoxically, hemodynamic parameters and myocardial function were not found to be altered.

Renal expression of heat shock proteins after brain death induction in rats

The majority of transplanted kidneys are derived from brain-dead patients. This nonphysiological state influences the hemodynamic and hormonal status of the donor. As a result, kidneys derived from brain-dead donors have inferior graft survival and increased graft function loss. Heat shock proteins (HSPs) are a family of stress-inducible proteins involved in maintaining cell homeostasis and regulating the immune system. We studied renal expression of the genes HO-1, HSP27, HSP40, and HSP70 after experimental brain death in rats. Brain death was induced in male F344 rats by slowly inflating a balloon catheter in the epidural space. Untreated rats were used as controls. Animals were humanely killed after 4 hours of brain death. Kidneys were analysed using RT-PCR, Western blotting, and immunohistochemistry. RT-PCR showed an increase in expression of genes coding for HO-1 (3.6-fold; P < .05) and HSP70 (2.7-fold; P < .05) after brain death. Western blotting also revealed an increase in HO-1 protein levels (4.6-fold; P < .001) but changes in HSP70 protein expression were not detected. Immunohistochemistry showed increments of HO-1 protein expression in the renal cortical tubules of brain-dead rats. HSP70 was predominantly increased in renal distal tubules of brain-dead rats treated for hypotension. No changes were observed in renal HSP27 and HSP40 expression after brain death. Renal stress caused by brain death induces expression of the cytoprotective genes HO-1 and HSP70, but not of HSP27 and HSP40. The up-regulation of these cytoprotective genes could be part of a recuperative mechanism induced by stress associated with brain death.

Functional evidence of reversible ischemic injury immediately after the sympathetic storm associated with experimental brain death

Background Acute brain death from increased intracranial pressure results in a transient increase in myocardial adenosine and lactate, which indicates that oxygen demand exceeds oxygen delivery during the sympathetic “storm”. The aim of this study was to determine the functional significance of this period of ischemia. Methods Brain death was inflicted on 40 Westran pigs (36.5–68.0 kg) by inflating a 21-ml subdural balloon over 3 minutes. In 38 animals, micromanometry and sonomicrometry were used to obtain left ventricular pressure-volume loops to determine the preload recruitable stroke work (PRSW) relationship. Data files were recorded before and at 15-minute intervals after beginning balloon inflation. Plasma troponin I was measured before and 60 minutes after beginning balloon inflation in the 38 instrumented and 2 non-instrumented animals. Results All animals experienced the classical sympathetic storm. The slope of the PRSW relationship decreased, and the volume-axis intercept shifted to the right 15 minutes after beginning balloon inflation ( p < 0.0001). Progressive incremental recovery (leftward shift) occurred between subsequent time points ( p ≤ 0.0018). In the instrumented animals, the mean plasma troponin I level increased from 1.4 ± 1.6 μg/liter to 2.8 ± 2.3 μg/liter ( p < 0.001). However, troponin I was not detected before or after induction of brain death in the plasma of either non-instrumented animal ( p = 0.001). Conclusions The sympathetic storm produced transient contractile dysfunction, consistent with ischemic injury. However, troponin I release reflected surgical instrumentation and not brain death.