AT2 activation does not influence brain damage in the early phase after experimental traumatic brain injury in male mice.

Antagonism of the angiotensin II type 1 receptor (AT1) improves neurological function and reduces brain damage after experimental traumatic brain injury (TBI), which may be partly a result of enhanced indirect angiotensin II type 2 receptor (AT2) stimulation. AT2 stimulation was demonstrated to be neuroprotective via anti-inflammatory, vasodilatory, and neuroregenerative mechanisms in experimental cerebral pathology models. We recently demonstrated an upregulation of AT2 after TBI suggesting a protective mechanism. The present study investigated the effect of post-traumatic (5 days after TBI) AT2 activation via high and low doses of a selective AT2 agonist, compound 21 (C21), compared to vehicle-treated controls. No differences in the extent of the TBI-induced lesions were found between both doses of C21 and the controls. We then tested AT2-knockdown animals for secondary brain damage after experimental TBI. Lesion volume and neurological outcomes in AT2-deficient mice were similar to those in wild-type control mice at both 24 h and 5 days post-trauma. Thus, in contrast to AT1 antagonism, AT2 modulation does not influence the initial pathophysiological mechanisms of TBI in the first 5 days after the insult, indicating that AT2 plays only a minor role in the early phase following trauma-induced brain damage.

Influence of rosuvastatin treatment on cerebral inflammation and nitro-oxidative stress in experimental lung injury in pigs.

BACKGROUND: Many patients with acute respiratory distress syndrome (ARDS) suffer from cognitive impairment after hospital discharge. Different mechanisms have been implicated as potential causes for this impairment, inter alia cerebral inflammation. A class of drugs with antioxidant and anti-inflammatory properties are ß-HMG-CoA-reductase inhibitors ("statins"). We hypothesized that treatment with rosuvastatin attenuates cerebral cytokine mRNA expression and nitro-oxidative stress in an animal model of acute lung injury. METHODS: After approval of the institutional and state animal care committee, we performed this prospective randomized controlled animal study in accordance with the international guidelines for the care and use of laboratory animals. Thirty-two healthy male pigs were randomized to one of four groups: lung injury by central venous injection of oleic acid (n = 8), statin treatment before and directly after lung injury (n = 8), statin treatment after lung injury (n = 8), or ventilation-only controls (n = 8). About 18 h after lung injury and standardized treatment, the animals were euthanised, and the brains and lungs were collected for further examinations. We determined histologic lung injury and cerebral and pulmonal cytokine and 3-nitrotyrosine production. RESULTS: We found a significant increase in hippocampal IL-6 mRNA after lung injury (p < 0.05). Treatment with rosuvastatin before and after induction of lung injury led to a significant reduction of hippocampal IL-6 mRNA (p < 0.05). Cerebral 3-nitrotyrosine was significantly higher in lung-injured animals compared with all other groups (p < 0.05 vs. animals treated with rosuvastatin after lung injury induction; p < 0.001 vs. all other groups). 3-Nitrotyrosine was also increased in the lungs of the lung-injured pigs compared to all other groups (p < 0.05 each). CONCLUSIONS: Our findings highlight cerebral cytokine production and nitro-oxidative stress within the first day after induction of lung injury. The treatment with rosuvastatin reduced IL-6 mRNA and 3-nitrotyrosine concentration in the brains of the animals. In earlier trials, statin treatment did not reduce mortality in ARDS patients but seemed to improve quality of life in ARDS survivors. Whether this is attributable to better cognitive function because of reduced nitro-oxidative stress and inflammation remains to be elucidated.

Levosimendan increases brain tissue oxygen levels after cardiopulmonary resuscitation independent of cardiac function and cerebral perfusion.

Prompt reperfusion is important to rescue ischemic tissue; however, the process itself presents a key pathomechanism that contributes to a poor outcome following cardiac arrest. Experimental data have suggested the use of levosimendan to limit ischemia-reperfusion injury by improving cerebral microcirculation. However, recent studies have questioned this effect. The present study aimed to investigate the influence on hemodynamic parameters, cerebral perfusion and oxygenation following cardiac arrest by ventricular fibrillation in juvenile male pigs. Following the return of spontaneous circulation (ROSC), animals were randomly assigned to levosimendan (12 µg/kg, followed by 0.3 µg/kg/min) or vehicle treatment for 6 h. Levosimendan-treated animals showed significantly higher brain PbtO2 levels. This effect was not accompanied by changes in cardiac output, preload and afterload, arterial blood pressure, or cerebral microcirculation indicating a local effect. Cerebral oxygenation is key to minimizing damage, and thus, current concepts are aimed at improving impaired cardiac output or cerebral perfusion. In the present study, we showed that NIRS does not reliably detect low PbtO2 levels and that levosimendan increases brain oxygen content. Thus, levosimendan may present a promising therapeutic approach to rescue brain tissue at risk following cardiac arrest or ischemic events such as stroke or traumatic brain injury.

Effect of fluid resuscitation on cerebral integrity: A prospective randomised porcine study of haemorrhagic shock.

BACKGROUND: The treatment of haemorrhagic shock is a challenging task. Colloids have been regarded as standard treatment, but their safety and benefit have been the subject of controversial debates. Negative effects, including renal failure and increased mortality, have resulted in restrictions on their administration. The cerebral effects of different infusion regimens are largely unknown. OBJECTIVES: The current study investigated the impact of gelatine-polysuccinate, hydroxyethyl starch (HES) and balanced electrolyte solution (BES) on cerebral integrity, focusing on cerebral inflammation, apoptosis and blood flow in pigs. DESIGN: Randomised experimental study. SETTING: University-affiliated large animal research unit. ANIMALS: Twenty-four juvenile pigs aged 8 to 12 weeks. INTERVENTION: Haemorrhagic shock was induced by controlled arterial blood withdrawal to achieve a combination of relevant blood loss (30 to 40 ml kg-1) and haemodynamic deterioration. After 30 min of shock, fluid resuscitation was started with either gelatine-polysuccinate, HES or BES. The animals were then monitored for 4 h. MAIN OUTCOME MEASURES: Cerebral perfusion and diffusion were measured via arterial-spin-labelling MRI. Peripheral tissue perfusion was evaluated via white light spectroscopy. Cortical and hippocampal samples were collected at the end of the experiment. The numbers of cerebral cell nuclei were counted and mRNA expression of markers for cerebral apoptosis [glucose transporter protein type 1 (SLC2A), lipocalin 2 (LCN-2), aquaporin-4 (AQP4)] and inflammation [IL-6, TNF-α, glial fibrillary acidic protein (GFAP)] were determined. RESULTS: The three fluid protocols all stabilised the macrocirculation. Fluid resuscitation significantly increased the cerebral perfusion. Gelatine-polysuccinate and HES initially led to a higher cardiac output but caused haemodilution. Cerebral cell counts (as cells µm-2) were lower after colloid administration in the cortex (gelatine-polysuccinate, 1.8 ±â€Š0.3; HES, 1.9 ±â€Š0.4; each P < 0.05 vs. BES, 2.3 ±â€Š0.2) and the hippocampus (gelatine-polysuccinate, 0.8 ±â€Š0.2; HES, 0.9 ±â€Š0.2; each P < 0.05 vs. BES, 1.1 ±â€Š0.1). After gelatine-polysuccinate, the hippocampal SLC2A and GFAP were lower. After gelatine-polysuccinate, the cortical LCN-2 and TNF-α expression levels were increased (each P < 0.05 vs. BES). CONCLUSION: In a porcine model, fluid resuscitation by colloids, particularly gelatine-polysuccinate, was associated with the occurrence of cerebral injury. ETHICAL APPROVAL NUMBER: 23 177-07/G 15-1-092; 01/2016.

Targeted fibre-optical intrabronchial lipopolysaccharide administration in pigs - a methodical refinement for improved accuracy in respiratory research.

OBJECTIVE: To establish and evaluate a standardized method of targeted, intrabronchial drug delivery in pigs. STUDY DESIGN: Randomized controlled trial. ANIMALS: A total of 16 German Landrace pigs (Sus scrofa), age range 12‒16 weeks, and weighing 28‒35 kg. METHODS: The animals were anaesthetized, intubated, and instrumented with extended cardiovascular monitoring. Lung injury was induced by administering via a flexible fibre-optic endoscope using 100 mL saline solution containing either 20 mg of Escherichia coli lipopolysaccharide (E. coli LPS) (n = 8) or no additive (sham, n = 8) into the two distal mainstem bronchi. The animals were monitored for 8 hours and arterial oxygenation, inspiratory pressure and arterial blood pressure were measured repeatedly. Post-mortem, lung tissue was prepared for histologic damage scoring and determination of proinflammatory cytokines Interleukin-6 (IL-6) and tumour necrosis factor alpha (TNFα). Statistical analyses were performed using inter-group analysis of variance and Student's t tests. Data are presented as mean ± standard deviation. A p value <0.05 was considered significant. RESULTS: The targeted application of LPS led to significant deterioration of oxygenation consistent with mild-to-moderate acute respiratory distress syndrome (ARDS) and hypotension (Horowitz ratio: sham 2 hour, 300 ± 39; LPS 2 hour, 193.7 ± 52; p < 0.001). Histologic analyses identified increased inflammation and oedema in the tissues of the animals in the LPS group IL-6 sham: 6.4 ± 4.4 × 10-5 pg mL-1; IL-6 LPS: 2.8 ± 2.4 × 10-4 pg mL-1, p = 0.015. CONCLUSIONS: The targeted application of agents via flexible fibre-optic endoscopy is a valid, reliable method of causing controlled lung damage in a porcine model. The data presented suggest the feasibility and possible advantages of controlled application and could expand the array of techniques used to help understand the critical condition of ARDS. In addition, a targeted approach could help reduce animal numbers used for this purpose.

Experimental lung injury induces cerebral cytokine mRNA production in pigs.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is an important disease with a high incidence among patients admitted to intensive care units. Over the last decades, the survival of critically ill patients has improved; however, cognitive deficits are among the long-term sequelae. We hypothesize that acute lung injury leads to upregulation of cerebral cytokine synthesis. METHODS: After approval of the institutional and animal care committee, 20 male pigs were randomized to one of three groups: (1) Lung injury by oleic acid injection (OAI), (2) ventilation only (CTR) or (3) untreated. We compared neuronal numbers, proportion of neurons with markers for apoptosis, activation state of Iba-1 stained microglia cells and cerebral mRNA levels of different cytokines between the groups 18 hours after onset of lung injury. RESULTS: We found an increase in hippocampal TNFalpha (p < 0.05) and IL-6 (p < 0.05) messenger RNA (mRNA) in the OAI compared to untreated group as well as higher hippocampal IL-6 mRNA compared to control (p < 0.05). IL-8 and IL-1beta mRNA showed no differences between the groups. We found histologic markers for beginning apoptosis in OAI compared to untreated (p < 0.05) and more active microglia cells in OAI and CTR compared to untreated (p < 0.001 each). CONCLUSION: Hippocampal cytokine transcription increases within 18 hours after the induction of acute lung injury with histological evidence of neuronal damage. It remains to be elucidated if increased cytokine mRNA synthesis plays a role in the cognitive decline observed in survivors of ARDS.

Bi-Level ventilation decreases pulmonary shunt and modulates neuroinflammation in a cardiopulmonary resuscitation model.

BACKGROUND: Optimal ventilation strategies during cardiopulmonary resuscitation are still heavily debated and poorly understood. So far, no convincing evidence could be presented in favour of outcome relevance and necessity of specific ventilation patterns. In recent years, alternative models to the guideline-based intermittent positive pressure ventilation (IPPV) have been proposed. In this randomized controlled trial, we evaluated a bi-level ventilation approach in a porcine model to assess possible physiological advantages for the pulmonary system as well as resulting changes in neuroinflammation compared to standard measures. METHODS: Sixteen male German landrace pigs were anesthetized and instrumented with arterial and venous catheters. Ventricular fibrillation was induced and the animals were left untreated and without ventilation for 4 minutes. After randomization, the animals were assigned to either the guideline-based group (IPPV, tidal volume 8-10 ml/kg, respiratory rate 10/min, FiO21.0) or the bi-level group (inspiratory pressure levels 15-17 cmH2O/5cmH2O, respiratory rate 10/min, FiO21.0). Mechanical chest compressions and interventional ventilation were initiated and after 5 minutes, blood samples, including ventilation/perfusion measurements via multiple inert gas elimination technique, were taken. After 8 minutes, advanced life support including adrenaline administration and defibrillations were started for up to 4 cycles. Animals achieving ROSC were monitored for 6 hours and lungs and brain tissue were harvested for further analyses. RESULTS: Five of the IPPV and four of the bi-level animals achieved ROSC. While there were no significant differences in gas exchange or hemodynamic values, bi-level treated animals showed less pulmonary shunt directly after ROSC and a tendency to lower inspiratory pressures during CPR. Additionally, cytokine expression of tumour necrosis factor alpha was significantly reduced in hippocampal tissue compared to IPPV animals. CONCLUSION: Bi-level ventilation with a constant positive end expiratory pressure and pressure-controlled ventilation is not inferior in terms of oxygenation and decarboxylation when compared to guideline-based IPPV ventilation. Additionally, bi-level ventilation showed signs for a potentially ameliorated neurological outcome as well as less pulmonary shunt following experimental resuscitation. Given the restrictions of the animal model, these advantages should be further examined.

Standardized Model of Ventricular Fibrillation and Advanced Cardiac Life Support in Swine.

Cardiopulmonary resuscitation after cardiac arrest, independent of its origin, is a regularly encountered medical emergency in hospitals as well as preclinical settings. Prospective randomized trials in human subjects are difficult to design and ethically ambiguous, which results in a lack of evidence-based therapies. The model presented in this report represents one of the most common causes of cardiac arrests, ventricular fibrillation, in a standardized setting in a large animal model. This allows for reproducible observations and various therapeutic interventions under clinically accurate conditions, hence facilitating the generation of better evidence and eventually the potential for improved medical treatment.

Fluid resuscitation-related coagulation impairment in a porcine hemorrhagic shock model.

BACKGROUND: Fast and effective treatment of hemorrhagic shock is one of the most important preclinical trauma care tasks e.g., in combat casualties in avoiding severe end-organ damage or death. In scenarios without immediate availability of blood products, alternate regimens of fluid resuscitation represent the only possibility of maintaining sufficient circulation and regaining adequate end-organ oxygen supply. However, the fluid choice alone may affect the extent of the bleeding by interfering with coagulation pathways. This study investigates the impact of hydroxyethyl starch (HES), gelatine-polysuccinate (GP) and balanced electrolyte solution (BES) as commonly used agents for fluid resuscitation on coagulation using a porcine hemorrhagic shock model. METHODS: Following approval by the State and Institutional Animal Care Committee, life-threatening hemorrhagic shock was induced via arterial blood withdrawal in 24 anesthetized pigs. Isovolumetric fluid resuscitation with either HES, GP or BES (n = 3 × 8) was performed to compensate for the blood loss. Over four hours, hemodynamics, laboratory parameters and rotational thromboelastometry-derived coagulation were analyzed. As secondary endpoint the porcine values were compared to human blood. RESULTS: All the agents used for fluid resuscitation significantly affected coagulation. We measured a restriction of laboratory parameters, clot development and clot firmness, particularly in HES- and GP-treated animals. Hemoglobin content dropped in all groups but showed a more pronounced decline in colloid-treated pigs. This effect was not maintained over the four-hour monitoring period. CONCLUSION: HES, GP, and BEL sufficiently stabilized the macrocirculation, but significantly affected coagulation. These effects were most pronounced after colloid and particularly HES administration. Despite suitability for rapid hemodynamic stabilization, colloids have to be chosen with caution, because their molecular properties may affect coagulation directly and as a consequence of pronounced hemodilution. Our comparison of porcine and human coagulation showed increased coagulation activity in pig blood.

In vitro testing of a funnel-shaped tip catheter model to decrease clot migration during mechanical thrombectomy.

One limitation of mechanical thrombectomy (MT) is clot migration during procedure. This might be caused by abruption of the trapped thrombus at the distal access catheter (DAC) tip during stent-retriever retraction due to the cylindrical shaped tip of the DAC. Aiming to solve this problem, this study evaluates the proof-of-concept of a new designed funnel-shaped tip, in an experimental in vitro setting. Two catheter models, one with a funnel-shaped tip and one with a cylindrical-shaped tip, were compared in an experimental setup. For MT a self-made vessel model and thrombi generated from pig's blood were used. MT was performed 20 times for each device using two different stent-retrievers, 10 times respectively. For the funnel-shaped model: for both stent-retrievers (Trevo XP ProVue 3/20 mm; Trevo XP ProVue 4/20 mm) MT was successful at first pass in 9/10 (90%), respectively. For the cylindrical-shaped model: MT was successful at first pass in 5/10 (50%) with the smaller stent-retriever and in 6/10 (60%) with the larger stent-retriever. The experiments show a better recanalization rate for funnel-shaped tips, than for cylindrical-shaped tips. These results are indicating a good feasibility for this new approach, thus the development of a prototype catheter seems reasonable.

Short-Time Ocular Ischemia Induces Vascular Endothelial Dysfunction and Ganglion Cell Loss in the Pig Retina.

Visual impairment and blindness are often caused by retinal ischemia-reperfusion (I/R) injury. We aimed to characterize a new model of I/R in pigs, in which the intraocular pathways were not manipulated by invasive methods on the ocular system. After 12 min of ischemia followed by 20 h of reperfusion, reactivity of retinal arterioles was measured in vitro by video microscopy. Dihydroethidium (DHE) staining, qPCR, immunohistochemistry, quantification of neurons in the retinal ganglion cell layer, and histological examination was performed. Retinal arterioles of I/R-treated pigs displayed marked attenuation in response to the endothelium-dependent vasodilator, bradykinin, compared to sham-treated pigs. DHE staining intensity and messenger RNA levels for HIF-1α, VEGF-A, NOX2, and iNOS were elevated in retinal arterioles following I/R. Immunoreactivity to HIF-1α, VEGF-A, NOX2, and iNOS was enhanced in retinal arteriole endothelium after I/R. Moreover, I/R evoked a substantial decrease in Brn3a-positive retinal ganglion cells and noticeable retinal thickening. In conclusion, the results of the present study demonstrate that short-time ocular ischemia impairs endothelial function and integrity of retinal blood vessels and induces structural changes in the retina. HIF-1α, VEGF-A, iNOS, and NOX2-derived reactive oxygen species appear to be involved in the pathophysiology.

Random allogeneic blood transfusion in pigs: characterisation of a novel experimental model.

BACKGROUND: Organ cross-talk describes interactions between a primary affected organ and a secondarily injured remote organ, particularly in lung-brain interactions. A common theory is the systemic distribution of inflammatory mediators that are released by the affected organ and transferred through the bloodstream. The present study characterises the baseline immunogenic effects of a novel experimental model of random allogeneic blood transfusion in pigs designed to analyse the role of the bloodstream in organ cross-talk. METHODS: After approval of the State and Institutional Animal Care Committee, 20 anesthetized pig were randomized in a donor and an acceptor (each n = 8): the acceptor animals each received high-volume whole blood transfusion from the donor (35-40 ml kg-1). Four animals received balanced electrolyte solution instead of blood transfusion (control group; n = 4). Afterwards the animals underwent extended cardiorespiratory monitoring for eight hours. Post mortem assessment included pulmonary, cerebral and systemic mediators of early inflammatory response (IL-6, TNF-alpha, iNOS), wet to dry ratio, and lung histology. RESULTS: No adverse events or incompatibilities occurred during the blood transfusion procedures. Systemic cytokine levels and pulmonary function were unaffected. Lung histopathology scoring did not display relevant intergroup differences. Neither within the lung nor within the brain an up-regulation of inflammatory mediators was detected. High volume random allogeneic blood transfusion in pigs neither impaired pulmonary integrity nor induced systemic, lung, or brain inflammatory response. CONCLUSION: This approach can represent a novel experimental model to characterize the blood-bound transmission in remote organ injury.

Standardized Hemorrhagic Shock Induction Guided by Cerebral Oximetry and Extended Hemodynamic Monitoring in Pigs.

Hemorrhagic shock ranks among the main reasons for severe injury-related death. The loss of circulatory volume and oxygen carriers can lead to an insufficient oxygen supply and irreversible organ failure. The brain exerts only limited compensation capacities and is particularly at high risk of severe hypoxic damage.This article demonstrates the reproducible induction of life-threatening hemorrhagic shock in a porcine model by means of calculated blood withdrawal. We titrate shock induction guided by near-infrared spectroscopy and extended hemodynamic monitoring to display systemic circulatory failure, as well as cerebral microcirculatory oxygen depletion. In comparison to similar models that primarily focus on predefined removal volumes for shock induction, this approach highlights a titration by means of the resulting failure of macro- and microcirculation.

Oleic Acid-Injection in Pigs As a Model for Acute Respiratory Distress Syndrome.

The acute respiratory distress syndrome is a relevant intensive care disease with an incidence ranging between 2.2% and 19% of intensive care unit patients. Despite treatment advances over the last decades, ARDS patients still suffer mortality rates between 35 and 40%. There is still a need for further research to improve the outcome of patients suffering from ARDS. One problem is that no single animal model can mimic the complex pathomechanism of the acute respiratory distress syndrome, but several models exist to study different parts of it. Oleic acid injection (OAI)-induced lung injury is a well-established model for studying ventilation strategies, lung mechanics and ventilation/perfusion distribution in animals. OAI leads to severely impaired gas exchange, deterioration of lung mechanics and disruption of the alveolo-capillary barrier. The disadvantage of this model is the controversial mechanistic relevance of this model and the necessity for central venous access, which is challenging especially in smaller animal models. In summary, OAI-induced lung injury leads to reproducible results in small and large animals and hence represents a well-suited model for studying ARDS. Nevertheless, further research is necessary to find a model that mimics all parts of ARDS and lacks the problems associated with the different models existing today.

Ultra-low tidal volume ventilation-A novel and effective ventilation strategy during experimental cardiopulmonary resuscitation.

BACKGROUND: The effects of different ventilation strategies during CPR on patient outcomes and lung physiology are still poorly understood. This study compares positive pressure ventilation (IPPV) to passive oxygenation (CPAP) and a novel ultra-low tidal volume ventilation (ULTVV) regimen in an experimental ventricular fibrillation animal model. STUDY DESIGN: Prospective randomized controlled trial. ANIMALS: 30 male German landrace pigs (16-20 weeks). METHODS: Ventricular fibrillation was induced in anesthetized and instrumented pigs and the animals were randomized into three groups. Mechanical CPR was initiated and ventilation was either provided by means of standard IPPV (RR: 10/min, Vt: 8-9 ml/kg, FiO2: 1,0, PEEP: 5 mbar), CPAP (O2-Flow: 10 l/min, PEEP: 5 mbar) or ULTVV (RR: 50/min, Vt: 2-3 ml/kg, FiO2: 1,0, PEEP: 5 mbar). Guideline-based advanced life support was applied for a maximum of 4 cycles and animals achieving ROSC were monitored for 6 h before terminating the experiment. Ventilation/perfusion ratios were performed via multiple inert gas elimination, blood gas analyses were taken hourly and extended cardiovascular measurements were collected constantly. Brain and lung tissue samples were taken and analysed for proinflammatory cytokine expression. RESULTS: ULTVV provided sufficient oxygenation and ventilation during CPR while demanding significantly lower respiratory and intrathoracic pressures. V/Q mismatch was significantly decreased and lung injury was mitigated in surviving animals compared to IPPV and CPAP. Additionally, cerebral cytokine expression was dramatically reduced. CONCLUSION: Ultra-low-volume ventilation during CPR in a porcine model is feasible and may provide lung-protective benefits as well as neurological outcome improvement due to lower inflammation. Our results warrant further studies and might eventually lead to new therapeutic options in the resuscitation setting.

Lung injury does not aggravate mechanical ventilation-induced early cerebral inflammation or apoptosis in an animal model.

INTRODUCTION: The acute respiratory distress syndrome is not only associated with a high mortality, but also goes along with cognitive impairment in survivors. The cause for this cognitive impairment is still not clear. One possible mechanism could be cerebral inflammation as result of a "lung-brain-crosstalk". Even mechanical ventilation itself can induce cerebral inflammation. We hypothesized, that an acute lung injury aggravates the cerebral inflammation induced by mechanical ventilation itself and leads to neuronal damage. METHODS: After approval of the institutional and state animal care committee 20 pigs were randomized to one of three groups: lung injury by central venous injection of oleic acid (n = 8), lung injury by bronchoalveolar lavage in combination with one hour of injurious ventilation (n = 8) or control (n = 6). Brain tissue of four native animals from a different study served as native group. For six hours all animals were ventilated with a tidal volume of 7 ml kg-1 and a scheme for positive end-expiratory pressure and inspired oxygen fraction, which was adapted from the ARDS network tables. Afterwards the animals were killed and the brains were harvested for histological (number of neurons and microglia) and molecular biologic (TNFalpha, IL-1beta, and IL-6) examinations. RESULTS: There was no difference in the number of neurons or microglia cells between the groups. TNFalpha was significantly higher in all groups compared to native (p < 0.05), IL-6 was only increased in the lavage group compared to native (p < 0.05), IL-1beta showed no difference between the groups. DISCUSSION: With our data we can confirm earlier results, that mechanical ventilation itself seems to trigger cerebral inflammation. This is not aggravated by acute lung injury, at least not within the first 6 hours after onset. Nevertheless, it seems too early to dismiss the idea of lung-injury induced cerebral inflammation, as 6 hours might be just not enough time to see any profound effect.

Effect of gelatin-polysuccinat on cerebral oxygenation and microcirculation in a porcine haemorrhagic shock model.

BACKGROUND: During early treatment of haemorrhagic shock maintenance of cerebral and end-organ oxygen supply by fluid resuscitation is mandatory. Gelatin-polysuccinat (GP) recently regained attention despite a still unclear risk profile and widely unknown effects on cerebral and peripheral microcirculation. This study investigates the effects of GP versus balanced electrolyte solution (BEL) with focus on cerebral regional oxygen saturation and peripheral microcirculation in a porcine haemorrhagic shock model. METHODS: After Animal Care Committee approval haemorrhagic shock was induced by arterial blood withdrawal in 27 anaesthetized pigs. Consequently, the animals received rapid fluid resuscitation by either GP or BEL to replace the removed amount of blood, or remained untreated (n = 3 × 9). Over two hours cerebral regional oxygen saturation by near-infrared spectroscopy and peripheral buccal microcirculation by combined white-light spectrometry and laser-Doppler flowmetry were recorded. Secondary parameters included extended haemodynamics, spirometry, haematological and blood gas parameters. RESULTS: Both fluid resuscitation regimes sufficiently stabilized the macro- and microcirculation in haemorrhagic shock with a more pronounced effect following GP infusion. GP administration led to a persisting, critical impairment of cerebral regional oxygen saturation through considerable haemodilution. Survival rates were 100% in both fluid resuscitation groups, but only 33% in the untreated control. CONCLUSION: Equal amounts of GP and BEL sufficiently stabilize systemic circulation and microcirculatory perfusion. Forced fluid resuscitation by GP should be applied with caution to prevent haemodilution-induced impairment of cerebral oxygen delivery.

Endexpiratory lung volume measurement correlates with the ventilation/perfusion mismatch in lung injured pigs.

BACKGROUND: In acute respiratory respiratory distress syndrome (ARDS) a sustained mismatch of alveolar ventilation and perfusion (VA/Q) impairs the pulmonary gas exchange. Measurement of endexpiratory lung volume (EELV) by multiple breath-nitrogen washout/washin is a non-invasive, bedside technology to assess pulmonary function in mechanically ventilated patients. The present study examines the association between EELV changes and VA/Q distribution and the possibility to predict VA/Q normalization by means of EELV in a porcine model. METHODS: After approval of the state and institutional animal care committee 12 anesthetized pigs were randomized to ARDS either by bronchoalveolar lavage (n = 6) or oleic acid injection (n = 6). EELV, VA/Q ratios by multiple inert gas elimination and ventilation distribution by electrical impedance tomography were assessed at healthy state and at five different positive endexpiratory pressure (PEEP) steps in ARDS (0, 20, 15, 10, 5 cmH2O; each maintained for 30 min). RESULTS: VA/Q, EELV and tidal volume distribution all displayed the PEEP-induced recruitment in ARDS. We found a close correlation between VA/Q < 0.1 (representing shunt and low VA/Q units) and changes in EELV (spearman correlation coefficient -0.79). Logistic regression reveals the potential to predict VA/Q normalization (VA/Q < 0.1 less than 5%) from changes in EELV with an area under the curve of 0.89 with a 95%-CI of 0.81-0.96 in the receiver operating characteristic. Different lung injury models and recruitment characteristics did not influence these findings. CONCLUSION: In a porcine ARDS model EELV measurement depicts PEEP-induced lung recruitment and is strongly associated with normalization of the VA/Q distribution in a model-independent fashion. Determination of EELV could be an intriguing addition in the context of lung protection strategies.

Normalization with Corresponding Naïve Tissue Minimizes Bias Caused by Commercial Reverse Transcription Kits on Quantitative Real-Time PCR Results.

Real-time reverse transcription polymerase chain reaction (PCR) is the gold standard for expression analysis. Designed to improve reproducibility and sensitivity, commercial kits are commonly used for the critical step of cDNA synthesis. The present study was designed to determine the impact of these kits. mRNA from mouse brains were pooled to create serial dilutions ranging from 0.0625 µg to 2 µg, which were transcribed into cDNA using four different commercial reverse-transcription kits. Next, we transcribed mRNA from brain tissue after acute brain injury and naïve mice into cDNA for qPCR. Depending on tested genes, some kits failed to show linear results in dilution series and revealed strong variations in cDNA yield. Absolute expression data in naïve and trauma settings varied substantially between these kits. Normalization with a housekeeping gene failed to reduce kit-dependent variations, whereas normalization eliminated differences when naïve samples from the same region were used. The study shows strong evidence that choice of commercial cDNA synthesis kit has a major impact on PCR results and, consequently, on comparability between studies. Additionally, it provides a solution to overcome this limitation by normalization with data from naïve samples. This simple step helps to compare mRNA expression data between different studies and groups.