Sex differences on solid organ histological characteristics after brain death1

PURPOSE: To investigate gender differences in the evolution of the inflammatory process in rats subjected to brain death (BD). METHODS: Adult Wistar rats were divided into three groups: female; ovariectomized female; and male rats. BD was induced using intracranial balloon inflation and confirmed by maximal pupil dilatation, apnea, absence of reflex, and drop of mean arterial pressure. Six hours after BD, histological evaluation was performed in lungs, heart, liver and kidneys, and levels of inflammatory proteins, estrogen, progesterone, and corticosterone were determined in plasma. RESULTS: In the lungs, females presented more leukocyte infiltration compared to males (p<0.01). Ovariectomized female rat lungs were more hemorrhagic compared to other groups (p<0.001). In the heart, females had higher leukocyte infiltration and tissue edema compared to males (p<0.05). In the liver and kidneys, there were no differences among groups. In female group estradiol and progesterone were sharply reduced 6 hours after BD (p<0.001) to values observed in ovariectomized females and males. Corticosterone levels were similar. CONCLUSIONS: Sex hormones influence the development of inflammation and the status of organs. The increased inflammation in lungs and heart of female rats might be associated with the acute reduction in female hormones triggered by BD.

Effects of ethyl pyruvate on leukocyte-endothelial interactions in the mesenteric microcirculation during early sepsis treatment

OBJECTIVES: Experimental studies on sepsis have demonstrated that ethyl pyruvate is endowed with antioxidant and anti-inflammatory properties. This study aimed to investigate the effects of ethyl pyruvate on leukocyte-endothelial interactions in the mesenteric microcirculation in a live Escherichia coli-induced sepsis model in rats. METHODS: Male Wistar rats were administered an intravenous suspension of E. coli bacteria or were subjected to a sham procedure. Three hours after bacterial infusion, the rats were randomized into the following groups: a control group without treatment, a group treated with lactated Ringer’s solution (4 mL/kg, i.v.), and a group treated with lactated Ringer’s solution (4 mL/kg, i.v.) plus ethyl pyruvate (50 mg/kg). At 24 h after bacterial infusion, leukocyte-endothelial interactions were investigated using intravital microscopy, and the expression of P-selectin and intercellular adhesion molecule-1 was evaluated via immunohistochemistry. White blood cell and platelet counts were also determined at baseline and 3 h and 24 h after E. coli inoculation. RESULTS: The non-treated and lactated Ringer’s solution-treated groups exhibited increases in the numbers of rolling leukocytes (∼2.5-fold increase), adherent cells (∼3.0-fold), and migrated cells (∼3.5-fold) compared with the sham group. In contrast, treatment with Ringer’s ethyl pyruvate solution reduced the numbers of rolling, adherent and migrated leukocytes to the levels observed in the sham group. Additionally, the expression of P-selectin and intercellular adhesion molecule-1 was significantly increased on mesenteric microvessels in the non-treated group compared with the sham group (p<0.001). The expression of both adhesion molecules was reduced in the other groups, with ethyl pyruvate being more effective than lactated Ringer’s solution. Infusion of bacteria caused significant leukopenia (3 h), followed ...

Mesenteric hypoperfusion and inflammation induced by brain death are not affected by inhibition of the autonomic storm in rats

OBJECTIVES: Brain death is typically followed by autonomic changes that lead to hemodynamic instability, which is likely associated with microcirculatory dysfunction and inflammation. We evaluated the role of the microcirculation in the hemodynamic and inflammatory events that occur after brain death and the effects of autonomic storm inhibition via thoracic epidural blockade on mesenteric microcirculatory changes and inflammatory responses. METHODS: Male Wistar rats were anesthetized and mechanically ventilated. Brain death was induced via intracranial balloon inflation. Bupivacaine (brain death-thoracic epidural blockade group) or saline (brain death group) infusion via an epidural catheter was initiated immediately before brain death induction. Sham-operated animals were used as controls (SH group). The mesenteric microcirculation was analyzed via intravital microscopy, and the expression of adhesion molecules was evaluated via immunohistochemistry 180 min after brain death induction. RESULTS: A significant difference in mean arterial pressure behavior was observed between the brain death-thoracic epidural blockade group and the other groups, indicating that the former group experienced autonomic storm inhibition. However, the proportion of perfused small vessels in the brain death-thoracic epidural blockade group was similar to or lower than that in the brain death and SH groups, respectively. The expression of intercellular adhesion molecule 1 was similar between the brain death-thoracic epidural blockade and brain death groups but was significantly lower in the SH group than in the other two groups. The number of migrating leukocytes in the perivascular tissue followed the same trend for all groups. CONCLUSIONS: Although thoracic epidural blockade effectively inhibited the autonomic storm, it did not affect mesenteric hypoperfusion or inflammation induced by brain death. .

Local and remote ischemic preconditioning protect against intestinal ischemic/reperfusion injury after supraceliac aortic clamping

OBJECTIVES: This study tests the hypothesis that local or remote ischemic preconditioning may protect the intestinal mucosa against ischemia and reperfusion injuries resulting from temporary supraceliac aortic clamping. METHODS: Twenty-eight Wistar rats were divided into four groups: the sham surgery group, the supraceliac aortic occlusion group, the local ischemic preconditioning prior to supraceliac aortic occlusion group, and the remote ischemic preconditioning prior to supraceliac aortic occlusion group. Tissue samples from the small bowel were used for quantitative morphometric analysis of mucosal injury, and blood samples were collected for laboratory analyses. RESULTS: Supraceliac aortic occlusion decreased intestinal mucosal length by reducing villous height and elevated serum lactic dehydrogenase and lactate levels. Both local and remote ischemic preconditioning mitigated these histopathological and laboratory changes. CONCLUSIONS: Both local and remote ischemic preconditioning protect intestinal mucosa against ischemia and reperfusion injury following supraceliac aortic clamping. .

Influence of brain death and associated trauma on solid organ histological characteristics / Influência da morte encefálica e do trauma associado nas características histológicas de órgãos sólidos

PURPOSE: To evaluate histopathological alterations triggered by brain death and associated trauma on different solid organs in rats. METHODS: Male Wistar rats (n=37) were anesthetized with isoflurane, intubated and mechanically ventilated. A trepanation was performed and a balloon catheter inserted into intracraninal cavity and rapidly inflated with saline to induce brain death. After induction, rats were monitored for 30, 180, and 360 min for hemodynamic parameters and exsanguinated from abdominal aorta. Heart, lung, liver, and kidney were removed and fixed in paraffin to evaluation of histological alterations (H&E). Sham-operated rats were trepanned only and used as control group. RESULTS: Brain dead rats showed a hemodynamic instability with hypertensive episode in the first minute after the induction followed by hypotension for approximately 1 h. Histological analyses showed that brain death induces vascular congestion in heart (p<0.05), and lung (p<0.05); lung alveolar edema (p=0.001), kidney tubular edema (p<0.05); and leukocyte infiltration in liver (p<0.05). CONCLUSIONS: Brain death induces hemodynamic instability associated with vascular changes in solid organs and compromises most severely the lungs. However, brain death associated trauma triggers important pathophysiological alterations in these organs.

OBJETIVO: Avaliar as alterações histopatológicas desencadeadas pela morte encefálica e pelo trauma associado em diferentes órgãos sólidos em ratos. MÉTODOS: Ratos Wistar machos (n=37) foram anestesiados com isoflurano, entubados e mecanicamente ventilados. Foi realizada trepanação e um cateter foi inserido na cavidade intracraniana e insuflado rapidamente para induzir morte encefálica. Após a indução, os ratos foram monitorados por 30, 180 e 360 min para parâmetros hemodinâmicos e exsanguinados pela aorta abdominal. Coração, pulmão, fígado e rim foram removidos e fixados em parafina para avaliação de alterações histológicas (H&E). Ratos falso-operados foram apenas trepanados e usados como grupo controle. RESULTADOS: Ratos com morte encefálica apresentaram instabilidade hemodinâmica com episódio hipertensivo no primeiro minuto após a indução seguido de hipotensão por aproximadamente 1 hora. Análises histológicas demonstraram que a morte encefálica induz congestão vascular no coração (p<0,05) e pulmão (p<0,05); edema alveolar (p=0,001); edema tubular (p<0,05); e infiltrado leucocitário no fígado (p<0,05). CONCLUSÕES: A morte encefálica induz instabilidade hemodinâmica associada com mudanças vasculares em órgãos sólidos e compromete mais severamente os pulmões. Contudo, o trauma associado à morte encefálica desencadeia importantes alterações fisiopatológicas naqueles órgãos.

Paradoxical effects of brain death and associated trauma on rat mesenteric microcirculation: an intravital microscopic study

OBJECTIVE: Experimental findings support clinical evidence that brain death impairs the viability of organs for transplantation, triggering hemodynamic, hormonal, and inflammatory responses. However, several of these events could be consequences of brain death-associated trauma. This study investigated microcirculatory alterations and systemic inflammatory markers in brain-dead rats and the influence of the associated trauma. METHOD: Brain death was induced using intracranial balloon inflation; sham-operated rats were trepanned only. After 30 or 180 min, the mesenteric microcirculation was observed using intravital microscopy. The expression of Pselectin and ICAM-1 on the endothelium was evaluated using immunohistochemistry. The serum cytokine, chemokine, and corticosterone levels were quantified using enzyme-linked immunosorbent assays. White blood cell counts were also determined. RESULTS: Brain death resulted in a decrease in the mesenteric perfusion to 30 percent, a 2.6-fold increase in the expression of ICAM-1 and leukocyte migration at the mesentery, a 70 percent reduction in the serum corticosterone level and pronounced leukopenia. Similar increases in the cytokine and chemokine levels were seen in the both the experimental and control animals. CONCLUSION: The data presented in this study suggest that brain death itself induces hypoperfusion in the mesenteric microcirculation that is associated with a pronounced reduction in the endogenous corticosterone level, thereby leading to increased local inflammation and organ dysfunction. These events are paradoxically associated with induced leukopenia after brain damage.

Evaluation of the effects of ozone therapy in the treatment of intra-abdominal infection in rats

INTRODUCTION: The antibacterial effect of ozone (O3) has been described in the extant literature, but the role of O3 therapy in the treatment of certain types of infection remains controversial. OBJECTIVES: To evaluate the effect of intraperitoneal (i.p.) O3 application in a cecal ligation/puncture rat model on interleukins (IL-6, IL-10) and cytokine-induced neutrophil chemoattractant (CINC)-1 serum levels, acute lung injury and survival rates. METHODS: Four animal groups were used for the study: a) the SHAM group underwent laparotomy; b) the cecal ligation/puncture group underwent cecal ligation/puncture procedures; and c) the CLP+O2 and CLP+O3 groups underwent CLP+ corresponding gas mixture infusions (i.p.) throughout the observation period. IL-6, CINC-1 and IL-10 concentrations were determined by enzyme-linked immunosorbent assay (ELISA). Acute lung injury was evaluated with the Evans blue dye lung leakage method and by lung histology. P<0.05 was considered significant. RESULTS: CINC-1 was at the lowest level in the SHAM group and was lower for the CLP+O3 group vs. the CLP+O2 group and the cecal ligation/puncture group. IL-10 was lower for the SHAM group vs. the other three groups, which were similar compared to each other. IL-6 was lower for the SHAM group vs. all other groups, was lower for the CLP+O3 or CLP+O2 group vs. the cecal ligation/puncture group, and was similar for the CLP+O3 group vs. the CLP+O2 group. The lung histology score was lower for the SHAM group vs. the other groups. The Evans blue dye result was lower for the CLP+O3 group vs. the CLP+O2 group and the cecal ligation/puncture group but similar to that of the SHAM group. The survival rate for the CLP+O3 group was lower than for the SHAM group and similar to that for the other 2 groups (CLP and CLP+O2). CONCLUSION: Ozone therapy modulated the inflammatory response and acute lung injury in the cecal ligation/puncture infection model in rats, although there was no ...

Effects of different peep levels on mesenteric leukocyte-endothelial interactions in rats during mechanical ventilation

INTRODUCTION: Mechanical ventilation with positive end expiratory pressure (PEEP) improves oxygenation and treats acute pulmonary failure. However, increased intrathoracic pressure may cause regional blood flow alterations that may contribute to mesenteric ischemia and gastrointestinal failure. We investigated the effects of different PEEP levels on mesenteric leukocyte-endothelial interactions. METHODS: Forty-four male Wistar rats were initially anesthetized (Pentobarbital I.P. 50mg/kg) and randomly assigned to one of the following groups: 1) NAIVE (only anesthesia; n=9), 2) PEEP 0 (PEEP of 0 cmH2O, n=13), 3) PEEP 5 (PEEP of 5 cmH2O, n=12), and 4) PEEP 10 (PEEP of 10 cmH2O, n=13). Positive end expiratory pressure groups were tracheostomized and mechanically ventilated with a tidal volume of 10 mL/kg, respiratory rate of 70 rpm, and inspired oxygen fraction of 1. Animals were maintained under isoflurane anesthesia. After two hours, laparotomy was performed, and leukocyte-endothelial interactions were evaluated by intravital microscopy. RESULTS: No significant changes were observed in mean arterial blood pressure among groups during the study. Tracheal peak pressure was smaller in PEEP 5 compared with PEEP 0 and PEEP 10 groups (11, 15, and 16 cmH2O, respectively; p<0.05). After two hours of MV, there were no differences among NAIVE, PEEP 0 and PEEP 5 groups in the number of rollers (118±9,127±14 and 147±26 cells/10minutes, respectively), adherent leukocytes (3±1,3±1 and 4±2 cells/100µm venule length, respectively), and migrated leukocytes (2±1,2±1 and 2±1 cells/5,000µm², respectively) at the mesentery. However, the PEEP 10 group exhibited an increase in the number of rolling, adherent and migrated leukocytes (188±15 cells / 10 min, 8±1 cells / 100 µm and 12±1 cells / 5,000 µm², respectively; p<0.05). CONCLUSIONS: High intrathoracic pressure was harmful to mesenteric microcirculation in the experimental model of rats with normal lungs and ...

Mesenteric microcirculatory dysfunctions and translocation of indigenous bacteria in a rat model of strangulated small bowel obstruction

PRUPOSE: Bacterial translocation has been shown to occur in critically ill patients after extensive trauma, shock, sepsis, or thermal injury. The present study investigates mesenteric microcirculatory dysfunctions, the bacterial translocation phenomenon, and hemodynamic/metabolic disturbances in a rat model of intestinal obstruction and ischemia. METHODS: Anesthetized (pentobarbital 50 mg/kg, i.p.) male Wistar rats (250-350 g) were submitted to intestinal obstruction or laparotomy without intestinal obstruction (Sham) and were evaluated 24 hours later. Bacterial translocation was assessed by bacterial culture of the mesenteric lymph nodes (MLN), liver, spleen, and blood. Leukocyte-endothelial interactions in the mesenteric microcirculation were assessed by intravital microscopy, and P-selectin and intercellular adhesion molecule (ICAM)-1 expressions were quantified by immunohistochemistry. Hematocrit, blood gases, lactate, glucose, white blood cells, serum urea, creatinine, bilirubin, and hepatic enzymes were measured. RESULTS: About 86 percent of intestinal obstruction rats presented positive cultures for E. coli in samples of the mesenteric lymph nodes, liver, and spleen, and 57 percent had positive hemocultures. In comparison to the Sham rats, intestinal obstruction induced neutrophilia and increased the number of rolling (~2-fold), adherent (~5-fold), and migrated leukocytes (~11-fold); this increase was accompanied by an increased expression of P-selectin (~2-fold) and intercellular adhesion molecule-1 (~2-fold) in the mesenteric microcirculation. Intestinal obstruction rats exhibited decreased PaCO2, alkalosis, hyperlactatemia, and hyperglycemia, and increased blood potassium, hepatic enzyme activity, serum urea, creatinine, and bilirubin. A high mortality rate was observed after intestinal obstruction (83 percent at 72 h vs. 0 percent in Sham rats). CONCLUSION: Intestinal obstruction and ischemia in rats is a relevant model for ...

In vivo observation of mesenteric leukocyte-endothelial interactions after cecal ligation/puncture and surgical sepsis source control

PURPOSE: Cecal ligation and puncture (CLP) has been used as a useful model for the induction of polymicrobial sepsis. Necrotic tissue resection and peritoneal lavage (REL) are the surgical procedures for controlling perforated appendicitis. The aim of this study was to evaluate leukocyte-endothelial interactions in the rat mesentery in vivo after CLP and REL. METHODS: Thirty-seven male Wistar rats (250-300 g) underwent laparotomy and were randomly assigned to the following groups: 1) SHAM; 2) CLP: animals submitted to CLP, 3) CLP+REL: animals submitted to CLP and REL. Mesenteric leukocyte-endothelial interactions were studied by intravital microscopy assessed once in each animal (3-5 postcapillary venules, 15-25 æm diameter) 24 hours after intervention. Follow-up was performed in all animals; this included analysis of glycemia, lactate, hematocrit, white blood cell count as well as a functional score that was the sum of scoring on the following parameters: alertness, mobility, piloerection, diarrhea, encrusted eyes, and dirty nose and tail. RESULTS: None of the animals showed significant changes in body weight (265 ± 20 g) or in hematocrit levels (46 percent ± 2 percent) during the experimental protocol. Compared to SHAM animals, CLP animals showed an increased number of rolling (2x), adherent, and migrating leukocytes (7x) in the mesenteric microcirculation, an increase in blood glucose (136 ± 8 mg/dL), lactate (3.58 ± 0.94 mmol/L), white cell count (23,570 ± 4,991 cells/mm³) and functional alterations (score 11 ± 1), characterized by impaired alertness and mobility, and presence of piloerection, diarrhea, encrusted eyes, and dirty nose and tail. The REL procedure normalized the number of rolling, adherent, and migrated leukocytes in the mesentery; glycemia; lactate; and white blood cell count. The REL procedure also improved the functional score (7 ± 1). CONCLUSION: Local and systemic inflammation was induced by CLP, while REL completely...

OBJETIVO: O procedimento de ligadura cecal e perfuração (CLP) tem sido usado como um modelo útil de indução de sepse polimicrobiana. A ressecção do tecido necrosado e lavagem peritoneal (REL) são procedimentos cirúrgicos freqüentemente utilizados para controlar uma apendicite perfurada. O objetivo desse estudo foi avaliar in vivo as interações leucócito-endotélio no mesentério de ratos após a CLP e REL. MÉTODOS: Trinta e sete ratos Wistar machos (250-300 g) foram submetidos à laparotomia e aleatoriamente divididos em grupos: 1) SHAM, 2) CLP: ratos submetidos à CLP, 3) CLP+REL: animais submetidos à CLP e REL. As interações leucócito-endotélio no mesentério foram estudadas através de microscopia intravital somente uma vez em cada animal (3-5 vênulas pós-capilares, 15-25 æm diâmetro), 24-horas após as intervenções. A evolução clínica foi realizada em todos os animais, incluindo glicemia, lactato, hematócrito, número total de células brancas e um escore funcional, o qual foi considerado como a somatória dos seguintes parâmetros: estado de alerta, mobilidade, piloereção, diarréia, olhos encrustados, e nariz e cauda sujos. RESULTADOS: Os animais não apresentaram alterações significantes no peso (265 ± 20 g) e hematócrito (46 ± 2 por cento) ao longo do estudo. Comparados ao SHAM, os animais CLP apresentaram aumento no número de leucócitos em rolamento (2x), aderidos (7x) e migrados (7x) na microcirculação mesentérica, aumentos da glicemia (136 ± 8 mg/dL), lactato (3,58 ± 0,94 mmol/L), leucocitose (23.570 ± 4.991 células/mm³) e alterações clínicas (escore 11±1), caracterizadas por comprometimento do estado de alerta e mobilidade, e presença de piloereção, diarréia, olhos encrustados, nariz e cauda sujos. REL normalizou o número de leucócitos em rolamento, aderidos e migrados no mesentério, a glicemia, o lactato e o número de leucócitos circulantes. REL também melhorou o escore clínico (7 ± 1). CONCLUSÃO: A CLP induziu inflamação local e sistêmica. A...