Olmesartan Ameliorates Organ Injury and Mortality in Rats With Peritonitis-Induced Sepsis.

INTRODUCTION: Sepsis and related complications lead to high morbidity and mortality in humans and animals. Olmesartan medoxomil (OLM), a nonpeptide angiotensin II type 1 receptor blocker, has antiinflammatory and antioxidative effects in various experimental animal models. The present study aimed to investigate whether OLM protects against sepsis in a clinically relevant model of polymicrobial sepsis induced by cecal ligation and puncture (CLP). METHODS: Sepsis was induced by CLP in anesthetized rats. OLM was administered intraperitoneally 3 h after CLP onset. Hemodynamic, biochemical, and inflammatory parameters were analyzed. RESULTS: The administration of OLM in CLP rats significantly improved their survival rate. Moreover, OLM mitigated CLP-induced hypotension and organ injury (indicated by biochemical parameters), but not tachycardia. OLM significantly reduced the plasma levels of interleukin-6 and nitric oxide. CONCLUSIONS: OLM markedly attenuated CLP-induced hypotension and organ injury, and hence improved survival by inhibiting the inflammatory response and nitrosative stress in this clinically relevant model of sepsis.

Angiotensin-(1-7) attenuates organ injury and mortality in rats with polymicrobial sepsis.

BACKGROUND: Sepsis and related multiple organ dysfunction result in high morbidity and mortality. Angiotensin (Ang)-(1-7), a biologically active peptide, has various opposing effects of Ang II. Because the effect of Ang-(1-7) on sepsis is unknown, in this study we aimed to determine the impact of Ang-(1-7) on pathophysiologic changes in a clinically relevant model of polymicrobial sepsis induced by cecal ligation and puncture (CLP). METHODS: Sepsis was induced by CLP in rats under anesthesia. Rats were randomized to one of the following five groups: (1) sham-operated group, (2) Ang-(1-7) (1 mg/kg intravenously infused for 1 h) at 3 h and 6 h after sham operation, (3) CLP, (4) Ang-(1-7) at 3 h after CLP, and (5) Ang-(1-7) at 3 h and 6 h after CLP. Rats were observed for 24 h after CLP surgery and then killed for subsequent histological examination. RESULTS: Ang-(1-7) significantly improved the survival of septic rats (83.3% vs. 36.4% at 24 h following CLP; p = 0.009). Ang-(1-7) attenuated the CLP-induced decreased arterial pressure and organ dysfunction, indicated by diminished biochemical variables and fewer histological changes. Ang-(1-7) significantly reduced the level of plasma interleukin-6 and pulmonary superoxide production (p < 0.05). Moreover, caspase-3 and cytoplasmic IκB expression in liver was significantly lower in the Ang-(1-7)-treated CLP rats (p < 0.05). CONCLUSIONS: In this clinically relevant model of sepsis, Ang-(1-7) ameliorates CLP-induced organ dysfunction and improves survival, possibly through suppressing the inflammatory response, oxidative stress, and apoptosis, suggesting that Ang-(1-7) could be a potential novel therapeutic approach to treatment of peritonitis and polymicrobial sepsis.

Levosimendan attenuates multiple organ injury and improves survival in peritonitis-induced septic shock: studies in a rat model.

INTRODUCTION: The aim of this study was to investigate the effects of levosimendan on rodent septic shock induced by cecal ligation and puncture (CLP). METHODS: Three hours after peritonitis-induced sepsis, male Wistar rats were randomly assigned to receive an intravenous infusion of levosimendan (1.2 µg/kg/min for 10 min and then 0.3 µg/kg/min for 6 h) or an equivalent volume of saline and vehicle (5% dextrose) solution. RESULTS: The levosimendan-treated CLP animals had significantly higher arterial pressure and lower biochemical indices of liver and kidney dysfunction compared to the CLP animals (P < 0.05). Plasma interleukin-1ß, nitric oxide and organ superoxide levels in the levosimendan-treated CLP group were less than those in CLP rats treated with vehicle (P < 0.05). In addition, the inducible nitric oxide synthase (iNOS) in lung and caspase-3 expressions in spleen were significantly lower in the levosimendan-treated CLP group (P < 0.05). The administration of CLP rats with levosimendan was associated with significantly higher survival (61.9% vs. 40% at 18 h after CLP, P < 0.05). At postmortem examination, the histological changes and neutrophil filtration index in liver and lung were significantly attenuated in the levosimendan-treated CLP group (vs. CLP group, P < 0.05). CONCLUSIONS: In this clinically relevant model of septic shock induced by fecal peritonitis, the administration of levosimendan had beneficial effects on haemodynamic variables, liver and kidney dysfunction, and metabolic acidosis. (1) Lower levels of interleukin-1ß, nitric oxide and superoxide, (2) attenuation of iNOS and caspase-3 expressions, and (3) decreases of neutrophil infiltration by levosimendan in peritonitis-induced sepsis animals suggest that anti-inflammation and anti-apoptosis effects of levosimendan contribute to prolonged survival.

Loss of plasma fibrinogen contributes to platelet hyporeactivity in rats with septic shock.

INTRODUCTION: Dysregulated host response to infection causes life-threatening organ dysfunction. Excessive inflammation and abnormal blood coagulation can lead to disseminated intravascular coagulation (DIC) and multiple-organ failure in the late sepsis stages. Platelet function impairment in sepsis contributes to bleeding, secondary infection, and tissue injury. Platelet transfusion is considered in patients with sepsis with DIC and bleeding; however, its benefits are limited and of low quality. Fibrinogen plays a crucial role in platelet function, and establishing a fibrin network binds to activated integrin αIIbß3 and promotes outside-in signaling that amplifies platelet functions. However, the role of fibrinogen in sepsis-induced platelet dysfunction remains unclear. MATERIALS AND METHODS: We evaluated the effects of fibrinogen on platelet hyporeactivity during septic shock in adult male Wistar rats using lipopolysaccharide (LPS) injection and cecal ligation and puncture (CLP) surgery. Changes in the hemodynamic, biochemical, and coagulation parameters were examined. Platelet activation and aggregation were measured using whole-blood assay, 96-well plate-based aggregometry, and light-transmission aggregometry. Additionally, platelet adhesion, spreading, and fibrin clot retraction were evaluated. RESULTS: Rats with LPS- and CLP-induced sepsis displayed considerable decreases in plasma fibrinogen levels and platelet aggregation, adhesion, spreading, and clot retraction. The aggregation of platelets obtained from rats with sepsis was markedly augmented by fibrinogen supplementation. Additionally, fibrinogen administration improved platelet adhesion, spreading, and clot retraction in rats with sepsis. CONCLUSIONS: Fibrinogen supplementation could serve as a potential therapeutic intervention for alleviating platelet hyporeactivity in patients with sepsis and bleeding.

Possible biomarkers of early mortality in peritonitis-induced sepsis rats.

BACKGROUND: Sepsis induced by cecal ligation and puncture (CLP) is accompanied by circulatory failure, multiple organ dysfunction syndrome, metabolic acidosis, and electrolyte imbalance in rats. However, it remains uncertain which parameters can be used to predict the mortality of septic rats. Thus, the aim of this study was to examine which possible biomarkers were associated with mortality in the CLP-induced sepsis model. MATERIALS AND METHODS: After the carotid artery and vein were cannulated, rats were subsequently subjected to CLP or sham operation. The changes of hemodynamics, biochemical variables, blood gas, and electrolytes were monitored during the 18-h observation. RESULTS: The CLP surgery caused circulatory failure, multiple organ dysfunction syndrome, metabolic acidosis, electrolyte imbalance, and death. Compared with survivors, nonsurvivors showed significant difference in (1) blood glucose; (2) lactate dehydrogenase, alanine aminotransferase, aspartate aminotransferase, creatinine, and blood urea nitrogen in serum; and (3) base excess, HCO3(-), PaCO2, potassium, and calcium in whole blood at 9 h after CLP. No significant difference in blood pressure, heart rate, pressor response to noradrenaline, rectal temperature, total protein, albumin, PaO2, and sodium was observed between nonsurvivors and survivors. However, after multifactor dimensionality reduction analysis, the union of HCO3(-) and blood glucose had the biggest testing balanced accuracy. CONCLUSIONS: These results indicate that HCO3(-) plus blood glucose serves as the best biomarker of early death in rats with CLP-induced sepsis. Thus, these parameters could guide experimental procedures for making the right interventions when utilizing CLP as a sepsis model in rats.

Hyperoncotic albumin attenuates lung and intestine injuries caused by peritonitis-induced sepsis in rats.

BACKGROUND: Hyperoncotic albumin may be a therapeutic option to improve tissue perfusion and organ injury in sepsis. To clarify the hypothesis and its mechanism, hyperoncotic albumin was administered to the rats in a polymicrobial sepsis-peritonitis model. MATERIALS AND METHODS: Peritonitis was induced by a surgery of cecal ligation and puncture (CLP) in 27 male Wistar rats. For control purposes, sham operations without ligating and puncturing the cecum were performed in 20 rats. Three hours later, rats were randomized to receive intravenously 3 mL/kg of 5% albumin, 25% albumin, or normal saline. All the hemodynamic and biochemical parameters were measured during the 18-h observation. RESULTS: In septic rats, 25% albumin attenuated hypotension, vascular hyporeactivity to norepinephrine, and the elevated serum levels of lactate dehydrogenase and blood urea nitrogen. However, these improvements were not noted in CLP rats after 5% albumin treatment. In addition, 25% albumin decreased metabolic acidosis and improved the CLP-induced hypoperfusion in the intestine and kidney. Superoxide levels in the aorta and lung and the protein expression of inducible nitric oxide synthase in the lung were also attenuated by 25% albumin in CLP rats. Microscopic findings confirmed that 25% albumin attenuated the substantial swelling and cell infiltration in the intestine and lung caused by CLP. CONCLUSIONS: In this sepsis rat model, 25% albumin reduced macro- and microhemodynamic changes and attenuated intestine and lung injuries in peritonitis-induced sepsis.

Stimulation of angiotensin II type 2 receptor attenuates organ injury in rats with polymicrobial sepsis.

BACKGROUND: Both inflammation and oxidative stress contribute to the pathogenesis of sepsis and its associated organ damage. Angiotensin-(1-7), acting through the Mas receptor and angiotensin II-type 2 receptors (AT2R), could attenuate organ dysfunction and improve survival in rats with sepsis. However, the role of AT2R in inflammation and oxidative stress in rats with sepsis is unclear. Therefore, this study examined the modulatory effects and molecular mechanism of AT2R stimulation in rats with polymicrobial sepsis. METHODS: Male Wistar rats underwent cecal ligation and puncture (CLP) or sham surgery followed by the administration of saline or CGP42112 (a selective, high-affinity agonist of AT2R, 50 µg/kg intravenously) at 3 hours after sham surgery or CLP. The changes in hemodynamics, biochemical variables, and plasma levels of chemokines and nitric oxide were detected during the 24-hour observation. Organ injury was evaluated by histological examination. RESULTS: We found that CLP evoked delayed hypotension, hypoglycemia, and multiple organ injuries, characterized by elevated plasma biochemical parameters and histopathological changes. These effects were attenuated by treatment with CGP42112. CGP42112 significantly attenuated plasma chemokines and nitric oxide production and reduced liver inducible nitric oxide synthase and nuclear factor kappa-B expression. More importantly, CGP42112 significantly improved the survival of rats with sepsis (50% vs. 20% at 24 h after CLP, p < 0.05). CONCLUSION: The protective effects of CGP42112 may be related to anti-inflammatory responses, suggesting that the stimulation of AT2R is a promising therapeutic candidate for the treatment of sepsis.

Antimicrobial peptide cathelicidin LL-37 preserves intestinal barrier and organ function in rats with heat stroke.

Global warming increases the incidence of heat stroke (HS) and HS causes the reduction of visceral blood flow during hyperthermia, leading to intestinal barrier disruption, microbial translocation, systemic inflammation and multiple organ failure. Cathelicidin LL-37 exhibits antimicrobial activities, helps innate immunity within the gut to maintain intestinal homeostasis, and augments intestinal wound healing and barrier function. Thus, we evaluated the effects and possible mechanisms of cathelicidin LL-37 on HS. Wistar rats were placed in a heating-chamber of 42 ̊C to induce HS. Changes in rectal temperature, hemodynamic parameters, and survival rate were measured during the experimental period. Blood samples and ilea were collected to analyze the effects of LL-37 on systemic inflammation, multiple organ dysfunction, and intestinal injury. Furthermore, LS174T and HT-29 cells were used to assess the underlying mechanisms. Our data showed cathelicidin LL-37 ameliorated the damage of intestinal cells induced by HS. Intestinal injury, systemic inflammation, and nitrosative stress (high nitric oxide level) caused by continuous hyperthermia were attenuated in HS rats treated with cathelicidin LL-37, and hence, improved multiple organ dysfunction, coagulopathy, and survival rate. These beneficial effects of cathelicidin LL-37 were attributed to the protection of intestinal goblet cells (by increasing transepithelial resistance, mucin-2 and Nrf2 expression) and the improvement of intestinal barrier function (less cyclooxygenase-2 expression and FITC-dextran translocation). Interestingly, high cathelicidin expression in the ileal samples of inflammatory bowel disease patients was associated with better clinical outcome. These results suggest that cathelicidin LL-37 could prevent heat stress-induced intestinal damage and heat-related illnesses.

Improvement in heat stress-induced multiple organ dysfunction and intestinal damage through protection of intestinal goblet cells from prostaglandin E1 analogue misoprostol.

AIMS: Heat stroke is a life-threatening disorder triggered by thermoregulatory failure. Hyperthermia-induced splanchnic hypoperfusion has been reported to induce intestinal barrier dysfunction and systemic immune response that ultimately cause multiple-organ failure and death. Intestinal goblet cells contribute greatly to the formation of mucus barrier, which hinders translocation of gut microorganisms. Studies have reported that misoprostol can not only alleviate ischemic injury but also protect GI mucosal layer. Therefore, we evaluated the effects of misoprostol on intestinal goblet cells after heat stress and on multiple-organ dysfunction in heat stroke rats. MAIN METHODS: Heat stress was established in the heating chamber and followed by misoprostol treatment. Changes in hemodynamics, organ function indices, inflammation, oxidative stress, and survival rate were analyzed. Furthermore, ilea and LS174T cells were used to examine intestinal functions. KEY FINDINGS: Heat stress caused dysfunction of intestinal goblet cells and damage to ilea by increasing oxidative stress and apoptosis. Increased nitrosative stress and inflammation accompanied by hypotension, hypoperfusion, tachycardia, multiple-organ dysfunction, and death were observed in the heat stroke rat model. Treatment of LS174T cells with misoprostol not only decreased oxidative stress and apoptosis but also reduced cytotoxicity caused by heat stress. Moreover, misoprostol prevented disruption of the enteric barrier, multiple-organ injury, and death in rats with heat stroke. SIGNIFICANCE: This study indicates that misoprostol could alleviate intestinal damage and organ injury caused by heat stress and be a potential therapy for heat-related illnesses.

Angiotensin-(1-7) treatment blocks lipopolysaccharide-induced organ damage, platelet dysfunction, and IL-6 and nitric oxide production in rats.

Sepsis can lead to shock, multiple organ failure, and even death. Platelets play an active role in the pathogenesis of sepsis-induced multiple organ failure. Angiotensin (Ang)-(1-7), a biologically active peptide, counteracts various effects of Ang II and attenuates inflammatory responses, reactive oxygen species production, and apoptosis. We evaluated the effects of Ang-(1-7) on organ injury and platelet dysfunction in rats with endotoxaemia. We treated male Wistar rats with saline or lipopolysaccharide (LPS, 10 mg, intravenously) then Ang-(1-7) (1 mg/kg, intravenous infusion for 3 h beginning 30 min after LPS administration). We analysed several haemodynamic, biochemical, and inflammatory parameters, as well as platelet counts and aggregation. Ang-(1-7) improved hypotension and organ dysfunction, and attenuated plasma interleukin-6, chemokines and nitric oxide production in rats after LPS administration. The LPS-induced reduction in platelet aggregation, but not the decreased platelet count, was restored after Ang-(1-7) treatment. The protein expression of iNOS and IκB, but not phosphorylated ERK1/2 and p38, was diminished in Ang-(1-7)-treated LPS rats. The histological changes in liver and lung were significantly attenuated in Ang-(1-7)-treated LPS rats. Our results suggest that Ang-(1-7) ameliorates endotoxaemic-induced organ injury and platelet dysfunction, likely through the inhibition of the inflammatory response and nitric oxide production.

Low-dose hydralazine improves endotoxin-induced coagulopathy and multiple organ dysfunction via its anti-inflammatory and anti-oxidative/nitrosative properties.

Coagulopathy is the major cause of organ injury as well as a strong predictor of mortality in septic patients. Systemic inflammatory response and redox imbalance are regarded as the major causes of sepsis-induced coagulopathy. There is growing evidence that a vasodilator hydralazine has beneficial effects on heart failure, hypertension, and ischemia/reperfusion injury via its antioxidant and anti-inflammatory properties. However, the effects of hydralazine on sepsis have not been examined. Therefore, we evaluated the effects of low-dose hydralazine on coagulopathy and multiple organ dysfunction in septic rats induced by endotoxin. Sepsis-induced coagulopathy was established by intravenous injection of rats with lipopolysaccharide (LPS). The changes of blood pressure, heart rate, blood glucose, hemostatic variables, prothrombin time, organ function indices, interleukin-6 (IL-6) concentration, and nitric oxide (NO) level were assessed during the experimental period. In addition, the aortas, lungs, livers, and kidneys were dissected to analyze superoxide levels and protein expressions. LPS induced (i) coagulopathy, multiple organ dysfunction, and circulatory failure successfully, and (ii) excessive superoxide, NO, and IL-6 production, accompanied by the overexpression of iNOS and Wnt5a in animals. Treatment of LPS-induced septic rats with low-dose hydralazine not only improved coagulopathy but also ameliorated multiple organ dysfunction. These could be due to attenuation of the overproduction of superoxide, NO, and IL-6, which were attributed to reduction of the overexpression of iNOS and Wnt5a. Thus, these findings indicate that low-dose hydralazine could be a potential therapy for sepsis-induced coagulopathy and multiple organ dysfunction via its anti-inflammatory and anti-oxidative/nitrosative properties.

Combined effects of propofol and dexamethasone on rats with endotoxemia.

OBJECTIVE: To clarify the effect of combined treatment with propofol and dexamethasone on hemodynamics, organ injury, and survival rate in rats with endotoxemia. DESIGN: Randomized, prospective animal experiment. SETTING: Academic research laboratory. SUBJECTS: Male Wistar rats. INTERVENTIONS: Rats were divided into five groups: a control group, a group of conscious rats infused with Escherichia coli lipopolysaccharide, two groups of lipopolysaccharide rats treated with either propofol or dexamethasone, and a group of lipopolysaccharide rats with combined treatment of propofol and dexamethasone. MEASUREMENTS AND MAIN RESULTS: All hemodynamic and biochemical variables were measured during the 6-hr observation. Propofol plus dexamethasone attenuated hypotension and delayed hypoglycemia and metabolic acidosis caused by coadministration of E. coli lipopolysaccharide. In addition, propofol plus dexamethasone attenuated the lipopolysaccharide-induced multiple organ dysfunctions, such as lung, liver, and kidney. The increases in serum tumor necrosis factor-alpha, tissue nitric oxide, and superoxide anion levels were attenuated by propofol plus dexamethasone in lipopolysaccharide rats. Microscopic findings confirmed that propofol plus dexamethasone attenuated the substantial swelling and cell infiltration in lung and kidney caused by endotoxin. The 22-hr survival rate after endotoxin injection was markedly increased in lipopolysaccharide rats with combined treatment compared with the lipopolysaccharide rats (80% vs. 0%). CONCLUSIONS: The combined treatment with propofol plus dexamethasone reduced mortality rate and attenuated organ injury in conscious rats treated with lipopolysaccharide. These protective effects may be associated with their anti-inflammatory capacity and antioxidant activity.

Therapeutic effects of melatonin on peritonitis-induced septic shock with multiple organ dysfunction syndrome in rats.

The pathogenesis of multiple organ dysfunction syndrome (MODS) in septic shock is complicated and not fully understood. Some studies show that an overproduction of nitric oxide (NO) leads to the refractory hypotension and multiple organ failure, while other studies suggest that free radicals, e.g. superoxide (O(2)(-)), contribute to the detrimental effect on vascular responsiveness and tissue/organ damage. Thus, this study was performed on the Wistar rat by using cecal ligation and puncture (CLP) to induce septic shock-associated MODS. We evaluated the effect of an antioxidant melatonin in CLP-induced septic rats and demonstrated that melatonin (3 mg/kg, i.v. at 3, 6, 12 hr after CLP) significantly (a) attenuated hyporeactivity to norepinephrine and delayed hypotension, (b) reduced plasma index of hepatic and renal dysfunction, (c) diminished plasma NO and interleukin-1beta (IL-1beta) concentrations as well as aortic O(2)(-) levels, (d) reduced marked infiltration of polymorphonuclear neutrophils (PMNs) in the lung and liver tissues, and (e) promoted the survival rate at 18 hr to twofold compared with the CLP alone group. The current study underlined the inhibition of plasma NO and IL-1beta as well as aortic O(2)(-) production and the reduction of PMN infiltration may lead to the amelioration of MODS, which may contribute to the beneficial effect of antioxidants (e.g. melatonin in this study) in conscious rats with peritonitis-induced lethality. Thus, the antioxidant could be a novel agent for the treatment of septic animals or patients in the early stage.

Low-dose bupivacaine-fentanyl spinal anesthesia for lower extremity surgery in a nonparturient with Eisenmenger's syndrome.

Perioperative mortality in patients with Eisenmenger's syndrome is very high, particularly following cesarean section. This case report describes the successful use of low-dose bupivacaine-fentanyl spinal anesthesia for lower extremity surgery in a nonparturient with Eisenmenger's syndrome. A 21-year-old woman with Eisenmenger's syndrome was scheduled to have a fibular head tumor excision. After placement of routine monitor and an arterial line, we inserted an epidural catheter at the L3-L4 interspace to cover a potential inadequate block and then we administered 6 mg of hyperbaric bupivacaine 0.5% with 20 microg of fentanyl intrathecally via a 27-gauge needle at the L4-L5 interspace. There were no hypotension, respiratory depression, hypoxemia, and other severe hemodynamic alterations. No drug was administered via the epidural catheter in the 2-hour operative period and the postoperative course was uneventful. Therefore, we propose that intrathecal opioids combined with local anesthetics may be an alternative anesthetic method in patients with Eisenmenger's syndrome.

Levosimendan mitigates coagulopathy and organ dysfunction in rats with endotoxemia.

BACKGROUND: In patients with severe sepsis, pro-inflammatory cytokines and subsequent activation of tissue factors trigger a cascade of events that lead to coagulation dysfunction and multiple organ failure. It has been shown that levosimendan has protective effects against tissue injury caused by endotoxin. The purpose of this study was to evaluate the effects of levosimendan on consumptive coagulopathy and organ dysfunction in an endotoxemic animal model induced by lipopolysaccharide (LPS). METHODS: Forty-six male adult Wistar rats were randomly divided into four groups: 1) control group (n = 10), an intravenous infusion of 5% dextrose 1.2 mL/kg for 20 min and 0.03 mL/kg/min for 4 h; 2) the levosimendan-treated control group (n = 12), an intravenous levosimendan infusion (24 µg/kg for 20 min plus 0.6 µg/kg/min for 4 h); 3) the LPS group (n = 12), an intravenous LPS (4 mg/kg) infusion followed by dextrose administration; and 4) the levosimendan-treated LPS group (n = 12), an intravenous LPS infusion followed by levosimendan treatment. Various parameters of hemodynamics, biochemistry, hemostasis and inflammatory response were examined during the experimental period. RESULTS: The administration of levosimendan significantly attenuated (i) consumptive coagulopathy displayed by thromboelastography, (ii) the decreases of platelet count and plasma fibrinogen level, (iii) injury in the lung, liver and kidney, and (iv) the rise in plasma interleukin-6 in rats treated with LPS. CONCLUSION: The treatment of LPS rats with levosimendan was found to reduce organ injury and coagulopathy. These protective effects may be attributed to the anti-inflammatory effects of this drug.

Therapeutic Effects of Procainamide on Endotoxin-Induced Rhabdomyolysis in Rats.

Overt systemic inflammatory response is a predisposing mechanism for infection-induced skeletal muscle damage and rhabdomyolysis. Aberrant DNA methylation plays a crucial role in the pathophysiology of excessive inflammatory response. The antiarrhythmic drug procainamide is a non-nucleoside inhibitor of DNA methyltransferase 1 (DNMT1) used to alleviate DNA hypermethylation. Therefore, we evaluated the effects of procainamide on the syndromes and complications of rhabdomyolysis rats induced by lipopolysaccharide (LPS). Rhabdomyolysis animal model was established by intravenous infusion of LPS (5 mg/kg) accompanied by procainamide therapy (50 mg/kg). During the experimental period, the changes of hemodynamics, muscle injury index, kidney function, blood gas, blood electrolytes, blood glucose, and plasma interleukin-6 (IL-6) levels were examined. Kidneys and lungs were exercised to analyze superoxide production, neutrophil infiltration, and DNMTs expression. The rats in this model showed similar clinical syndromes and complications of rhabdomyolysis including high levels of plasma creatine kinase, acute kidney injury, hyperkalemia, hypocalcemia, metabolic acidosis, hypotension, tachycardia, and hypoglycemia. The increases of lung DNMT1 expression and plasma IL-6 concentration were also observed in rhabdomyolysis animals induced by LPS. Treatment with procainamide not only inhibited the overexpression of DNMT1 but also diminished the overproduction of IL-6 in rhabdomyolysis rats. In addition, procainamide improved muscle damage, renal dysfunction, electrolytes disturbance, metabolic acidosis, hypotension, and hypoglycemia in the rats with rhabdomyolysis. Moreover, another DNMT inhibitor hydralazine mitigated hypoglycemia, muscle damage, and renal dysfunction in rhabdomyolysis rats. These findings reveal that therapeutic effects of procainamide could be based on the suppression of DNMT1 and pro-inflammatory cytokine in endotoxin-induced rhabdomyolysis.

Coagulation abnormalities in sepsis.

Although the pathophysiology of sepsis has been elucidated with the passage of time, sepsis may be regarded as an uncontrolled inflammatory and procoagulant response to infection. The hemostatic changes in sepsis range from subclinical activation of blood coagulation to acute disseminated intravascular coagulation (DIC). DIC is characterized by widespread microvascular thrombosis, which contributes to multiple organ dysfunction/failure, and subsequent consumption of platelets and coagulation factors, eventually causing bleeding manifestations. The diagnosis of DIC can be made using routinely available laboratory tests, scoring algorithms, and thromboelastography. In this cascade of events, the inhibition of coagulation activation and platelet function is conjectured as a useful tool for attenuating inflammatory response and improving outcomes in sepsis. A number of clinical trials of anticoagulants were performed, but none of them have been recognized as a standard therapy because recombinant activated protein C was withdrawn from the market owing to its insufficient efficacy in a randomized controlled trial. However, these subgroup analyses of activated protein C, antithrombin, and thrombomodulin trials show that overt coagulation activation is strongly associated with the best therapeutic effect of the inhibitor. In addition, antiplatelet drugs, including acetylsalicylic acid, P2Y12 inhibitors, and glycoprotein IIb/IIIa antagonists, may reduce organ failure and mortality in the experimental model of sepsis without a concomitant increased bleeding risk, which should be supported by solid clinical data. For a state-of-the-art treatment of sepsis, the efficacy of anticoagulant and antiplatelet agents needs to be proved in further large-scale prospective, interventional, randomized validation trials.

Effects of gabexate mesilate on coagulopathy and organ dysfunction in rats with endotoxemia: a potential use of thrombelastography in endotoxin-induced sepsis.

Sepsis and its associated multiple organ failure are related to high mortality in critical patients. Several studies have reported that gabexate mesilate, a synthetic inhibitor of trypsin-like serine protease, protects tissues/organs against injury in the models of endotoxemia. The aim of this study was to examine whether gabexate mesilate could attenuate coagulopathy and organ dysfunction in lipopolysaccharide (LPS)-induced sepsis model by using thrombelastography (TEG). LPS (7.5  mg/kg/h, intravenouly for 4  h) was administered to male adult Wistar rats. Some of the LPS rats received a continuous infusion of gabexate mesilate (10  mg/kg/h, intravenously for 8.5  h) for 30  min before the LPS administration. Variable parameters of hemodynamics, biochemistry, hemostasis and inflammatory response were measured for 6  h after the LPS infusion. TEG variables (R-time, K-time, α-angle, and maximal amplitude) were also measured. The pretreatment of LPS rats with gabexate mesilate significantly attenuated the lung, liver and kidney dysfunction, consumptive coagulopathy, the increases in serum tumor necrosis factor-α, interleukin-6, plasma thrombin-antithrombin complex and plasminogen activator inhibitor-1, and neutrophils infiltration score in lung, liver and kidney, compared with the LPS alone group. In addition, TEG parameters correlated with tissue and liver injury in the late phase of endotoxemia. In particular, a strong negative correlation between maximal amplitude at 4  h and Ln (lactate dehydrogenase) at 6  h after LPS infusion was noted (r = -0.752, P < 0.001, R = 0.566). These results indicate that beneficial effects of anticoagulants (e.g. gabexate mesilate) in endotoxemia could be monitored by TEG per se.

Low-dose dexamethasone ameliorates circulatory failure and renal dysfunction in conscious rats with endotoxemia.

Corticosteroids have long been proposed as a therapeutic adjuvant in septic renal dysfunction because of their anti-inflammatory properties and favorable results from animal experiments. However, some reports suggested the potential for harm associated with the administration of early high-dose corticosteroids in patients with severe sepsis and septic shock. Thus, we examined the effects of low-dose dexamethasone (0.01 and 0.1 mg/kg) on hemodynamics and renal function in conscious rats with endotoxemia. Intravenous injection of rats with endotoxin (E. coli lipopolysaccharide, LPS, 1 mg/kg) caused hypotension, vascular hyporeactivity, and tachycardia as well as renal dysfunction. Circulatory failure and renal dysfunction caused by LPS were significantly attenuated in the dexamethasone 0.1 mg/kg-treated group. The nitric oxide (NO) production in plasma and renal tissue and the iNOS protein expression in the kidney were suppressed by cotreatment of LPS rats with dexamethasone, 0.1 mg/kg. Light microscopy showed that 0.1 mg/kg dexamethasone reduced marked infiltration of neutrophils in renal tissues from LPS rats. Moreover, the survival rate at 18 h was significantly increased in the dexamethasone 0.1 mg/kg-treated group when compared with the LPS group. These results suggest that the beneficial effects of low-dose dexamethasone (0.1 mg/kg) in conscious rats with endotoxic shock are associated with amelioration of circulatory failure and renal dysfunction, and this is attributed to inhibition of NO production.

Honokiol protects rat brain from focal cerebral ischemia-reperfusion injury by inhibiting neutrophil infiltration and reactive oxygen species production.

We have previously shown that honokiol, an active component of Magnolia officinalis, displayed protective effect against focal cerebral ischemia-reperfusion (FCI/R) injury in rats. Production of reactive oxygen species (ROS) and infiltration of neutrophils to injured tissue play deleterious roles during cerebral ischemia. To study the mechanism(s) in mediating neuroprotective effect of honokiol, FCI/R-induced neutrophil infiltration and lipid peroxidation in brain tissue, and activation of neutrophils in-vitro were examined. Intravenous administration of honokiol (0.01-1.0 microg/kg) 15 min before (pretreatment) or 60 min after (post-treatment) middle cerebral artery occlusion reduced the total infarcted volume by 20-70% in dose-dependent manner. Pretreatment or post-treatment of honokiol at concentration of 0.1 and 1.0 microg/kg significantly decreased the neutrophil infiltration in the infarcted brain. Time course of neutrophil infiltration was performed in parallel with the lipid peroxidation in infracted brain tissue during FCI/R injury. The results indicate that honokiol can protect brain tissue against lipid peroxidation and neutrophil infiltration during FCI/R injury and cerebral infarction induced by FCI/R is accompanied with a prominent neutrophil infiltration to the infarcted area during FCI/R course. In-vitro, honokiol (0.1-10 microM) significantly diminished fMLP (N-formyl-methionyl-leucyl-phenylalanine)- or PMA (phorbol-12-myristate-13-acetate)-induced neutrophil firm adhesion, a prerequisite step behind neutrophil infiltration, and ROS production in neutrophils. Intracellular calcium overloading activates calcium-stimulated enzymes and further exaggerates FCI/R injury. Honokiol (0.1-10 microM) impeded the calcium influx induced by fMLP (a receptor agonist), AlF(4)(-) (a G-protein activator) or thapsigargin (an intracellular calcium pool releaser). Therefore, we conclude that the amelioration of FCI/R injury by honokiol can be attributed to its anti-oxidative and anti-inflammatory actions through, at least in part, limiting lipid peroxidation and reducing neutrophil activation/infiltration by interfering firm adhesion, ROS production, and calcium overloading that may be primed/activated during FCI/R injury.