Inhibition of intraluminal pancreatic enzymes with nafamostat mesilate improves clinical outcomes after hemorrhagic shock in swine.

BACKGROUND: Recent studies suggest that intraluminal pancreatic enzymes play a major role in the initiation of the inflammatory cascade by the gut after hemorrhagic shock. Previous animal models have shown that the inhibition of enteral pancreatic enzymes with a serine protease inhibitor, nafamostat mesilate (NM), decreases leukocyte activation and transfusion requirements after hemorrhagic shock. The objective of this study was to determine whether enteroclysis with NM would improve the clinical outcomes in swine after hemorrhagic shock and intestinal hypoperfusion. METHODS: Thirty-three male Yucatan minipigs weighing 25 kg to 30 kg underwent a controlled hemorrhage of 25 mL/kg with mesenteric clamp for further gut ischemia. Animals were allocated to three groups: (1) shock only (n = 15), (2) shock + enteroclysis with 100 mL/kg GoLYTELY (GL) as a carrier (n = 11), and (3) shock + enteroclysis with GL + 0.37 mmol/L NM (GL+NM, n = 7). Animals were resuscitated, recovered from anesthesia, observed for 3 days, and graded on a daily 4-point clinical scoring system. A score of 0 indicated a moribund state or early death, and a score of 4 indicated normal behavior. RESULTS: Pigs treated with GL + NM had significantly higher mean postoperative recovery scores (3.8 +/- 0.4, essentially normal behavior with no early deaths) compared with animals within the shock only and shock + GL groups (2.1 +/- 1 with one early death and 2.2 +/- 1.2 with two early deaths, respectively, analysis of variance p < 0.003). CONCLUSION: The inhibition of intraluminal pancreatic enzymes using enteroclysis with the serine protease inhibitor, NM, after hemorrhagic shock significantly improves the clinical outcome.

Inhibition of enteral enzymes by enteroclysis with nafamostat mesilate reduces neutrophil activation and transfusion requirements after hemorrhagic shock.

BACKGROUND: The gut origin of the inflammatory response in trauma patients has been difficult to define. "In vivo" generation of neutrophil-activating factors by gut proteases may be a cause of multiorgan failure after hemorrhagic shock, and can be prevented with the serine protease inhibitor nafamostat mesilate (Futhan). The objective of this study was to determine the effect of nafamostat mesilate given by enteroclysis on enteric serine protease activity, neutrophil activation, and transfusion requirements during hemorrhagic shock. METHODS: Sixteen pigs weighing 21 to 26 kg were divided into control and treatment groups. A laparotomy was performed under anesthesia, and catheters were placed in the duodenum, midjejunum, and terminal ileum. Pigs were bled 30 mL/kg over 30 minutes and maintained at a mean arterial pressure of 30 mm Hg for 60 minutes. Shed blood was then used to maintain a mean arterial pressure of 45 mm Hg for another 3 hours. Treated animals received 100 mL/kg of 0.37 mmol/L nafamostat mesilate in GoLYTELY through the duodenal catheter at 1 L/h. Control animals received GoLYTELY only. Samples of enteral content and blood were taken at baseline, after shock, and at 30-minute intervals during resuscitation. Animals were killed after 3 hours of resuscitation. Enteral trypsin-like activity at the three gut sites was measured by spectrophotometry. Activation of naive human neutrophils by pig plasma was measured by the percentage of cells having pseudopods larger than 1 microm on microscopy. Lung, liver, and small bowel were analyzed by histology and myeloperoxidase assay. RESULTS: Both control and nafamostat mesilate-treated groups had significant reductions in protein and protease levels in the duodenum during enteroclysis; however, only nafamostat mesilate-treated animals had persistent suppression of protease activity throughout the experiment. Nafamostat mesilate-treated animals had a lower transfusion requirement of shed blood, 18.1 +/- 4.5 mL/kg versus 30 +/- 0.43 mL/kg (p = 0.002). Nafamostat mesilate-treated animals had significantly less neutrophil activation than controls at 150 minutes after resuscitation (33.7 +/- 6.48% vs. 42.4 +/- 4.57%,p = 0.01) and 180 minutes after resuscitation (31.1 +/- 3.31% vs. 46.9 +/- 4.53%, p = 0.0002). Lung myeloperoxidase activity was lower in nafamostat mesilate-treated animals (0.31 +/- 0.14) than in control animals (0.16 +/- 0.04, p = 0.04). Histology of liver and small intestine showed less injury in nafamostat mesilate-treated animals. CONCLUSION: Nafamostat mesilate given by means of enteroclysis with GoLYTELY significantly reduces enteral protease levels, leukocyte activation, and transfusion requirements during resuscitation from hemorrhagic shock. This strategy may have clinical promise.

The modulation of hepatic injury and heat shock expression by inhibition of inducible nitric oxide synthase after hemorrhagic shock.

The role of nitric oxide (NO) in maintaining homeostasis and regulating organ function during hemorrhagic shock is complex. The inducible NO synthase (iNOS) has been hypothesized to play a critical role in the pathophysiologic consequences of severe hemorrhage. Heat shock protein (HSP) expression is increased by hemorrhage and is a marker of the magnitude of ischemic injury in the liver. HSP induction is protective against injury in animal models of inflammation and is regulated by NO in hepatocytes. To clarify the role of iNOS in hepatic injury and its relationship to HSP expression in hemorrhagic shock, NOS was inhibited with L-N-6-(1-iminoethyl) lysine (L-NIL), which is reported to be a selective inhibitor of the inducible NOS isoform. Doses of 50 microg/kg or 150 microg/kg were infused over 1 h at the end of compensated shock. Plasma ornithine carbamoyltransferase (OCT), a specific marker of liver injury, was significantly reduced after hemorrhage with low-dose L-NIL (7.1+/-1.5 IU/L) compared to saline-treated control rats (13.0+/-1.5 IU/L, P < 0.005), while high-dose L-NIL significantly increased OCT release (35.9+/-7.2 IU/L, P< 0.05 versus shock alone) despite a greater MAP after resuscitation. HSP expression (HSP-72 and HSP-32) after hemorrhage was increased by L-NIL treatment at the highest dose. We conclude that excessive NO production from iNOS contributes to shock-induced hepatic injury. Our data suggest HSP expression may reflect the degree of ischemic injury after hemorrhage.

Induction of heme-oxygenase-1 prevents the systemic responses to hemorrhagic shock.

Oxidant-mediated reperfusion injury of the gut is a major contributor of the systemic inflammatory response in hemorrhagic shock. Recent studies have suggested that heme-oxygenase-1 (HO-1) represents an endogenous protective mechanism against oxidant stress. We assessed whether HO-1 induction modulates the synthesis of tumor necrosis factor-alpha (TNF-alpha) in hemorrhagic shock. In rats submitted to hemorrhagic shock, pretreatment with hemoglobin (Hb) increased HO-1 mRNA expression in macrophages. This increased expression was associated with a decreased expression of TNF-alpha mRNA, as well as decreased plasma concentrations of TNF-alpha. These effects of Hb were reduced by the HO-1 inhibitor tin-protoporphyrin (Sn-PP 20 micromol/kg), while Sn-PP had no effect in the absence of Hb. In parallel, Hb pretreatment reduced pulmonary edema, vascular injury, and increased mesenteric blood flow, and these effects were reduced by Sn-PP. Thus, induction of HO-1 is protective in hemorrhagic shock, possibly through its antioxidant properties. Interventions that induce HO-1 may be beneficial in the treatment of shock states, leading to a reduced systemic inflammatory response.

BN 52021, PAF-receptor antagonist, improves diminished antioxidant defense system of lungs in experimentally induced haemorrhagic shock.

The purpose of our experiments was to determine whether BN 52021, PAF-receptor antagonist, has any influence on the production of reactive oxygen species in the lung induced by haemorrhagic shock. The results showed, that the activities of Cu,Zn-superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in the lung after haemorrhagic shock were decreased in comparison with control and sham groups while the activity of glutathione reductase (GSSG-R) was unchanged. The content of SH-groups was decreased while thiobarbituric acid reacting substances (TBA-rs) after haemorrhagic shock were increased. Infusion of BN 52021 (5.0 mg/kg i.v.) in haemorrhagic animals caused significantly increased activities of Cu,Zn-SOD, GSH-Px, GSSG-R in the lung. The SH-group level was slightly increased while TBA-rs returned to normal values.

[Research on energy metabolism enzyme in liver of the rabbit during hemorrhagic shock and effect of acupuncture at "renzhong" acupoint by quantitative histochemistry].

The energy crisis has been considered to act an important role during shock recent years. We all know the liver can supply for energy substrate when body is in an emergency condition. According to the theory of Traditional Chinese Medicine, Renzhong acupoint is a very important acupoint to treat "Jue Zheng" (including shock). We divided 36 male rabbits into three groups randomly, they were normal, acupuncture and nonacupuncture. The normal control was anesthetized only with sodium pentobarbital (35mg/kg). The acupuncture and nonacupuncture groups were operated under conscious with method of improving Wiggers(1950). After bleeding 20ml/kg, these animals were in shock. Then the acupuncture group was applied with acupuncture at "Renzhong" and the nonacupuncture group was not during hemorrhagic shock. The samples of liver were excised and stained using enzyme histochemical method. All of them were determined with MPV-2 and Univar-SMSP. Two dehydrogenases (LDH and SDH) in hepatocytes were observed in this work. The results showed that the enzyme's activity in nonacupuncture both LDH and SDH were the weakest among three groups (P less than 0.01 and P less than 0.05). But in acupuncture group, they were stronger than nonacupuncture group and near normal control. These suggest that acupuncture at "Renzhong" acupoint can increase activity of LDH and SDH during hemorrhagic shock. It is useful to resistance shock in a certain degree.

[The effect of severity and duration of hemorrhagic hypotension and the narcosis on the plasma levels of lysosomal enzymes in dogs (author's transl)]. / Die Wirkung von Grad und Zeitdauer der hämorrhagischen Hypotension und der Narkose auf die lysosomalen Plasmaenzymakivitäten bei Hunden

The effect of degree and duration of hemorrhagic hypotension on the lysosomal enzyme and LAP levels of plasma of the anaesthesied or wakeful dog has been examined. We ascertained that in a state of hypovolaemia a reduction by 10 mm Hg of the blood pressure resulting significantly increased enzyme activity during the same time. A similar change was caused by prolonging the time. In the anaesthesised group there was the enzyme activity after 120 min of haemorrhagic hypotension lower.