Stress and the immune system: preliminary observations in rheumatoid arthritis using an in vivo marker of immune activity

This study measured volumetric liver blood flow and galactose clearance concurrently during orthotopic liver transplant in human subjects. Ultrasound transit time flowmeters measured hepatic artery and portal vein flow 1-3 h after reperfusion. Galactose (100 mg/min) was infused over 45-60 min to steady state for calculation of clearance. Mean +/- S.D. total volumetric flow was 1966 +/- 831 ml/min with portal flow contributing 86%. Mean galactose clearance was 1988 +/- 641 ml/min. There was a significant correlation (p < 0.05, r = 0.61) between volumetric total liver blood flow and galactose clearance. The data show that: (i) the newly transplanted liver is capable of metabolizing galactose within 1-3 h of reperfusion; and (ii) liver blood flow is high in the newly implanted liver. The clinical importance of this observation is that there is increased clearance of high first pass substances by the transplanted liver which may be of importance in patient management.     

Neutrophil and macrophage activation and anaphylatoxin formation in orthotopic liver transplantation without the use of veno-venous bypass

BACKGROUND: Activation of neutrophils and activation of complement may be an aetiologic factor behind circulatory insufficiency in association with reperfusion of the grafted liver. METHODS: Neutrophil and macrophage activation (determined as PMN elastase and neopterin release) and complement activation were evaluated in 15 consecutive patients undergoing orthotopic liver transplantation without the use of veno-venous bypass. RESULTS: The PMN elastase concentrations were increased at the end of the anhepatic phase, 2, 5 and 30 min after start of reperfusion and 6 and 24 h postoperatively. There were significantly higher PMN elastase concentrations in patients with circulatory instability (postreperfusion syndrome) compared with those without postreperfusion syndrome. The neopterin concentration was increased 2 min after the start of reperfusion and remained elevated until 6 h postoperatively. The plasma complement C3a concentrations were increased at the end of the anhepatic phase and 2, 5 and 30 min after the start of reperfusion. The plasma C3a levels were higher in patients with postreperfusion syndrome compared to those without. CONCLUSIONS: Activation of neutrophils and macrophages and of the complement cascade with the formation of biologically active substances may be one explanation for the circulatory instability often seen in patients undergoing orthotopic liver transplantation.     

Role of the donor liver in the origin of platelet disorders and hyperfibrinolysis in liver transplantation

We investigated the role of the donor liver in the origin of platelet disorders and hemostatic defects in liver transplantation. Eighteen pigs received an orthotopic or a heterotopic, auxiliary liver graft. Liver biopsies were taken for electron microscopic studies 5-10 min after reperfusion in nine animals. Blood samples were taken from the first hepatic outflow and from the systemic circulation before and 5 min after graft recirculation. Electron microscopy did not show any evidence of microthrombi or platelet aggregation in the graft, either after orthotopic liver transplantation or after heterotopic liver transplantation. Most blood platelets, which were lying free in the sinusoids, showed cell processes and many seemed to have lost their granulae, suggesting a degree of platelet activation. There were also signs of phagocytosis of platelets by the Kupffer cells. In the hepatic outflow, platelet count was significantly lower (p < 0.05) and fibrinolytic activity significantly higher (p < 0.01), than systemic post-reperfusion values. There were no important changes in the coagulation parameters. No significant changes were found between the effects on hemostasis of orthotopic and auxiliary graft reperfusion. In the second part of the study evidence for platelet activation was found after graft reperfusion in human liver transplantation. Plasma levels of platelet factor-4 and beta-thromboglobulin increased significantly after graft reperfusion. These studies suggest that platelet disorders and increased fibrinolytic activity are the major components of the hemostatic defect after graft recirculation in liver transplantation. Sequestration of platelets in the graft is probably due to the accumulation of (activated and degranulated) platelets in the sinusoids and phagocytosis by Kupffer cells.     

A comparison of the efficacy and tolerability of dorzolamide and acetazolamide as adjunctive therapy to timolol. Oral to Topical CAI Study Group

BACKGROUND: Recent observations provide evidence that complement is involved in the pathophysiology of ischemia/reperfusion injury. In this study, we assessed the impact of complement inhibition on hepatic microcirculation and graft function using a rat model of liver transplantation. METHODS: Arterialized orthotopic liver transplantation was performed in Lewis rats after cold preservation (University of Wisconsin solution, 4 degrees C, 24 h). Eight animals received the physiological complement regulator soluble complement receptor type 1 (sCR1) intravenously 1 min before reperfusion. Controls received Ringer's solution (n=8). Microvascular perfusion, leukocyte adhesion, and Kupffer cell phagocytic activity were studied 30-100 min after reperfusion by in vivo microscopy. RESULTS: Microvascular perfusion in hepatic sinusoids was improved in the sCR1 group (87+/-0.7% vs. 50+/-1%; P < 0.001). The number of adherent leukocytes was reduced in sinusoids (68.3+/-4.7 vs. 334.1+/-15.8 [adherent leukocytes per mm < or = liver surface]; P < 0.001) and in postsinusoidal venules after sCR1 treatment (306.6+/-21.8 vs. 931.6+/-55.9 [adherent leukocytes per mm < or = endothelial surface]; P < 0.001). Kupffer cell phagocytic activity was decreased in the sCR1 group compared to controls. Postischemic bile production reflecting hepatocellular function was increased by almost 200% (P = 0.004) after complement inhibition. Plasmatic liver enzyme activity was decreased significantly upon sCR1 treatment, indicating reduced parenchymal cell injury. CONCLUSIONS: Our results provide further evidence that the complement system plays a decisive role in hepatic ischemia/reperfusion injury. We conclude that complement inhibition by sCR1 represents an effective treatment to prevent reperfusion injury in liver transplantation.     

Electromagnetic fields influence NGF activity and levels following sciatic nerve transection

Activated neutrophils play an important role in reperfusion injury following hepatic ischemia. Neutrophil elastase is a powerful proteolytic enzyme. We investigated the possibility that ONO-5046. Na, which is a new recombinant-specific neutrophil elastase inhibitor, can reduce ischemia and reperfusion injury in the canine liver. Adult mongrel dogs (n = 19) were used in this experimental study. Seventy-five percent of the liver was resected after 60 min of vascular occlusion. The animals were divided into two groups. The ONO group (n = 8) was given ONO-5046. Na dissolved in saline starting 30 min prior to clamping the hepatic inflow and continuing for 4 h after reperfusion at a rate of 10 mg/kg/h. The nontreatment group (n = 11) received a saline solution for the same period. ALT and LDH levels were significantly lower (P < 0.05) in the ONO group than in the nontreatment group after reperfusion. Purine nucleoside phosphorylase and hyaluronic acid levels, which are markers of endothelial damage, were significantly lower (P < 0.05) in the ONO group than in the nontreatment group after reperfusion. Histologically, widely spread hepatocyte necrosis was found in dogs in the nontreatment group that died prematurely. Neutrophil infiltration of the sinusoids was less evident in the ONO group than in the nontreatment group. Neutrophil elastase inhibitor may prevent injuries of both endothelial and parenchymal cells in extended hepatectomy with vascular occlusion.     

Hepatic protein synthesis rate of liver specimens as a predictor of viability in rat cold ischemia liver transplantation model

BACKGROUND/AIMS: We have previously reported that the hepatic protein synthesis rate, calculated as the uptake rate of L-[4.5 3H] leucine by the protein fraction during a 10-min incubation of a 16-G needle biopsy specimen of liver tissue, represents a high level of liver function and is therefore useful for evaluating liver function. We investigated the hepatic protein synthesis rate level in a pre-transplant liver to learn if it might predict the outcome in a rat orthotopic liver transplantation model. METHODS: Grafts were stored, liver specimens were obtained using a 21-G Chiba type II skinny needle, and the hepatic protein synthesis rate was calculated. Subsequently, liver transplantation was performed, and the hepatic protein synthesis rate level of revascularized liver, tissue blood flow rate, serum alanine aminotransferase, lactate dehydrogenase, hyaluronic acid, ketone body rate, and 2-week survival were examined. RESULTS: The hepatic protein synthesis rate of pretransplant liver was correlated with parameters of post-transplant liver function: hepatic protein synthesis rate of the revascularized liver (r=0.92, p<0.0001), tissue blood flow rate (r=0.77, p<0.004), serum alanine aminotransferase (r=-0.69, p<0.003), lactate dehydrogenase (r=-0.54, p<0.03), hyaluronic acid (r=-0.86, p<0.0002), and ketone body rate (r=0.57, p<0.02). Pretransplant hepatic protein synthesis rate in survivors was 263.6+/-54.2 nmol/mg protein/10 min, while that in nonsurvivors was significantly lower at 162.0+/-39.0 (p<0.0001). When evaluation was made using a logistic regression model, the accuracy predicted using the value of hepatic protein synthesis rate was 95% (19/20). CONCLUSIONS: These results suggest that measuring the hepatic protein synthesis rate of the grafts with a 21-G Chiba type II skinny needle may be a predictive criterion in the assessment of graft viability.     

Short-term effects of transcatheter arterial chemoembolisation on metabolic activity of the liver of cirrhotic patients with hepatocellular carcinoma

BACKGROUND: Transcatheter arterial chemoembolisation, a procedure for the treatment of hepatocellular carcinoma, provokes a pronounced but transient increase in hepatic cytolysis parameters. A definite evaluation of the impairment of liver function after this treatment, performed by adequate techniques, is still lacking. AIMS: To assess and quantify the impairment of liver metabolic activity after arterial chemoembolisation in patients with cirrhosis. The variations of hepatic vein pressure gradient provoked by this procedure were evaluated. PATIENTS: 15 patients with cirrhosis (Child's class A and B) and hepatocellular carcinoma. METHODS: 17 transcatheter arterial chemoembolisations with epirubicin, iodised oil, and gelfoam were performed; liver function was assessed before, the following day, and after seven days measuring galactose elimination capacity; aminopyrine breath test was also performed in six patients before the procedure and seven days after. In 10 patients intrinsic hepatic clearance of indocyanine green and hepatic vein pressure gradient were measured by hepatic vein catheterisation before and 30 minutes after chemoembolisation. RESULTS: Intrinsic hepatic clearance of indocyanine green decreased significantly from (mean (SEM)) 355 (140) ml/min to 277 (98) ml/min after the procedure (p = 0.0007). Galactose elimination capacity did not show significant changes, being 4.00 (0.90) mg/min/kg body weight at baseline, 4.20 (0.90) mg/min/kg body weight after one day, and 3.95 (0.87) mg/min/kg body weight seven days after chemoembolisation. Aminopyrine breath test was 2.31 (1.09)% and remained unchanged after treatment, being 2.39 (2.04)% at day 7. Baseline hepatic vein pressure gradient was 17.0 (5.5) mm Hg, and 14.4 (3.7) mm Hg 30 minutes after chemoembolisation (p = 0.09). CONCLUSIONS: A single transcatheter chemoembolisation in cirrhotic patients was detected by galactose elimination capacity and aminopyrine breath test one and seven days after the procedure. Therefore it can be considered a safe therapeutic tool for hepatocellular carcinoma in Child's class A and B cirrhotic patients.     

Spinal reflexes in the determination of brain death

Tranexamic acid (TA) is a synthetic drug that inhibits fibrinolysis. It has been administered to decrease the use of blood products during cardiac surgery and orthotopic liver transplantation when infused in larger doses. A small-dose infusion of aprotinin causes a reduction in fibrinolysis and blood product requirement during orthotopic liver transplantation without apparent risk of intravascular thrombosis. This prospective study was designed to investigate whether a small-dose infusion of TA would be equally effective in reducing fibrinolysis and blood product transfusions during orthotopic liver transplantation. A double-blind, controlled study was undertaken to compare the efficacy of a small-dose TA infusion with that of a placebo. Thirty-two consecutive patients were randomized either to the TA group (n = 16), which received an intravenous infusion of 2 mg x kg(-1) x h(-1), or to the control group (n = 16), which received an identical volume of normal saline. Coagulation values were measured, a field rating was made by the surgeon, and a thromboelastogram was produced at four predetermined intervals throughout the case-before TA infusion was started, after portal vein ligation, 10 min after reperfusion, and at the end of surgery. Intraoperative transfusion requirements were recorded during the procedure and for the first 24 h postoperatively. A record was kept of any intraoperative epsilon-aminocaproic acid administered for uncontrolled fibrinolysis. The thromboelastogram clot lysis index was significant for lysis in the control group during both the anhepatic and the neohepatic phases (P < 0.01 and P < 0.05, respectively) when compared with the TA group. Fibrin degradation products were significantly increased (>20 microg/mL) in the control group at reperfusion (P < 0.03) and at the end of surgery (P < 0.01). D-dimers were also significantly increased (>1 mg/L) in the control group at the end of surgery (P < 0.04). Nine of the 16 control patients versus 3 of the 16 TA patients required epsilon-aminocaproic acid rescue for fibrinolysis. There were no other significant differences between groups. Transfusion requirements during surgery and for the first 24 h postoperatively did not differ significantly between the two groups. We conclude that the use of small-dose TA reduces fibrinolysis but not transfusion requirements during orthotopic liver transplantation.     

Effects of Carolina rinse and adenosine rinse on microvascular perfusion and intrahepatic leukocyte-endothelium interaction after liver transplantation in the rat

Flushing hepatic grafts immediately before revascularization with a specially designed rinse solution such as "Carolina rinse" has been reported to improve survival after liver transplantation in the rat. This study investigated the influence of Carolina rinse and adenosine rinse on early graft function, microcirculation, and leukocyte (WBC)-endothelial cell interaction of arterialized syngeneic orthotopic liver transplants in Lewis rats. Livers were preserved for 24 hr in University of Wisconsin solution and flushed immediately before reperfusion with either Ringer's lactate (group A: n = 7), Ringer's lactate + 0.2 mmol/liter adenosine (group B: n = 6), or Carolina rinse (group C: n = 7). Microvascular perfusion and WBC accumulation were assessed by intravital fluorescence microscopy. In group C, acinar perfusion was significantly improved, accompanied by a lower percentage of nonperfused sinusoids 1 hr after reperfusion (mean +/- SEM: 26 +/- 2% [group A], 21 +/- 2% [B], 11 +/- 1% [C], P < 0.01 for C vs. A or B). In addition, Carolina rinse and, to a lesser extent, adenosine rinse reduced the number of WBC sticking in sinusoids and postsinusoidal venules. Better graft function in group C was indicated by increased bile flow during the observation period of 90 min after reperfusion (0.5 +/- 0.3 ml/100 g liver [group A], 1.5 +/- 0.7 [B], 3.7 +/- 0.6 [C], P < 0.01 for C vs. A or B). We conclude that Carolina rinse is able to improve early excretory hepatocellular function, microvascular perfusion, and intrahepatic WBC accumulation after prolonged cold ischemia and reperfusion, but adenosine is unlikely to be the key component of this rinse solution.     

Impact of N-acetylcysteine on the hepatic microcirculation after orthotopic liver transplantation

Recent observations showed an improvement of hepatic macro- and microhemodynamics as well as survival rates after warm ischemia of the liver following treatment with N-acetylcysteine (NAC). In this study we assessed the influence of NAC on the hepatic microcirculation after orthotopic liver transplantation (OLT) using intravital fluorescence microscopy. OLT with simultaneous arterialization was performed in 16 male Lewis rats following cold storage in University of Wisconsin solution for 24 hr. Within the experimental group (n = 8) donors received NAC (400 mg/kg) 25 min before hepatectomy. In addition, high-dose treatment of recipients with NAC (400 mg/kg) was started with reperfusion. Control animals (n = 8) received an equivalent amount of Ringer's solution. Intravital fluorescence microscopy was performed 30-90 min after reperfusion assessing acinar and sinusoidal perfusion, leukocyte-endothelium interaction, and phagocytic activity. Treatment with NAC reduced the number of nonperfused sinusoid from 52.4 +/- 0.8% to 15.7 +/- 0.5% (p = 0.0001) (mean +/- SEM). Furthermore, we achieved a significant reduction of leukocytes adhering to sinusoidal endothelium (per mm2 liver surface) from 351.9 +/- 13.0 in controls to 83.6 +/- 4.2 in the experimental group (P = 0.0001). In postsinusoidal venules, treatment with NAC decreased the number of sticking leukocytes (per mm2 endothelium) from 1098.5 +/- 59.6 to 425.9 +/- 37.7 (P = 0.0001). Moreover, bile flow was significantly increased after therapy with NAC (4.3 +/- 1.2 vs. 2.2 +/- 0.7 ml/90 min x 100g liver) (P < 0.05). Phagocytic activity was not influenced by application of NAC. We conclude that high-dose therapy with NAC in OLT attenuates manifestations of microvascular perfusion failure early after reperfusion and should be considered as a means to reduce reperfusion injury.     

Blood concentrations of dopexamine in patients during and after orthotopic liver transplantation

Blood concentrations of dopexamine were measured in five female patients during and after orthotopic liver transplantation. Each patient received a continuous infusion of the drug (2 micrograms kg-1 min-1), starting at induction of anaesthesia and finishing 48 h afterwards. Blood concentrations of dopexamine increased rapidly at the start of the infusion, to a mean (range) value of 64 (40-150) ng ml-1 after 20 min. Blood concentrations of dopexamine increased further during the anhepatic period of surgery, to 236 (180-410) ng ml-1. On reperfusion of the donor liver, concentrations of dopexamine decreased rapidly, reaching similar values to the maximum seen during the dissection period. Steady-state was not reached during either the dissection or anhepatic periods. From 1-2 h after revascularisation the mean (range) steady-state concentration was 85 (69-102) ng ml-1 corresponding to a mean (range) clearance of 24 (20-29) ml min-1 kg -1. These results suggest that the liver plays a significant role in the clearance of dopexamine.     

Phospholipid signaling in leukocytes

The principal cause of primary non-function in orthotopic liver transplantation is thought to be preservation injury to the microvasculature. We, therefore, evaluated if effluent levels of hyaluronate, whose uptake is an endothelial cell marker, could predict early graft function and ultimate graft outcome in orthotopic liver transplantation. A total of 102 cases were studied in two phases. In the first phase, we attempted to determine if a correlation existed between effluent hyaluronate levels, early graft function and ultimate graft outcome. This phase of the study was also used to determine hypothetical cut-off values for hyaluronate which could discriminate between good and bad livers. Thirty-two livers orthotopically transplanted to randomly selected primary recipients were studied. After varying periods of static cold storage (4 degrees C) in University of Wisconsin solution, the livers were reinfused with cold (4 degrees C) lactated Ringer's solution. The first 50 ml of the reperfusion effluent was collected from the infrahepatic vena cava. Effluent samples were analyzed for hyaluronate. Linear regression analysis demonstrated a significant correlation between effluent hyaluronate levels and post-operative aspartate and alanine aminotransferase levels (p < 0.001 for both). Logistic regression demonstrated a highly significant correlation (p = 0.0056) between effluent hyaluronate levels and ultimate graft outcome. Generation of Receiver Characteristics Curves indicated that a level between 400 and 430 micrograms.l-1 could possibly discriminate between good livers and those at risk of early graft failure. The authenticity of this hyaluronate cut-off level was further confirmed in the second phase of the study where 70 consecutive primary crossmatch-negative transplants were performed. A highly significant difference was observed in peak aspartate and alanine aminotransferase levels in the first week (p < 0.0006 and p < 0.0005, respectively) between livers with effluent hyaluronate levels < or = 400 micrograms.l-1 and livers with hyaluronate levels higher than 400 micrograms.l-1. Logistic regression revealed a highly significant correlation between effluent hyaluronate levels and graft success (p = 0.0001). Since hyaluronate uptake by the microvascular endothelial cell is significantly greater than production, high hyaluronate effluent levels in failed livers would be due to decreased hyaluronate uptake by the injured microvascular endothelial cell. We therefore conclude that effluent hyaluronate levels may prove to be a reliable preoperative test to assess early graft function and outcome in clinical orthotopic liver transplantation.     

Hereditary neuralgic amyotrophy

Using a new ion-selective electrode, plasma concentration of ionized magnesium was measured in nine adult patients undergoing orthotopic liver transplantation. Baseline plasma ionized magnesium (IMg2+) concentration (0.49 +/- 0.07 mmol/L) was slightly below normal values (0.55-0.66 mmol/L, 95% CI): Six patients had ionized hypomagnesemia and two of these had total hypomagnesemia. Ionized IMg2+ concentration progressively decreased during the dissection (0.45 +/- 0.07 mmol/L, p < 0.05) and anhepatic stage (0.38 +/- 0.07 mmol/L, p < 0.05) and returned toward baseline values by 2 hours after graft reperfusion. Plasma ionized calcium levels and acid-base status were maintained within normal limits during surgery. Serum citrate concentration increased during the dissection (0.58 +/- 0.60 mmol/L) and anhepatic stages (1.18 +/- 0.78 mmol/L), the result of transfusion of citrate-rich blood products in the absence of adequate hepatic function, and gradually returned toward baseline values after graft reperfusion. IMg2+ concentration inversely correlated with the plasma citrate concentration (r2 = 0.54). The results of this study demonstrate that ionized hypomagnesemia invariably occurs during liver transplantation and suggest that this derangement may be a clinical concern, because magnesium is an important cofactor for the maintenance of cardiovascular homeostasis. The data further suggest the clinical importance of supplementation with magnesium based on the monitoring of plasma IMg2+ concentration.     

Evaluation of early graft function by hepatic venous hemoglobin oxygen saturation following orthotopic liver transplantation in the rat

This study was designed to evaluate the use of hepatic venous oxygen saturation (Shvo2) as a predictor of early graft function after liver transplantation. We examined the levels of Shvo2 and serum ALT after transplantation using the isogeneic rat orthotopic liver transplant model. Shvo2 levels 2 hr after reperfusion in the 6-hr cold ischemia (nonviable allograft) group were significantly lower than those in the 1-hr and 3-hr cold ischemia (viable allograft) groups. However, there was no significant difference in ALT levels among these groups. These results suggest that decreased hepatic blood flow due to microcirculatory disturbances may occur in the nonviable allograft even in the early phase of reperfusion and may be responsible for ischemic damage to parenchymal cells. Therefore, Shvo2 could provide a simple index of the initial graft status and be useful for a rapid etiological diagnosis of early postoperative graft dysfunction and for estimating the graft outcome after liver transplantation.     

ICAM-1 antisense oligodesoxynucleotides prevent reperfusion injury and enhance immediate graft function in renal transplantation

BACKGROUND: Ischemia-reperfusion injury after organ transplantation is a major cause of delayed graft function. We showed earlier that antisense oligodesoxynucleotides (ODN) for intercellular adhesion molecule-1 (ICAM-1) ameliorate reperfusion injury after acute ischemia. This study tested the hypothesis that antisense ODN for ICAM-1 prevents ischemia-reperfusion injury and facilitates immediate graft function in a rat autotransplantation model. METHODS: Both kidneys were removed from male Lewis rats and re-implanted the left kidney after 30 minutes of cold ischemia time. The warm ischemia time was 60 minutes. Sham operated, uninephrectomized animals served as controls for renal function and histology. ICAM-1 antisense ODN (5 mg/kg), reverse ODN, or saline-vehicle were administered to donor animals i.v. six hours before autotransplantation. Glomerular filtration rate (insulin clearance), and serum creatinine concentrations were measured 24 hours post-transplantation. Tubular necrosis severity was assessed by histological grading scale. ICAM-1 expression was determined by immunohistochemistry and Western blot. RESULTS: Antisense ODN decreased ICAM-1 expression and leukocyte infiltration significant. Antisense ODN-treated animals showed significantly less tubular necrosis, than controls. Serum creatinine of antisense ODN-treated animals (N = 6) was 0.55 +/- 0.02 mg/dl compared to 1.92 +/- 0.07 mg/dl in reverse ODN-treated controls (N = 6; P < 0.01), 24 hours after transplantation. Antisense ODN-treated animals had normal GFR (0.93 +/- 0.07 ml/min/kidney wt) compared to sham-operated animals (0.95 +/- 0.09 ml/min/kidney wt), while autotransplanted animals treated with reverse ODN or saline-vehicle were all anuric. The ischemia-reperfusion-induced up-regulation of MHC class II was totally prevented by antisense ODN. CONCLUSIONS: ICAM-1 inhibition ameliorates ischemia-reperfusion injury and prevents delayed graft function. Antisense ODN-treatment of donors or donor organs for ICAM-1 may be useful for the prevention of reperfusion injury in human renal transplantation.     

Ischemia/reperfusion injury in human kidney transplantation: an immunohistochemical analysis of changes after reperfusion

Organs used for transplantation undergo varying degrees of cold ischemia and reperfusion injury after transplantation. In renal transplantation, prolonged cold ischemia is strongly associated with delayed graft function, an event that contributes to inferior graft survival. At present, the pathophysiological changes associated with ischemia/reperfusion injury in clinical renal transplantation are poorly understood. We have performed an immunohistochemical analysis of pre- and postreperfusion biopsies obtained from cadaver (n = 55) and living/related donor (LRD) (n = 11) renal allografts using antibodies to adhesion molecules and leukocyte markers to investigate the intragraft changes after cold preservation and reperfusion. Neutrophil infiltration and P-selectin expression were detected after reperfusion in 29 of 55 (53%) and 24 of 55 (44%) cadaver renal allografts, respectively. In marked contrast, neutrophil infiltration was not observed in LRD allografts, and only 1 of 11 (9%) had an increased level of P-selectin after reperfusion. Immunofluorescent double-staining demonstrated that P-selectin expression resulted from platelet deposition and not from endothelial activation. No statistically significant association was observed between neutrophil infiltration and P-selectin expression in the glomeruli or intertubular capillaries despite the large number of cadaver renal allografts with postreperfusion changes. Neutrophil infiltration into the glomeruli was significantly associated with long cold ischemia times and delayed graft function. Elevated serum creatinine levels at 3 and 6 months after transplantation were also associated with the presence of neutrophils and platelets after reperfusion. Our results suggest that graft function may be influenced by early inflammatory events after reperfusion, which can be targeted for future therapeutic intervention.     

Arthroscopic repair of full-thickness tears of the rotator cuff

OBJECTIVE: The authors' goal was to determine the effects of specific binding and blockade of P- and E-selectins by a soluble P-selectin glycoprotein ligand-1 (PSGL-1) in rat models of hepatic in vivo warm ischemia and ex vivo cold ischemia. The authors also sought to determine the effect of selectin blockade on isograft survival in a syngeneic rat orthotopic liver transplant model. SUMMARY BACKGROUND DATA: Ischemia/reperfusion (I/R) injury is a major factor in poor graft function after liver transplantation, which may profoundly influence early graft function and late changes. It is hypothesized that I/R injury leads to the upregulation of P-selectin, which is then rapidly translocated to endothelial cell surfaces within 5 minutes of reperfusion of the liver, initiating steps leading to tethering of polymorphonuclear neutrophil leukocytes to the vascular intima. Local production by leukocytes of interleukin-1, tumor necrosis factor-alpha, or both induces P-selectin expression on the endothelium and continues the cascade of events, which increases cell adherence and infiltration of the organ. METHODS: To examine directly the effects of selectins in a warm hepatic I/R injury model, 100 microg of PSGL-1 or saline was given through the portal vein at the time of total hepatic inflow occlusion. The effects of PSGL-1 in cold ischemia were assessed using an isolated perfused rat liver after 6 hours of 4 degrees C storage in University of Wisconsin (UW) solution, with or without the instillation of PSGL-1 before the storage. To evaluate the effect of selectin blockade on liver transplant survival, syngeneic orthotopic liver transplants were performed between inbred male Sprague-Dawley rats after 24 hours of cold ischemic storage in UW solution. A separate group of animals received two doses of 100 microg of PSGL-1 through the portal vein before storage and before reperfusion of the transplanted liver. Recipient survival was assessed at 7 days, and the Kaplan-Meier product limit estimate method was used for univariate calculations of time-dependent recipient survival events. RESULTS: In an in vivo warm rat liver ischemia model, perfusion with PSGL-1 afforded considerable protection from I/R injury, as demonstrated by decreased transaminase release, reduced histologic hepatocyte damage, and suppressed neutrophil infiltration, versus controls (p < 0.05). When cold stored livers were reperfused, PSGL-1 reduced the degree of hepatocyte transaminase release, reduced neutrophil infiltration, and decreased histologic hepatocyte damage (p < 0.05 vs. UW-only controls). On reperfusion, livers treated with PSGL-1 demonstrated increased portal vein blood flow and bile production (p < 0.05 vs. UW-only controls). In addition, 90% of the rats receiving liver isografts stored in UW solution supplemented with PSGL-1 survived 7 days versus 50% of those whose transplanted syngeneic livers had been stored in UW alone (p < 0.05). CONCLUSIONS: Selectins play an important role in I/R injury of the liver. Early modulation of the interaction between P-selectin and its ligand decreases hepatocyte injury, neutrophil adhesion, and subsequent migration in both warm and cold rat liver ischemia models. In addition, the use of PSGL-1 before ischemic storage and before transplantation prevents hepatic injury, as documented by a significant increase in liver isograft survival. These findings have important clinical ramifications: early inhibition of alloantigen-independent mechanisms during the I/R damage may influence both short- and long-term survival of liver allografts.     

Transfusion requirements are correlated with the degree of proteolysis of von Willebrand factor during orthotopic liver transplantation

During orthotopic liver transplantation (OLT) excessive bleeding is the main cause of death and graft failure. The acute bleeding tendency that accompanies OLT, particularly during the anhepatic period and after reperfusion of the graft, is due to the depletion or functional abnormalities of several hemostasis components caused by the enhanced activity of enzymes such as plasmin, trypsin and leukocyte proteases. We surmised that enhanced proteolysis might also cause abnormalities of von Willebrand factor (vWF), and that these abnormalities are implicated in the bleeding tendency that develops during OLT. Therefore, the pattern of vWF proteolysis was studied with 16 patients with chronic liver disease, in serial blood samples obtained before OLT, during the anhepatic stage, after graft reperfusion and at the end of the surgical procedure. vWF became markedly degraded during the anhepatic and reperfusion stages, as shown by the partial loss of high molecular weight multimers, the relative decrease of the intact 225 kD subunit and the increase of the native proteolytic fragments of 176 and 140 kD. Novel proteolytic fragments also became detectable. Using monoclonal antibody epitope mapping, it could be demonstrated that some of the proteolytic fragments corresponded in apparent molecular mass to those produced in vitro by incubating purified vWF with plasmin or elastase, but other fragments could not be attributed to these proteases. During the anhepatic and reperfusion stages there was a significant correlation between the degree of vWF degradation and the total amount of blood components transfused to replace blood losses. To evaluate whether or not vWF degradation could be controlled by the administration of a broad-spectrum protease inhibitor such as aprotinin, 5 patients were given a bolus dose of 500,000 U before surgery followed by 100,000 U/h during surgery, 5 were given a 2,000,000 U bolus followed by 500,000 U/h, and no aprotinin was given to the remaining 6 patients. There were no differences in the patterns or degrees of vWF degradation between patients treated with aprotinin or not. In conclusion, there is a marked degradation of a key hemostasis protein during OLT. These alterations may be of clinical significance, because they are correlated with the transfusion requirements.