Preservation-reperfusion Injury Research Articles

Correlation of Histopathologic Findings of Non-Graft Threatening Preservation/Reperfusion Injury in Time-Zero Liver Needle Biopsies With Short-Term Post-transplantation Laboratory Alterations.

Background:Early post-transplantation alterations in liver tests are caused by a variety of etiologies including rejection, biliary or vascular complications, and preservation/reperfusion injury (PRI).Objectives:The aim of this study was to show the correlation between histopathologic changes of PRI and the alterations in liver tests in the early post-transplantation period.Materials and Methods:Between April 2013 and August 2014, histopathologic findings of protocol, time-zero, Tru-Cut, liver needle biopsies were evaluated in 94 cases of cadaveric liver transplantation. The histopathologic changes included ballooning degeneration, micro- and macro-vesicular steatosis, bilirubinostasis, apoptotic cells, bile plugs and neutrophilic infiltration. These histopathologic changes were compared with the early (15 days) post-transplantation liver laboratory findings.Results:Clinico-pathologic evaluation of all 94 cases was done by assessment of PRI findings in time-zero biopsies and possible causes of allograft injury were appraised. In 21 patients, a specific cause for allograft injury was found including rejection and/or surgical complications. In the remaining 73 cases, there was no specific cause for allograft injury and histopathologic findings of time-zero liver needle biopsies supported PRI. We classified liver laboratory tests alterations as: hepatocellular damage (elevation of transaminases and lactate dehydrogenase), cholestatic damage (elevation of alkaline phosphatase and total bilirubin) and mixed. Hepatocellular and cholestatic alterations in liver function tests were associated with the presence of marked apoptotic bodies and neutrophilic aggregates in time zero biopsies, respectively. On the other hand, macrovesicular steatosis was dominantly associated with mixed (hepatocellular and cholestatic) laboratory alterations of liver tests.Conclusions:Any discrepancy between histopathologic changes in time-zero biopsies and pattern of early liver laboratory alterations may be considered as a warning for causes other than PRI.

Histological and Clinicopathological Evaluation of Liver Allograft Biopsy: An Initial Experience of Fifty Six Biopsies.

Liver biopsy is gold standard for diagnosis of allograft dysfunction. The aim of study was to evaluate liver allograft biopsies performed for graft dysfunction, study the pattern of injury and intensity, and timeline of occurrence of graft dysfunction. Retrospective study was carried out of 56 liver allograft biopsies and their histological findings with clinical presentation were correlated. Totally 56 needle liver allograft biopsies from January 1210 to July 2014, obtained from 35 patients were studied for histological and clinicopathological evaluation. The mean age was 53.2±5.48 years. The most common original disease was alcoholic cirrhosis. The most common histological lesion was acute cellular rejection (ACR) in 31 (55.36%) biopsies followed by preservation-reperfusion injury (PRI) in 10 (17.86%) biopsies and drug toxicity in 8 (14.29%) biopsies. Chronic rejection was reported in 2 (3.57%) and recurrence of HCV in 3 (5.36%). Ischemic coagulative necrosis and acute cholangitis were seen in 1 (1.79 %) case each. Alcoholic cirrhosis was the most common etiology for end stage liver disease. ACR and PRI were the major complications in liver allograft biopsies at our centre.

Influence of preformed donor-specific antibodies and C4d on early liver allograft function

Introduction. The impact of preformed donor-specific antibodies (DSA) is incompletely understood in liver transplantation. The incidence and impact of preformed DSA on early post liver transplant were assessed and these were correlated with compliment fragment C4d on allograft biopsy. Methods. Pretransplant serum from 41 consecutive liver transplant recipients (brain dead donors; DBD = 27 and cardiac death donors; DCD = 14) were tested for class-specific anti-human leukocyte antigen (HLA) and compared against donor HLA types. Liver biopsies were taken during cold storage (t-1) and post-reperfusion (t0) stained with C4d and graded for preservation-reperfusion injury (PRI). Results. Of the 41 recipients, 8 (20%) had anti-HLA class I/II antibodies pretransplant, 3 (7%) were confirmed preformed DSA; classes I and II (n=1) and class I only (n=2). No biopsies showed definite evidence of antibody-mediated rejection. Graft biopsies in overall showed only mild PRI with ischemic hepatocyte C4d pattern similar in both positive and negative DSA patients. One DSA-positive (33%) compared with four DSA-negative patients (10%) had significant early graft dysfunction; severe PRI causing graft loss from primary nonfunction was seen only in DSA-negative group. Allograft biopsy of preformed DSA-positive patient demonstrated only minimal PRI; however, no identifiable cause could be attributed to graft dysfunction other than preformed DSA. Conclusion. Preformed DSA are present in 5–10% liver transplant recipients. There is no association between anti-HLA DSA and PRI and C4d, but preformed DSA may cause early morbidity. Larger studies on the impact of DSA with optimization of C4d techniques are required.

Gut Bacteria Drive Kupffer Cell Expansion via MAMP-Mediated ICAM-1 Induction on Sinusoidal Endothelium and Influence Preservation-Reperfusion Injury after Orthotopic Liver Transplantation

Bacteria in the gut microbiome shed microbial-associated molecule patterns (MAMPs) into the portal venous circulation, where they augment various aspects of systemic immunity via low-level stimulation. Because the liver is immediately downstream of the intestines, we proposed that gut-derived MAMPs shape liver immunity and affect Kupffer cell (KC) phenotype. Germ-free (GF), antibiotic-treated (AVMN), and conventional (CL) mice were used to study KC development, function, and response to the significant stress of cold storage, reperfusion, and orthotopic transplantation. We found that a cocktail of physiologically active MAMPs translocate into the portal circulation, with flagellin (Toll-like receptor 5 ligand) being the most plentiful and capable of promoting hepatic monocyte influx in GF mice. In MAMP-deficient GF or AVMN livers, KCs are lower in numbers, have higher phagocytic activity, and have lower major histocompatibility complex II expression. MAMP-containing CL livers harbor significantly increased KC numbers via induction of intercellular adhesion molecule 1 on liver sinusoidal endothelium. These CL KCs have a primed yet expected phenotype, with increased major histocompatibility complex class II and lower phagocytic activity that increases susceptibility to liver preservation/reperfusion injury after orthotopic transplantation. The KC number, functional activity, and maturational status are directly related to the concentration of gut-derived MAMPs and can be significantly reduced by broad-spectrum antibiotics, thereby affecting susceptibility to injury.

Erythropoietin as Additive of HTK Preservation Solution in Cold Ischemia/Reperfusion Injury of Steatotic Livers

Organ shortage in liver transplantation has justified usage of marginal donor livers to expand the donor organ pool. The particular susceptibility of steatotic livers to I/R injury necessitates optimal preservation conditions in order to minimize preservation-reperfusion injury for successful transplantation. The effect of erythropoietin (EPO) as additive to HTK preservation solution was studied in a mouse model. Lean and steatotic livers were harvested, stored for 24 h in 4°C HTK solution containing either EPO or saline and reperfused for 2 h with 37°C Krebs-Henseleit buffer. Livers without cold storage served as sham controls. Flushing of livers upon cold storage revealed a transaminase release, which was 2- to 10-fold higher in steatotic versus lean livers. EPO was effective in reducing the enzyme release to 50% in steatotic but not in lean livers. EPO prevented cold storage-induced denudation of the endothelial lining in steatotic livers, but aggravated it in lean livers. During reperfusion, steatotic livers presented with lower oxygen consumption and higher enzyme release than lean livers. In all livers, parameters of reperfusion injury remained unaffected by EPO. Expression of UCP2 was found markedly higher in steatotic livers. After I/R, steatotic livers revealed a significant drop of UCP2, whereas expression in lean livers was only slightly affected. EPO diminished Erk phosphorylation to almost the same extent in both mouse strains. Fortification of the preservation solution by EPO ameliorates cold ischemic injury of steatotic livers and may thus be considered for use as an adjunctive agent to increase the success of transplanting steatotic livers.

Cytoprotective Actions of FTY720 Modulate Severe Preservation Reperfusion Injury in RatRenal Transplants

Fingolimod (FTY720) is a potent agonist of sphingosine 1 phosphate receptors and thereby interferes with lymphocyte trafficking. We previously showed that FTY720 protects from mild preservation reperfusion injury induced by 4 hr of cold ischemia. The purpose of this study was to explore the role of FTY720 in ischemic injury and regeneration using a clinically relevant rat renal transplant model with 24 hr of cold ischemia. Donor kidneys were cold stored in the University of Wisconsin solution for 24 hr before transplantation into bilaterally nephrectomized syngeneic recipients (n=6 per group), which received 0.5 mg/kg/d FTY720 or vehicle through oral gavage. Grafts were harvested 2 or 7 days posttransplantation. Renal tissue was examined histologically, stained for apoptosis, proliferation, inflammatory cell infiltrates, and studied for transforming growth factor-beta, and tumor necrosis factor-alpha expression. Rat proximal tubular cells were incubated with 0.1 to 30 micromol/L of phosphorylated FTY720 to test for in vitro cytopathic effects. FTY720 induced peripheral lymphopenia and significantly reduced intragraft CD3 and ED1 infiltrates. Acute tubular damage scores and graft function were not influenced by FTY720. Tubular apoptosis was significantly reduced, whereas the number of proliferating cell nuclear antigen-positive tubular cells were markedly increased. FTY720 attenuated renal tumor necrosis factor-alpha and transforming growth factor-beta expression. In vitro, pharmacologic concentrations up to 1 micromol/L of phosphorylated FTY720 did not affect tubular cell viability. FTY720 confers tubular epithelial protection in the presence of severe preservation reperfusion injury. Beneficial effects may in part be due to reduction in cell-mediated immune mechanisms. Furthermore, FTY720 could be helpful in patients with delayed graft function.

Role of Postreperfusion Subcapsular Wedge Biopsies in Predicting Initially Poor Graft Function After Liver Transplantation

Preservation injury is a major contributing factor to primary allograft failure or poor initial graft function after an orthotopic liver transplant (OLT). We examined the histopathological findings from postreperfusion wedge biopsy specimens in relation to early graft function during the first postoperative week among OLT patients at our center. We reanalyzed subcapsular postreperfusion biopsy specimens from 88 patients to histologically grade the lesions. Grafts were grouped as good function, initial poor function (an alanine aminotransferase or aspartate aminotransferase level >1500 IU/L during week 1), or primary nonfunction (death or retransplantation). Only 1 patient experienced primary nonfunction; the remaining patients fell into the other 2 groups: ie, good function or initial poor function. When patients were compared using numerous morphologic and clinical features, no statistical relation was observed regarding clinical data on bile duct complications, donor type, graft volume, patient age, or type of stent. Histological features of neutrophilic infiltration of the subcapsular region, hepatocellular ballooning, and macro/microvesicular steatosis were not related to initial poor graft function; in contrast, there were prominent sinusoidal neutrophilic infiltrations and hepatocellular necrosis. Preservation-reperfusion injury (grade 2 or grade 3 neutrophilic infiltration) occurred in 78.6% of initial poor function patients and in 39.7% of good function patients. Subcapsular neutrophilic infiltration, a sign of surgical hepatitis, did not provide prognostic information about graft survival. Similar to other studies, we observed neutrophilic infiltration and necrosis away from the capsule to predict subsequent graft function.

Hemoglobin-Glutamer 200 Reduces Reperfusion Injury of the Cold Preserved Rat Liver by Induction of Heme Oxygenase-1

Microcirculatory failure after cold liver preservation and reperfusion impairs tissue oxygenation and causes additional organ damage. Hemoglobin-glutamer (HbG) 200 is a hemoglobin-based oxygen carrying solution capable to improve organ oxygenation. The aim of this study was to evaluate its potential to decrease reperfusion injury after cold liver preservation. Therefore, Wistar rat livers were stored at 4 degrees C for 24 h and reperfused in the isolated perfused rat liver model with a sanguineous perfusate for 180 min. The perfusate consisted of rat blood and Krebs-Henseleit solution (Group A), supplemented by either HES 6% (Group B), or HbG (Groups C and D). In Group D heme oxygenase (HO) activity was blocked by intraperitoneal tin protoporphyrin-IX application before organ harvest. HbG supplementation increased the perfusate hemoglobin by 3,3 g/dL. After 180 min reperfusion perfusate alanine aminotransferase levels (72 +/- 27 micro/L) were significantly reduced in Group C, compared with Groups A and B (140 +/- 28 micro/L and 203 +/- 62 micro/L, respectively). These results correlated with a significant increase of HO-1 expression and activity during reperfusion. These effects could be abolished by tin protoporphyrin-IX application. HbG has been proven to be effective to reduce cold liver preservation-reperfusion injury. The positive effect on reperfusion injury depends on the induction of HO-1, which increases the bilirubin production, an important antioxidant acting as intracellular radical scavenger.

Glutamine Donor Pretreatment in Rat Kidney Transplants with Severe Preservation Reperfusion Injury

Glutamine (GLN) has been shown to confer cytoprotection by enhancing endogenous heat shock protein (HSP) expression. We hypothesized that GLN donor pretreatment protects rat renal grafts against severe preservation reperfusion injury (PRI). GLN (0.75 g/kg) or saline was administered i.p. to male donor rats 24 h and 6 h before donor nephrectomy. Kidneys (n = 6/group) were cold-stored in UW solution for 40 h and transplanted into bilaterally nephrectomized syngeneic recipients. Grafts were removed after 24 h. Renal HSP 70 expression was determined by Western blotting. Graft function was assessed by serum creatinine. Renal cross sections were microscopically examined for acute tubular necrosis, apoptosis, tubular proliferation, and macrophage infiltration. GLN donor pretreatment significantly increased intragraft HSP 70 expression. Serum creatinine was not different between groups: 2.6 +/- 0.2 mg/dL (saline) versus 2.7 +/- 0.5 mg/dL (GLN). Both treatment groups showed severe tubular damage with significantly less papillary necrosis in the GLN group (P < 0.05). GLN significantly reduced the number of apoptotic tubular cells in the cortex, medulla, and papilla (P < 0.001 versus saline). Postinjury tubular proliferation, measured by PCNA antigen expression, and intragraft macrophage infiltration was not influenced by GLN. In rat renal grafts suffering severe PRI pharmacological preconditioning with GLN attenuates early structural damage, especially tubular cell apoptosis. Stimulation of renal HSP 70 expression could be an important mechanism of GLN-induced cytoprotection. Our findings may have implications for the treatment of delayed graft function in recipients of marginal donor kidneys.

The protective effect of heat shock protein 72 by Doxorubicin in cold ischemia-reperfusion injury of the rat liver

To observe induction of heat shock reaction by pretreatment of Doxorubicin (DXR) in long-term cold preservation-reperfusion injury of the rat liver. The rats were administered intravenously by DXR at a dose of 1 mg/kg body weight in DXR group and by saline in control group. After 48 hours, the rat liver was perfused by using cold University of Wisconsin (UW) solutions and was preserved in UW solution at 4 degrees C for 48 hours. Recipient liver was perfused for 1 and 3 hours after orthotopic liver transplantation. Tumor necrosis factor-alpha (TNF-alpha) mRNA, cytokine-induced neutrophil chemoattractant (CINC) mRNA, macrophage inflammatory protein (MIP-2) mRNA was measured by RT-PCR and heat shock protein 72 (HSP72), nuclear factor-kappaB (NF-kappaB) by Western blot. The serum levels of TNF-alpha, CINC, MIP-2 by ELISA and AST were measured. The survival rate of 7 days was observed. The expression of TNF-alpha mRNA, CINC mRNA and MIP-2 mRNA was stronger in control group than in DXR group. HSP72 was expressed in SA group but not in control group and oppositely NF-kappaB was expressed in control group but not in DXR group. Serum AST, TNF-alpha, CINC and MIP-2 concentrations were significantly lower in DXR group than in control group (P < 0.05). The survival rate of 7 days was significantly higher in DXR group than in control group (50% vs. 0%, P < 0.05). These data suggested that long-term cold ischemia-reperfusion injury was attenuated in liver graft with pretreatment of DXR. The induction of HSP72 may offer protection to hepatocytes by restraining the activation of NF-kappaB and inflammation.

The Nutritional Status Modulates Preservation-Reperfusion Injury in Rat Fatty Liver

The Nutritional Status Modulates Preservation-Reperfusion Injury in Rat Fatty Liver

The Nutritional Status Modulates Preservation-Reperfusion Injury in Rat Fatty Liver

Microcirculation disturbances are essential factors of preservation injury in fatty liver. However, hepatocyte injury is also markedly excessive in fatty liver resulting, at least in part, from energy metabolism impairment and oxidative stress. Thus, this study aimed to determine whether nutritional status influences preservation injury in fatty liver and whether energetic substrate supplementation, alone or with a vasodilator, is protective. Normal or fatty livers induced by a choline-deficient diet were isolated from fed and fasted rats, preserved in University of Wisconsin solution at 4 degrees C for 18 h, and then reperfused with Krebs-Henseleit solution at 37 degrees C for 120 min. Fasted rats with fatty liver were also treated as follows: (1) Glucose supplementation: rats had access to a glucose solution for 18 h prior procurement; (2) Prostaglandin (PG): alprostadil was continuously infused during reperfusion; (3) Combined treatment: Glucose supplementation + PG. Fasting-induced liver injury was significantly greater in fatty than normal liver. In fatty livers from fasted rats, all treatments reduced the alanine aminotransaminase release. Hepatic oxygen consumption improved in the glucose and glucose + PG groups, while PG infusion had no effect. Glucose supplementation did not affect portal pressure, which, in contrast, was reduced in livers receiving PG. Finally, all treatments lowered oxidative injury. Preservation injury in fatty liver is greatly related to nutritional status. Energetic substrate supplementation may represent a clinically feasible protective strategy and a multistep approach adding vasodilators could offer further benefit by acting on different pathogenetic mechanisms.

Ultrastructural evaluation of the protective effect of nitroglycerin in preservation-reperfusion injury of rat lungs

Aim of study Nitric oxide (NO) has been reported as a favorable protective supplement in donor lung preservation, but related ultrastructural studies are rare in the literature. This study was performed to assess the ultrastructural changes and to evaluate the protective effect of NO as donor nitroglycerin (NTG) treatment of ischemia-reperfusion injury in rat lungs. Materials and methods Fifteen Sprague-Dawley rats weighing 300 to 350 g were used in this study. The NTG group (n = 5) used intravenous administration followed by mixture in the University of Wisconsin (UW) solution. For the non-NTG group (n = 5), we injected the same amount of normal saline intravenously followed by admixture in the UW solution. The heart-lung blocks were removed, weighed, and kept in UW solution for 24 hours at 10°C. Reperfusion using human blood diluted in Krebs-Hensleit solution was done for 60 minutes. For the control group (n = 5), we injected the same amount of normal saline intravenously, and removed the lungs with no preservation and reperfusion procedures. Results The non-NTG group showed multiple patchy areas of alveolar collapse with marked swelling and destruction of type I epithelial cells, loss of type II cell surfactant granules, endothelial swelling and papillary projection, interstitial edema, and alveolar macrophages with active phagocytosis of the destroyed materials. The NTG group showed similar ultrastructural changes, but in a lesser severity compared with the non-NTG group. Conclusion Administration of the NTG reduced the ischemia-reperfusion injury in the rat donor lungs. Ultrastructural examination was an effective tool to evaluate the protective effect of NTG in ischemia-reperfusion procedures of donor lungs.

HEPATOCYTE GROWTH FACTOR IMPROVES SURVIVAL OF STEATOTIC LIVER TRANSPLANTATION IN RATS

P71 Background/Aims; Transplantation of liver with severe steatosis may result in primary nonfunction (PNF) of the graft. As described previously, human recombinant hepatocyte growth factor (hrHGF) protected fatty grafts against cold preservation injury in rats. In the present study, we examined the effects of hrHGF on survival and graft function in rat fatty liver transplantation model. Methods; Severe steatosis was induced by a 15-day choline-deficient diet in male Lewis rats. The fatty graft was orthotopically transplanted in normal male Lewis rat after 12 hr cold preservation in UW solution. In the HGF group, donor rats were pretreated with a single bolus of systemic injection of hrHGF (10μg/rat), and 30μg of hrHGF was also added to the perfusion/preservation solution. Animal survival, graft function and morphological changes were compared with those of the untreated group. Liver cell apoptosis was detected by TUNEL stain and liver regeneration was evaluated by BrdU stain at 2 hrs after reperfusion of the liver graft. Results; One week survival was significantly improved in the HGF group compared with the untreated group (83% vs. 33%, p<0.01, n=18/group). AST, ALT and LDH levels in the rinse effluent at the end of cold preservation and serum ALT and LDH at 2 hrs after reperfusion were significantly lower in the HGF group. Although there was no significant difference in hyaluronic acid (HA) level in the rinse effluent at the end of preservation between the two groups, exogenous HA uptake rate (HAUR) at 2 hrs after reperfusion was improved in the HGF group (0.62±0.05 vs. 0.79±0.11, p<0.05). hrHGF pretreatment also significantly reduced the sinusoidal endothelial cell (SEC) apoptotic index (1.65±0.60% vs. 4.90±1.25%, p<0.01) and increased BrdU incorporation index (2.07±0.62% vs. 0.06±0.04%, p<0.01) at 2 hrs after reperfusion. More severe sinusoidal SEC damage was observed by transmission electron microscope in the untreated group than that in HGF group. Conclusions; hrHGF prevented the death of PNF in steatotic liver transplantation by attenuating preservation–reperfusion injury and facilitating recovery of the graft, probably through its antiapoptotic properties and acceleration of liver regeneration at early stage after transplantation.

Impairment of mitochondrial oxidative phosphorylation in rat fatty liver exposed to preservation-reperfusion injury

Background/Aims As the impairment of the cellular energy metabolism contributes to the failure of fatty liver grafts after transplantation, we aimed to determine whether steatosis affects the oxidative phosphorylation activity during preservation. Methods Rat normal and fatty livers were preserved for 18 h and then reperfused with warm oxygenated solution. The oxidative phosphorylation, the F 0F 1-ATPase and the Complex I activities were assessed in isolated mitochondria before and after preservation, and during reperfusion. The ALT release and portal pressure were monitored during reperfusion. Results The baseline phosphorylation activity was similar in normal and steatotic mitochondria. After cold preservation, the respiratory control index and state 3 respiration decreased significantly only in steatotic livers. Reperfusion induced a further deterioration in either group. Contrary to normal liver, uncoupling of fatty liver mitochondria allowed the recovery of the maximal respiration rate only using succinate (Complex II-dependent substrate), but not glutamate-malate (Complex I-dependent). Complex I dysfunction was confirmed spectrophotometrically. The ATPase activity was also significantly lower in fatty livers. Finally, ALT release and portal pressure were greater in steatotic livers. Conclusions The alteration of the oxidative phosphorylation activity during preservation is greatly exacerbated by fatty infiltration likely resulting from damage of the respiratory chain Complex I and of the F 0F 1-ATP synthase.

Improvement of rat liver graft quality by pifithrin-alpha-mediated inhibition of hepatocyte necrapoptosis.

Early graft dysfunction due to ischemia reperfusion injury remains a major clinical challenge in liver transplantation. Because apoptosis may contribute to graft dysfunction, we studied whether transient inhibition of p53 is capable of improving graft quality by reducing apoptotic cell death. Rat livers were harvested and stored for 24 hours or 48 hours in a 4 degrees C solution containing either pifithrin-alpha (PFT-alpha), a specific p53-inhibitor, or the vehicle dimethyl-sulfoxide. Storage was followed by 2-hour reperfusion with 37 degrees C Krebs-Henseleit buffer in an isolated liver perfusion system. Besides caspase-3 activation, apoptosis was quantified using fluorescence microscopy and hematoxylin-eosin histology. Trypan blue allowed for assessment of cell membrane damage, indicating both secondary apoptosis and primary necrosis. Bile flow, oxygen consumption, K(+)-excretion and enzyme release served as indicators of overall graft quality. Upon 2-hour reperfusion, livers developed procaspase activation as well as a mixture of apoptotic and necrotic cell death, representing necrapoptosis. In livers that had been stored for 48 hours, necrapoptotic injury was more pronounced compared with that after 24-hour storage. PFT-alpha effectively attenuated caspase activation as well as hepatocellular apoptosis and necrosis. Attenuation of both modes of cell death by PFT-alpha was associated with improved liver function, metabolism, and integrity. Experiments with the caspase inhibitor z-VAD-fmk confirmed that apoptosis is one mode of cell death in cold ischemia reperfusion. In conclusion, inhibition of p53-dependent apoptosis by PFT-alpha reduces hepatic preservation-reperfusion injury and improves primary organ function and metabolism. Fortification of the preservation solution with PFT-alpha may represent a promising and easily applicable approach to mitigate reperfusion injury in liver transplants.

Time course of proteinuria after living-donor kidney transplantation.

After cadaveric kidney transplantation, preservation-reperfusion damage results in glomerular and tubular proteinuria. There are no data on the time course of proteinuria after living-donor (LD) transplantation. In 10 patients receiving a kidney graft from an LD, the excretion of high molecular weight proteins (albumin, transferrin, and immunoglobulin G) and low molecular weight proteins (beta2-microglobulin and alpha1-microglobulin) was measured at various time points during the first 5 days after transplantation. Immediately after restoration of the circulation, we observed a massive nonselective high molecular weight proteinuria, indicative of glomerular damage. This proteinuria rapidly decreased to slightly elevated values beyond 24 hr after transplantation. Low molecular weight proteinuria, reflecting tubular damage, was also prominent and remained grossly abnormal even at day 5. After LD transplantation, preservation-reperfusion injury causes massive proteinuria during the first 24 hr. Thereafter proteinuria rarely exceeds 1 g per day.

Biliary inorganic phosphate as a tool for assessing cold preservation-reperfusion injury: a study in the isolated perfused rat liver model.

Ischemia-reperfusion injury is a major cause of early graft dysfunction after liver transplantation. The bile flow has been suggested as an index of ischemic damage, and severely impaired bile flow seems to be predictive of poor survival in experimental studies. Looking for injury markers, biliary inorganic phosphate has the potential of being a useful endogenous marker of diminished hepatobiliary function because this anion is excreted in the bile by a paracellular pathway and it can detect changes in permeability. The goal of this study was to evaluate the effects of cold preservation-reperfusion of the liver on bile flow and bile inorganic phosphate and their relationship with storage-related graft failure. The isolated and perfused rat liver was used to evaluate the injury for ischemia-reperfusion. The intrahepatic resistance, lactate dehydrogenase release, and potassium and biliary inorganic phosphate concentration were used to estimate viability and function of freshly isolated or cold-preserved livers. The intrahepatic resistance and the bile flow were consistent and significantly decreased throughout the perfusion time in relation to the increment in storage. Inorganic phosphate is more concentrated in bile from preserved livers, showing an alteration in paracellular pathway, confirmed by the biliary excretion of horseradish peroxidase. After preservation, concentration and excretion of the paracellular marker were increased during the first peak. The second peak appears earlier in preserved livers (10 minutes) with a different shape but without changes in concentration. In conclusion, inorganic phosphate in bile shows changes in paracellular permeability as occurs in livers after 48 hours of cold preservation.

Ischemic preconditioning decreases C-X-C chemokine expression and neutrophil accumulation early after liver transplantation in rats.

Polymorphonuclear neutrophil (PMN) plays a major role in liver ischemia/reperfusion injury. Protective effect of ischemic preconditioning (IP) has been confirmed in liver ischemia/reperfusion injury. The purpose of this study was to investigate the effect of IP on C-X-C chemokine expression and PMNs recruitment early after liver transplantation. Male Sprague-Dawley rats were used as donors and recipients of orthotopic liver transplantation (OLT). The donor liver was stored 24 hours in University of Wisconsin (UW) solution at 4 degrees pre-implantation. IP was done by clamp of the portal vein and hepatic artery of the donor liver for 10 minutes followed by reperfusion for 10 minutes before harvesting. The neutrophilic infiltration in liver was quantified using a myeloperoxidase (MPO) assay. Intragraft expression of macrophage inflammatory protein-2 (MIP-2) mRNA was investigated with in situ hybridization. The serum levels of MIP-2 and tumor necrosis factor (TNF)-alpha were also monitored. After liver transplantation without IP, the hepatic MPO increased significantly compared with sham operated group. In IP group, PMN in liver indicated by MPO was reduced significantly. In situ hybridization showed no MIP-2 mRNA in sham group but dramatic expression in hepatocytes in non-IP group. In IP group, MIP-2 mRNA was significantly down-regulated. Similarly, serum MIP-2 and TNF-alpha levels were significantly elevated in non-IP group and both were reduced in IP group. IP might protect graft liver from preservation-reperfusion injury after OLT through down-regulating C-X-C chemokine expression of hepatocytes, and alleviating PMNs recruitment after reperfusion.

Fatty degeneration exacerbates mitochondrial dysfunction in the rat liver exposed to preservation-reperfusion injury

Fatty degeneration exacerbates mitochondrial dysfunction in the rat liver exposed to preservation-reperfusion injury