Coronary Microvascular Dysfunction in Acute Cholestasis-Induced Liver Injury.

BACKGROUND: Previous studies have shown cardiac abnormalities in acute liver injury, suggesting a potential role in the associated high mortality. METHODS: We designed an experimental study exploring the short-term effects of acute cholestasis-induced liver injury on cardiac function and structure in a rodent bile duct ligation (BDL) model to elucidate the potential interplay. Thirty-seven male Sprague-Dawley rats were subjected to BDL surgery (n = 28) or served as sham-operated (n = 9) controls. Transthoracic echocardiography, Doppler evaluation of the left anterior descending coronary artery, and myocardial contrast echocardiography were performed at rest and during adenosine and dobutamine stress 5 days after BDL. Immunohistochemical staining of myocardial tissue samples for hypoxia and inflammation as well as serum analysis were performed. RESULTS: BDL animals exhibited acute liver injury with elevated transaminases, bilirubin, and total circulating bile acids (TBA) 5 days after BDL (TBA control: 0.81 ± 2.54 µmol/L vs. BDL: 127.52 ± 57.03 µmol/L; p < 0.001). Concurrently, cardiac function was significantly impaired, characterized by reduced cardiac output (CO) and global longitudinal strain (GLS) in the echocardiography at rest and under pharmacological stress (CO rest control: 120.6 ± 24.3 mL/min vs. BDL 102.5 ± 16.6 mL/min, p = 0.041; GLS rest control: -24.05 ± 3.8% vs. BDL: -18.5 ± 5.1%, p = 0.01). Myocardial perfusion analysis revealed a reduced myocardial blood flow at rest and a decreased coronary flow velocity reserve (CFVR) under dobutamine stress in the BDL animals (CFVR control: 2.1 ± 0.6 vs. BDL: 1.7 ± 0.5 p = 0.047). Immunofluorescence staining indicated myocardial hypoxia and increased neutrophil infiltration. CONCLUSIONS: In summary, acute cholestasis-induced liver injury can lead to impaired cardiac function mediated by coronary microvascular dysfunction, suggesting that major adverse cardiac events may contribute to the mortality of acute liver failure. This may be due to endothelial dysfunction and direct bile acid signaling.

A Potential Association between Ribonuclease 1 Dynamics in the Blood and the Outcome in COVID-19 Patients.

The COVID-19 pandemic caused by the new SARS-CoV-2 coronavirus is the most recent and well-known outbreak of a coronavirus. RNase 1 is a small endogenous antimicrobial polypeptide that possesses antiviral activity against viral diseases. In this study, we investigated a potential association between ribonuclease 1 and the outcome in COVID-19 patients and the impact of increased and decreased RNase 1 levels serum during the course of the disease. Therefore, two patient populations, Cohort A (n = 35) and B (n = 80), were subclassified into two groups, in which the RNase 1 concentration increased or decreased from time point one to time point two. We show that the RNase 1 serum levels significantly increased in the increasing group of both cohorts (p = 0.0171; p < 0.0001). We detect that patients in the increasing group who died had significantly higher RNase 1 serum levels at both time points in Cohort A (p = 0.0170; p = 0.0393) and Cohort B (p = 0.0253; p = 0.0034) than patients who survived. Additionally, we measured a significant correlation of RNase 1 serum levels with serum creatinine as well as creatinine clearance in the increasing and decreasing group at both time points of Cohort A. Based on these results, there is now good evidence that RNase 1 may play a role in renal dysfunction associated with ICU COVID-19 patients and that increasing RNase 1 serum level may be a potential biomarker to predict outcome in COVID-19 patients.

DAMPs Released from Proinflammatory Macrophages Induce Inflammation in Cardiomyocytes via Activation of TLR4 and TNFR.

Cardiac dysfunction is a life-threatening complication in sepsis. Upon infection and cardiac stress, the cardiac macrophage population expands. Recruited macrophages exhibit a predominantly proinflammatory phenotype and release danger-associated molecular patterns (DAMPs) that contribute to cardiac dysfunction. However, the underlying pathomechanisms are highly complex and not fully understood. Here, we utilized an indirect macrophage-cardiomyocyte co-culture model to study the effects of proinflammatory macrophages on the activation of different cardiac receptors (TLR3, TLR4, and TNFR) and their role in cardiac inflammation and caspase-3/7 activation. The stimulation of cardiomyocytes with conditioned medium of LPS-stimulated macrophages resulted in elevated IL-6 protein concentrations and relative IL-6 and TNFα mRNA levels. Conditioned medium from LPS-stimulated macrophages also induced NFκB translocation and increased caspase-3/7 activation in cardiomyocytes. Analyzing the role of different cardiac receptors, we found that TLR4 and TNFR inhibition reduces cardiac inflammation and that the inhibition of TNFR prevents NFκB translocation into the nuclei of cardiomyocytes, induced by exposure to conditioned medium of proinflammatory macrophages. Moreover, we demonstrated that TLR3 inhibition reduces macrophage-mediated caspase-3/7 activation. Our results suggest that the immune response of macrophages under inflammatory conditions leads to the release of DAMPs, such as eRNA and cytokines, which in turn induce cardiomyocyte dysfunction. Thus, the data obtained in this study contribute to a better understanding of the pathophysiological mechanisms of cardiac dysfunction.

The Potential Impact of Heparanase Activity and Endothelial Damage in COVID-19 Disease.

SARS-CoV-2 was first detected in 2019 in Wuhan, China. It has been found to be the most pathogenic virus among coronaviruses and is associated with endothelial damage resulting in respiratory failure. Determine whether heparanase and heparan sulfate fragments, biomarkers of endothelial function, can assist in the risk stratification and clinical management of critically ill COVID-19 patients admitted to the intensive care unit. We investigated 53 critically ill patients with severe COVID-19 admitted between March and April 2020 to the University Hospital RWTH Aachen. Heparanase activity and serum levels of both heparanase and heparan sulfate were measured on day one (day of diagnosis) and day three in patients with COVID-19. The patients were classified into four groups according to the severity of ARDS. When compared to baseline data (day one), heparanase activity increased and the heparan sulfate serum levels decreased with increasing severity of ARDS. The heparanase activity significantly correlated with the lactate concentration on day one (r = 0.34, p = 0.024) and on day three (r = 0.43, p = 0.006). Heparanase activity and heparan sulfate levels correlate with COVID-19 disease severity and outcome. Both biomarkers might be helpful in predicting clinical course and outcomes in COVID-19 patients.

Comparison of the Hemocompatibility of an Axial and a Centrifugal Left Ventricular Assist Device in an In Vitro Test Circuit.

BACKGROUND: Hemocompatibility of left ventricular assist devices is essential for preventing adverse events. In this study, we compared the hemocompatibility of an axial-flow (Sputnik) to a centrifugal-flow (HeartMate 3) pump. METHODS: Both pumps were integrated into identical in vitro test circuits, each filled with 75 mL heparinized human blood of the same donor. During each experiment (n = 7), the pumps were operated with equal flow for six hours. Blood sampling and analysis were performed on a regular schedule. The analytes were indicators of hemolysis, coagulation activation, platelet count and activation, as well as extracellular vesicles. RESULTS: Sputnik induced higher hemolysis compared to the HeartMate 3 after 360 min. Furthermore, platelet activation was higher for Sputnik after 120 min onward. In the HeartMate 3 circuit, the platelet count was reduced within the first hour. Furthermore, Sputnik triggered a more pronounced increase in extracellular vesicles, a potential trigger for adverse events in left ventricular assist device application. Activation of coagulation showed a time-dependent increase, with no differences between both groups. CONCLUSIONS: This experimental study confirms the hypothesis that axial-flow pumps may induce stronger hemolysis compared to centrifugal pumps, coming along with larger amounts of circulating extracellular vesicles and a stronger PLT activation.

Repetitive Treatment with Volatile Anesthetics Does Not Affect the In Vivo Plasma Concentration and Composition of Extracellular Vesicles in Rats.

BACKGROUND: Anesthetic-induced preconditioning (AIP) with volatile anesthetics is a well-known experimental technique to protect tissues from ischemic injury or oxidative stress. Additionally, plasmatic extracellular vesicle (EV) populations and their cargo are known to be affected by AIP in vitro, and to provide organ protective properties via their cargo. We investigated whether AIP would affect the generation of EVs in an in vivo rat model. METHODS: Twenty male Sprague Dawley rats received a repetitive treatment with either isoflurane or with sevoflurane for a duration of 4 or 8 weeks. EVs from blood plasma were characterized by nanoparticle tracking analysis, transmission electron microscopy (TEM) and Western blot. A scratch assay (H9C2 cardiomyoblast cell line) was performed to investigate the protective capabilities of the isolated EVs. RESULTS: TEM images as well as Western blot analysis indicated that EVs were successfully isolated. The AIP changed the flotillin and CD63 expression on the EV surface, but not the EV concentration. The scratch assay did not show increased cell migration and/or proliferation after EV treatment. CONCLUSION: AIP in rats changed the cargo of EVs but had no effect on EV concentration or cell migration/proliferation. Future studies are needed to investigate the cargo on a miRNA level and to investigate the properties of these EVs in additional functional experiments.

Phosphatidylserine Supplementation as a Novel Strategy for Reducing Myocardial Infarct Size and Preventing Adverse Left Ventricular Remodeling.

Phosphatidylserines are known to sustain skeletal muscle activity during intense activity or hypoxic conditions, as well as preserve neurocognitive function in older patients. Our previous studies pointed out a potential cardioprotective role of phosphatidylserine in heart ischemia. Therefore, we investigated the effects of phosphatidylserine oral supplementation in a mouse model of acute myocardial infarction (AMI). We found out that phosphatidylserine increases, significantly, the cardiomyocyte survival by 50% in an acute model of myocardial ischemia-reperfusion. Similar, phosphatidylserine reduced significantly the infarcted size by 30% and improved heart function by 25% in a chronic model of AMI. The main responsible mechanism seems to be up-regulation of protein kinase C epsilon (PKC-ε), the main player of cardio-protection during pre-conditioning. Interestingly, if the phosphatidylserine supplementation is started before induction of AMI, but not after, it selectively inhibits neutrophil's activation, such as Interleukin 1 beta (IL-1ß) expression, without affecting the healing and fibrosis. Thus, phosphatidylserine supplementation may represent a simple way to activate a pre-conditioning mechanism and may be a promising novel strategy to reduce infarct size following AMI and to prevent myocardial injury during myocardial infarction or cardiac surgery. Due to the minimal adverse effects, further investigation in large animals or in human are soon possible to establish the exact role of phosphatidylserine in cardiac diseases.

The Role of Ribonuclease 1 and Ribonuclease Inhibitor 1 in Acute Kidney Injury after Open and Endovascular Thoracoabdominal Aortic Aneurysm Repair.

Acute kidney injury (AKI) is one of the most common post-operative complications and is closely associated with increased mortality after open and endovascular thoracoabdominal aortic aneurysm (TAAA) repair. Ribonuclease (RNase) 1 belongs to the group of antimicrobial peptides elevated in septic patients and indicates the prediction of two or more organ failures. The role of RNase 1 and its antagonist RNase inhibitor 1 (RNH1) after TAAA repair is unknown. In this study, we analyzed RNase 1 and RNH1 serum levels in patients undergoing open (n = 14) or endovascular (n = 19) TAAA repair to determine their association with post-operative AKI and in-hospital mortality. Increased RNH1 serum levels after open TAAA repair as compared with endovascular TAAA repair immediately after surgery and 12, 48, and 72 h after surgery (all p < 0.05) were observed. Additionally, elevated RNase 1 and RNH1 serum levels 12, 24, and 48 h after surgery were shown to be significantly associated with AKI (all p < 0.05). RNH1 serum levels before and RNase 1 serum levels 12 h after TAAA repair were significantly correlated with in-hospital mortality (both p < 0.05). On the basis of these findings, RNase 1 and RNH1 may be therapeutically relevant and may represent biomarkers for post-operative AKI and in-hospital mortality.

The Role of Macrophage Migration Inhibitory Factor in Remote Ischemic Conditioning Induced Hepatoprotection in a Rodent Model of Liver Transplantation.

BACKGROUND: Macrophage migration inhibitory factor (MIF) is an important stress-regulating mediator of acute ischemia/reperfusion (I/R) injury and ischemic conditioning. The present study aimed to investigate whether MIF is involved in the effects of remote ischemic conditioning (RIC) in a rat model of orthotopic liver transplantation (OLT). METHODS: OLTs were performed in male Lewis rats (245 g-340 g). Recipients were allocated in a randomized fashion into three experimental groups: remote preconditioning-RIPC, remote post-conditioning-RIPOST, control. RIC was applied as 4×5-5 min I/R via clamping of the infrarenal aorta. Animals were followed for 1, 3, 24, 168 h post-reperfusion (n = 6 recipient/group/time point). Graft micro- and macrocirculation and hepatocellular damage were assessed. Messenger ribonucleic acid (mRNA) expression, serum, and tissue protein levels of MIF, as well as additional markers of I/R injury, were measured. RESULTS: RIC resulted in a prominent downregulation of MIF mRNA, serum, and tissue protein. Compared with control, hepatocellular damage was significantly mitigated after RIPC or RIPOST (serum ALT; RIPC, RIPOST vs. Control, P = 0.008, P = 0.030, respectively). Graft circulation was better preserved in the RIC groups. Furthermore, there was a significant positive correlation between serum MIF and transaminase levels (r = 0.330; P = 0.02). RIC showed a significant effect on iNOS and STAT5 mRNA expressions. Supporting findings were obtained from the measurements of tissue CXCL12 mRNA expression and pAkt/Akt, pErk/Erk. CONCLUSION: In this sophisticated experimental model of OLT, RIC-induced hepatoprotective effects were associated with a downregulation of MIF at mRNA and protein levels, suggesting the role of MIF as a mediator in RIC-induced protection following OLT.

In vitro head-to-head comparison of anticoagulation properties of two heparin brands in a human blood miniature mock loop.

OBJECTIVES: The first aim was the development of a human blood miniature mock-loop system consisting of 2 identical extracorporeal circuits, which enable systematic head-to-head comparisons of test substances. In a second step, we evaluated the suitability of the mock-loop system, by comparing 2 different brands of heparin (ROTEXMEDICA vs B.BRAUN), which have showed different anticoagulation capacities in the clinic. METHODS: For 1 experiment (18 in total), blood of the same healthy human donor was divided into 2 portions (2 × 50 ml), heparinized with 37.5 IU⋅ml-1 of the competing test substances and diluted to a haematocrit value of 20-25%. Each mock loop was filled with 70 ml, and in vivo heparin degradation was simulated in 3 different groups by protamine application, representing 0%, 50% and 100% heparin antagonization. At baseline, 5, 60, 120, 240 and 360 min, blood samples were taken to perform thromboelastometry, flow cytometry, haemolysis and general haemostasis analysis. RESULTS: Blood pressure, blood flow and blood temperature within the loops remained stable for 6 h in all groups. After 6 h, in the 100% antagonized ROTEXMEDICA heparin group, significantly increased haemolysis (148.7 ± 80 mg⋅dl-1 vs 57.5 ± 15.8 mg⋅dl-1), activated platelets (8 ± 3.8% vs 3.3 ± 0.7%), D-dimers (7376 ± 7144 ng ml-1 vs 576.2 ± 190 ng ml-1) and fulminant blood clots were detected. CONCLUSIONS: Our in vitro system is suitable for the detection of reduced anticoagulation capacity of a test drug, which was reported in vivo previously.

Soluble CD74 Reroutes MIF/CXCR4/AKT-Mediated Survival of Cardiac Myofibroblasts to Necroptosis.

Background Although macrophage migration inhibitory factor ( MIF ) has been demonstrated to mediate cardioprotection in ischemia/reperfusion injury and antagonize fibrotic effects through its receptor, CD 74, the function of the soluble CD 74 receptor ectodomain ( sCD 74) and its interaction with circulating MIF have not been explored in cardiac disease. Methods and Results Cardiac fibroblasts were isolated from hearts of neonatal mice and differentiated into myofibroblasts. Co-treatment with recombinant MIF and sCD 74 induced cell death ( P<0.001), which was mediated by receptor-interacting serine/threonine-protein kinase ( RIP) 1/ RIP 3-dependent necroptosis ( P=0.0376). This effect was specific for cardiac fibroblasts and did not affect cardiomyocytes. Gene expression analyses using microarray and RT - qPCR technology revealed a 4-fold upregulation of several interferon-induced genes upon co-treatment of myofibroblasts with sCD 74 and MIF (Ifi44: P=0.011; Irg1: P=0.022; Clec4e: P=0.011). Furthermore, Western blot analysis confirmed the role of sCD 74 as a modulator of MIF signaling by diminishing MIF -mediated protein kinase B ( AKT) activation ( P=0.0197) and triggering p38 activation ( P=0.0641). We obtained evidence that sCD 74 inhibits MIF -mediated survival pathway through the C-X-C chemokine receptor 4/ AKT axis, enabling the induction of CD 74-dependent necroptotic processes in cardiac myofibroblasts. Preliminary clinical data revealed a lowered sCD 74/ MIF ratio in heart failure patients (17.47±10.09 versus 1.413±0.6244). Conclusions These findings suggest that treatment of cardiac myofibroblasts with sCD 74 and MIF induces necroptosis, offering new insights into the mechanism of myofibroblast depletion during scar maturation. Preliminary clinical data provided first evidence about a clinical relevance of the sCD 74/ MIF axis in heart failure, suggesting that these proteins may be a promising target to modulate cardiac remodeling and disease progression in heart failure.

Limb remote ischemic conditioning of the recipient protects the liver in a rat model of arterialized orthotopic liver transplantation.

BACKGROUND: Ischemic-reperfusion (IR) injury still represents a major concern in clinical transplantation, especially in the era of extreme organ shortage and extended criteria donor organs. In the present study we aimed to investigate the hepatoprotective effects of remote ischemic conditioning (RIC) in a rat model of arterialized orthotopic liver transplantation (OLT). METHODS: Male Lewis rats were used (n = 144 / 72 OLT cases; 240-340g) as donors and recipients. Livers were flushed and stored in 4°C HTK-solution for 8h before implantation. Recipients were randomly allocated into three experimental groups: RIC 1, RIC 2, Control. In RIC 1, RIC 2 groups, RIC was applied in the recipient before hepatectomy or after reperfusion (4x5-5min IR via clamping the infrarenal aorta), respectively. Animals were sacrificed at 1, 3, 24, 168h post-reperfusion (n = 6 recipient/group/time point). Hepatocellular injury, graft circulation, serum cytokines, tissue redox-stress and adenosine-triphosphate (ATP) levels have been assessed. Additional markers were analyzed, using Western blotting and reverse-transcription polymerase chain reaction. RESULTS: RIC 1 group showed significantly (p<0.05) improved portal venous and microcirculation flow as well as velocity. RIC has significantly reduced tissue injury according to the serum levels of transaminases and results of histopathological evaluation. Reduced TUNEL-staining (p<0.01 RIC 1-2 vs. Control) and elevated pBAD/BAD ratio was detected in the RIC groups (p<0.01 RIC 1 vs. Control). Supporting findings were obtained from measurements of serum IL-10 as well as tissue malondialdehyde and ATP levels. Hemoxygenase-1 (HO-1) mRNA-expression was significantly higher in RIC 1 compared to Control (p<0.05 RIC 1 vs. Control). CONCLUSION: These results suggest that RIC might confer potent protection against the detrimental effects of IR injury including tissue damage, apoptosis, graft circulation, inflammation, tissue energetic status in OLT. HO-1 overexpression might play an orchestrating role in RIC mediated organ protection. An earlier intervention (RIC 1 protocol) was more effective than remote conditioning after graft reperfusion.

Characterization of extracellular vesicles derived from cardiac cells in an in vitro model of preconditioning.

Preconditioning is a promising technique to protect the heart from ischaemia-reperfusion injury. In this context, the crosstalk between different cardiac cell types and especially the exchange of cardioprotective mediators has come into the focus of current research. Recently, extracellular vesicles (EVs), nano-sized structures, emerged as possible communication mediators. They are taken up by recipient cells and can alter gene expression or activate intracellular signal cascades. It has been shown that all cardiac cell types are able to secrete EVs, but so far the influence of an in vitro preconditioning stimulus on EV concentration and composition has not been investigated. Therefore, we stimulated primary cardiac myocytes and fibroblasts from neonatal rats, as well as H9c2 cells, with two known in vitro preconditioning stimuli: hypoxia or isoflurane. EVs were isolated from cell culture supernatants 48 h after stimulation by differential centrifugation and size exclusion chromatography. They were characterized by transmission electron microscopy, tunable resistive pulse sensing, miRNA array and Western blot analysis. The detected EVs had the typical cup-shaped morphology and a size of about 150 nm. No significant differences in EV concentration were observed between the different groups. The protein and miRNA load was affected by in vitro preconditioning with isoflurane or hypoxia. EV markers like Alix, CD63, flotillin-1 and especially heat shock protein 70 were significantly up-regulated by the treatments. Several miRNAs like miR-92b-3p, miR-761 and miR-101a-5p were also significantly affected. A migration assay confirmed the physiological benefit of these EVs. Taken together, our findings show that a model of in vitro preconditioning of cardiac cells does not influence EV concentration but strongly regulates the EV cargo and affects migration. This might indicate a role for EV-mediated communication in isoflurane- and hypoxia-induced in vitro preconditioning.

Argon Induces Protective Effects in Cardiomyocytes during the Second Window of Preconditioning.

Increasing evidence indicates that argon has organoprotective properties. So far, the underlying mechanisms remain poorly understood. Therefore, we investigated the effect of argon preconditioning in cardiomyocytes within the first and second window of preconditioning. Primary isolated cardiomyocytes from neonatal rats were subjected to 50% argon for 1 h, and subsequently exposed to a sublethal dosage of hypoxia (<1% O2) for 5 h either within the first (0-3 h) or second window (24-48 h) of preconditioning. Subsequently, the cell viability and proliferation was measured. The argon-induced effects were assessed by evaluation of mRNA and protein expression after preconditioning. Argon preconditioning did not show any cardioprotective effects in the early window of preconditioning, whereas it leads to a significant increase of cell viability 24 h after preconditioning compared to untreated cells (p = 0.015) independent of proliferation. Argon-preconditioning significantly increased the mRNA expression of heat shock protein (HSP) B1 (HSP27) (p = 0.048), superoxide dismutase 2 (SOD2) (p = 0.001), vascular endothelial growth factor (VEGF) (p < 0.001) and inducible nitric oxide synthase (iNOS) (p = 0.001). No difference was found with respect to activation of pro-survival kinases in the early and late window of preconditioning. The findings provide the first evidence of argon-induced effects on the survival of cardiomyocytes during the second window of preconditioning, which may be mediated through the induction of HSP27, SOD2, VEGF and iNOS.

Comparison of Aerobic Preservation by Venous Systemic Oxygen Persufflation or Oxygenated Machine Perfusion of Warm-Ischemia-Damaged Porcine Kidneys.

BACKGROUND/AIM: The global shortage of donor organs for transplantation has necessitated the expansion of the organ pool through increased use of organs from less ideal donors. Venous systemic oxygen persufflation (VSOP) and oxygenated machine perfusion (OMP) have previously demonstrated beneficial results compared to cold storage (CS) in the preservation of warm-ischemia-damaged kidney grafts. The aim of this study was to compare the efficacy of VSOP and OMP for the preservation of warm-ischemia-damaged porcine kidneys using the recently introduced Ecosol preservation solution compared to CS using Ecosol or histidine-tryptophan-ketoglutarate solution (HTK). MATERIALS AND METHODS: Kidneys from German Landrace pigs (n = 5/group) were retrieved and washed out with either Ecosol or HTK after 45 min of clamping of the renal pedicle. As controls, kidneys without warm ischemia, cold stored for 24 h in HTK, were employed. Following 24 h of preservation by VSOP, OMP, CS-Ecosol, or CS-HTK, renal function and damage were assessed during 1 h using the isolated perfused porcine kidney model. RESULTS: During reperfusion, urine production was significantly higher in the VSOP and OMP groups than in the CS-HTK group; however, only VSOP could demonstrate lower urine protein concentrations and fractional excretion of sodium, which did not differ from the non-warm-ischemia-damaged control group. VSOP, CS-Ecosol, and controls showed better maintenance of the acid-base balance than CS-HTK. Reduced lipid peroxidation, as reflected in postreperfusion tissue thiobarbituric acid-reactive substance levels, was observed in the VSOP group compared to the OMP group, and the VSOP and CS-Ecosol groups had concentrations similar to the controls. The ratio of reduced to oxidized glutathione was higher in the VSOP, OMP, and CS-Ecosol groups than in the CS-HTK group and controls, with a higher ratio in the VSOP than in the OMP group. CONCLUSION: VSOP was associated with mitigation of oxidative stress in comparison to OMP and CS. Preservation of warm-ischemia-damaged porcine kidneys by VSOP was improved compared to OMP and CS, and was comparable to preservation of non-warm-ischemia-damaged cold-stored kidneys.

Improved preservation of warm ischemia-damaged porcine kidneys after cold storage in Ecosol, a novel preservation solution.

BACKGROUND: Ecosol, an extracellular-type, colloid-based preservation solution, has recently been introduced for washout, cold storage, and machine perfusion preservation of kidney grafts. Here, we assessed the efficacy of Ecosol compared to the widely used Histidine-Tryptophan-Ketoglutarate solution (HTK) for 24-h cold storage preservation of warm ischemia-damaged kidney grafts. MATERIAL AND METHODS: Before recovery, warm ischemia was induced by clamping the renal pedicle for 45-min. Thereafter, kidneys were washed-out and cold-stored for 24-h in Ecosol or HTK solution. Kidneys recovered without warm ischemia and cold-stored for 24-h in HTK served as controls (n=5). Renal function and damage parameters were assessed during 1-h normothermic reperfusion using the isolated perfused porcine kidney model. RESULTS: Renal function did not differ between Ecosol and controls and was significantly reduced in HTK compared to controls. Total output of urine was higher in Ecosol compared to HTK. Intrarenal resistance and urine protein concentrations in Ecosol were lower compared to HTK and equal to controls. In the Ecosol group, oxygen consumption during reperfusion was higher and reduced tissue lipid peroxidation products were detected compared to HTK. CONCLUSIONS: The preservation quality of warm ischemia-damaged, cold-stored porcine kidneys was improved using the recently developed Ecosol preservation solution compared to HTK.

Determination of the preferred conditions for the isolated perfusion of porcine kidneys.

BACKGROUND: The isolated perfused porcine kidney (IPPK) model has been the method of choice for the early preclinical evaluation of kidney graft preservation techniques. The preferred reperfusion conditions have not yet been determined. Here, we examined the effects of pressure- or flow-controlled perfusion and oxygenation by pure oxygen or carbogen (95% O2/5% CO2) on normothermic reperfusion in the IPPK model. METHODS: Porcine kidneys were cold-stored for 24 h in histidine-tryptophan-ketoglutarate solution and reperfused for 1 h with normothermic whole blood/Krebs-Henseleit buffer medium (20/80%). Kidneys (n = 5/group) were flow-controlled reperfused with pure oxygen (1 ml/min/g; Flow-O2) or pressure-controlled reperfused (85 mm Hg mean arterial pressure) and oxygenated with either pure oxygen (Pressure-O2) or carbogen (Pressure-O2/CO2). Renal function and damage were assessed during reperfusion and NGAL and HIF-1α levels were analyzed using an ELISA. RESULTS: Pressure-O2 and Pressure-O2/CO2 were associated with significantly better renal hemodynamics and acid-base homeostasis compared to Flow-O2. Urine protein concentrations and the fractional excretion of sodium were lower with both Pressure-O2 and Pressure-O2/CO2 than with Flow-O2. NGAL and HIF-1α levels were also lower with Pressure-O2 and Pressure-O2/CO2 than with Flow-O2. Only Pressure-O2/CO2 could demonstrate a significantly increased urine production compared to Flow-O2. The structural integrity was well preserved in the Pressure-O2 and Pressure-O2/CO2 groups, whereas diffuse and global glomerular destruction was observed in the Flow-O2 group. CONCLUSION: In the IPPK model, the application of pressure-controlled reperfusion with carbogen oxygenation, and to a lesser extent with pure oxygen, maintained physiological renal function for 1 h, thus providing a reliable and reproducible ex vivo evaluation of kidney preservation quality.

The role of macrophage migration inhibitory factor in anesthetic-induced myocardial preconditioning.

INTRODUCTION: Anesthetic-induced preconditioning (AIP) is known to elicit cardioprotective effects that are mediated at least in part by activation of the kinases AMPK and PKCε as well as by inhibition of JNK. Recent data demonstrated that the pleiotropic cytokine macrophage migration inhibitory factor (MIF) provides cardioprotection through activation and/or inhibition of kinases that are also known to mediate effects of AIP. Therefore, we hypothesized that MIF could play a key role in the AIP response. METHODS: Cardiomyocytes were isolated from rats and subjected to isoflurane preconditioning (4 h; 1.5 vol. %). Subsequently, MIF secretion and alterations in the activation levels of protective kinases were compared to a control group that was exposed to ambient air conditions. MIF secretion was quantified by ELISA and AIP-induced activation of protein kinases was assessed by Western blotting of cardiomyocyte lysates after isoflurane treatment. RESULTS: In cardiomyocytes, preconditioning with isoflurane resulted in a significantly elevated secretion of MIF that followed a biphasic behavior (30 min vs. baseline: p = 0.020; 24 h vs. baseline p = 0.000). Moreover, quantitative polymerase chain reaction demonstrated a significant increase in MIF mRNA expression 8 h after AIP. Of note, activation of AMPK and PKCε coincided with the observed peaks in MIF secretion and differed significantly from baseline. CONCLUSIONS: These results suggest that the pleiotropic mediator MIF is involved in anesthetic-induced preconditioning of cardiomyocytes through stimulation of the protective kinases AMPK and PKCε.