Andreas Lun

Methylprednisolone therapy in deceased donors reduces inflammation in the donor liver and improves outcome after liver transplantation: a prospective randomized controlled trial.

Objective:To investigate potential beneficial effects of donor treatment with methylprednisolone on organ function and outcome after liver transplantation. Summary Background Data:It is proven experimentally and clinically that the brain death of the donor leads to increased levels of inflammatory cytokines and is followed by an intensified ischemia/reperfusion injury after organ transplantation. In experiments, donor treatment with steroids successfully diminished these effects and led to better organ function after transplantation. Methods:To investigate whether methylprednisolone treatment of the deceased donor is applicable to attenuate brain death-associated damage in clinical liver transplantation we conducted a prospective randomized treatment-versus-control study in 100 deceased donors. Donor treatment (n = 50) consisted of 250 mg methylprednisolone at the time of consent for organ donation and a subsequent infusion of 100 mg/h until recovery of organs. A liver biopsy was taken immediately after laparotomy and blood samples were obtained after brain death diagnosis and before organ recovery. Cytokines were assessed by real-time reverse transcriptase-polymerase chain reaction. Soluble serum cytokines were measured by cytometric bead array system. Results:After methylprednisolone treatment, steroid plasma levels were significantly higher (P < 0.05), and a significant decrease in soluble interleukins, monocyte chemotactic protein-1, interleukin-2, interleukin-6, tumor necrosis factor-&agr;, and inducible protein-10 was observed. Methylprednisolone treatment resulted in a significant downregulation of intercellular adhesion molecule-1, tumor necrosis factor-&agr;, major histocompatibility complex class II, Fas-ligand, inducible protein-10, and CD68 intragraft mRNA expression. Significantly ameliorated ischemia/reperfusion injury in the posttransplant course was accompanied by a decreased incidence of acute rejection. Conclusions:Our present study verifies the protective effect of methylprednisolone treatment in deceased donor liver transplantation, suggesting it as a potential therapeutical approach.

The role of automated urine particle flow cytometry in clinical practice

Urine particle flow cytometers (UFC) have improved count precision and accuracy compared to visual microscopy and offer significant labor saving. The absence of an internationally recognized reference measurement procedure, however, is a serious drawback to their validation. Chamber counting by phase contrast microscopy of supravitally-stained uncentrifuged urine is considered the best candidate for reference. The UF-100 (Sysmex Corporation, Japan) identifies RBC, WBC, squamous epithelial cells, transitional epithelial and renal tubular cells (SRC), bacteria, hyaline and inclusional casts, yeast-like cells, crystals and spermatozoa, using argon laser flow cytometry. Evaluations have established acceptable linearity over useful working ranges, with an imprecision that is consistently and significantly less than microscopy, and with negligible carry-over. Comparisons of UFC with chamber counts, quantitative urine microscopy, sediment counts, test strips, bacterial culture and urine density are reviewed. Clinical studies include diagnosis and monitoring of urinary tract infection; localization of the sites of hematuria; and diagnosis, monitoring and exclusion of renal disease. The most popular approach is to combine test strips with UFC for primary screening either always by both methods or by using test strips for analytes unrelated to particles analyzed by UFC. Expert systems now exist combining both test modalities based on user definable decision rules. The implementation of such a strategy significantly reduces microscopy review and saves time and expense without diminishing clinical utility.

Effects of Endothelin Receptor Antagonists on the Progression of Diabetic Nephropathy

Background: Diabetic nephropathy is the leading cause of end-stage renal disease in European countries and is associated with an enhanced renal synthesis of endothelin (ET)-1. ETs are – beside its potent vasoconstrictor properties – very potent profibrotic acting paracrine hormones especially in the kidney. Methods: We analyzed in rats with streptozotocin-induced diabetes the effects of an ETA-type (ETA) receptor antagonist (LU 135252) in comparison to a combined ETA/ETB receptor antagonist (LU 224332) on the expression of interstitial and glomerular collagen type I, III and IV as well as on fibronectin and laminin by quantitative immunohistochemistry using a computer-aided image analysis system. Global glomerular matrix deposition was analyzed after PAS staining. In addition to the morphometric examination of the kidneys, we also investigated GFR, urinary albumin and total protein excretion. The diabetic rats were treated for 36 weeks. Results: Treatment with either LU 135252 or LU 224332 normalized the amount of PAS-positive material within the glomeruli. The expression of glomerular fibronectin and type IV collagen was increased 36 weeks after induction of diabetes. The overexpression of these two matrix proteins within the glomeruli of diabetic rats was completely abolished by both ET receptor antagonists, whereas protein excretion was only reduced by about 50% as compared to diabetic rats without treatment. Conclusion: The present study indicates that ETA receptor antagonists as well as combined ETA/ETB receptor antagonists reduce proteinuria and completely normalize the renal matrix protein expression in hyperglycemic rats with streptozotocin-induced diabetes. The antifibrotic effect seems to be mediated via the ETA receptor. ET receptor antagonists might be a new approach in the treatment of diabetic nephropathy.

DHEA-S plasma levels and incidence of Alzheimer's disease

BACKGROUND Cross-sectional studies controlling for age and gender reported a relationship between Alzheimers disease and low dehydroepiandrosterone sulphate (DHEA-S) plasma levels. Prospective data with sufficient control for confounding factors are lacking. METHODS A nested case-control study examined baseline DHEA-S in participants of the Berlin Aging Study. Cases (n = 14) developed dementia of the Alzheimer type within 3 years. Control group A (n = 14) was matched for gender, age, multimorbidity, and immobility. Control group B (n = 13) was matched for gender and age and comprised participants free from multimorbidity, immobility, multimedication, need of help, incontinence, visual impairment, hearing impairment, and depression. RESULTS The mean plasma DHEA-S concentration of case subjects was 1.02 +/- 0.61 mumol/L. Both control groups had higher mean DEHA-S levels, in control group A, it was 1.89 +/- 1.24 mumol/L (p = .012) and in control group B 1.70 +/- 1.38 mumol/L (p = .093). CONCLUSIONS This population-based prospective study supports the role of DHEA-S as a risk factor for Alzheimers disease.

Renal Elimination of Troponin T and Troponin I

Cardiovascular complications represent the predominant cause of death in patients in the terminal stage of renal failure. Increased concentrations of cardiac troponin T (cTnT) may be a valuable predictor of cardiac risk (1)(2). However, cardiac troponin I (cTnI), clinical symptoms, and electrocardiogram (ECG) indications may be absent in patients with a positive cTnT. This may be attributable to instability of the cTnI molecule (3) or dissimilar glomerular filtration of cTnT and cTnI (4). Positive cTnT values are of cardiac origin because the second generation of cTnT assays will not detect cTnT isoforms expressed in the skeletal muscle of hemodialysis patients (5). We therefore measured the cardiac troponins cTnT and cTnI in the plasma and urine of selected patients differing in their kidney function. We examined 24 patients with increased plasma cTnT. Patients were grouped according to their basic disease and renal function as follows: Group A included five patients (patients 1–5) who had suffered an acute myocardial infarction and three patients (patients 6–8) with cardiac damage as a result of heart surgery, all with normal or only slightly restricted glomerular filtration rate of >80 mL/min. All eight patients were male, with a mean (SD) age of 63 (11) years. Patients had clinically typical chest pain (with the exception of patient 8), and electrocardiography (ECG) showed signs of old myocardial infarction, signs of ST-segment reduction or elevation >0.1 mV with and without chest pain, or signs any arrhythmia of unknown origin. Group B included two patients (patients 9 and 10) who had suffered an acute myocardial infarction and six patients (patients 11–16) with cardiac damage as a result of heart surgery; all had a substantially restricted glomerular filtration rate (only patient 9 had values for creatinine and creatinine clearance that were within the appropriate reference intervals). All patients in …

Histone H1-mediated transfection: serum inhibition can be overcome by Ca2+ ions.

AbstractPurpose. One of the drawbacks of polycationic and cationic liposomalgene transfer is its sensitivity to serum. Gene therapy requires thetransfectant-DNA complex to be resistant to serum as well as blood.Since Ca2+ has proved to be an efficient cofactor of polycationic genetransfer, we decided to investigate its effects on transfection in thepresence of serum. Methods. We studied transgene expression of luciferase gene (pCMVLuc) on ECV 304 human endothelial cells using H1 histone andDOSPER as transfectants in the presence of 0-100% fetal calf serum. Results. H1-and DOSPER-mediated transfection was found to beinhibited by serum above the concentration of 10%. If 2 mM Ca2+ or2 mM Ca2+/0.1 mM chloroquine was included in the culture mediumwhich replace the transfection mixture and was left on the cells for24 hours postincubation, the inhibiting effect of even 100% serumwas overcome. Conclusions. A high serum level does not interfere with binding anduptake of H1- and DOSPER-DNA complexes, but inhibits subsequentsteps such as endosomal escape. Ca2+ in the form of nascent calciumphosphate microprecipitates and other lysosomolytical agents facilitateendosomal/lysosomal release by their fusigenic and membranolyticactivity.

Renal endothelin system in diabetes : Comparison of angiotensin-converting enzyme inhibition and endothelin-A antagonism

An activated renal endothelin (ET) system is implicated in the pathogenesis of renal fibrosis, as recently shown in ET-1 transgenic mice. Because progressive renal fibrosis is also a major finding in diabetic nephropathy, we analyzed the activity of the renal ET system in rats with streptozotocin-induced diabetes mellitus and the effect of blocking the ETA receptor, using the orally active ETA antagonist LU 135252. The effects of long-term treatment with LU 135252 were compared with those of an ACE inhibitor. Plasma and urinary ET-1 concentrations were measured. Progression of diabetic nephropathy was analyzed by measuring urinary albumin and protein excretion. Urinary ET-1 excretion was significantly elevated as early as 7 days after induction of diabetes and increased further. The daily urine volume was significantly correlated with urine ET-1 excretion. Treatment with LU 135252 significantly decreased the ET-1 excretion by more than 50%, whereas ACE inhibition resulted only in a mild decrease. Albumin excretion was significantly decreased after ACE inhibition, whereas ETA inhibition resulted in a nonsignificant decrease. Urinary ET and albumin excretion probably reflect independent mechanisms of renal damage in diabetes.

Hyperosmotic stress enhances cytokine production and decreases phagocytosis in vitro

IntroductionHyperglycemia is associated with negative outcomes in various settings of critical illness; infectious complications, especially, seem to be increased. On the other hand, intensive insulin therapy (IIT) has been shown to improve outcome in clinical trials. Whether normoglycemia itself or the application of insulin is responsible for the observed findings is unknown. We therefore tested the effect of glucose and insulin on various immune functions in vitro.MethodsHuman peripheral blood mononuclear cells (PBMCs) were incubated ex vivo with low doses of lipopolysaccharide (LPS). PBMCs were incubated with various osmotic agents, insulin, or a combination of both. Interleukin (IL)-6 and IL-1 cytokine response was measured by enzyme-linked immunosorbent assay. In addition, we investigated the effects of glucose on phagocytosis and oxidative burst in human granulocytes.ResultsIncreasing concentrations of both glucose and mannitol significantly enhanced LPS-induced cytokine production. Insulin alone did not alter cytokine production and had only a minor influence in combination with glucose. Phagocytosis and oxidative burst were significantly reduced with increasing concentrations of glucose and mannitol.ConclusionHyperglycemia may lead to inflammation by enhancing cytokine production via the direct effects of hyperosmotic stress. Impaired phagocytosis and oxidative burst under hyperglycemia may weaken defense mechanisms of the host. Our in vitro findings may help to explain the beneficial effects of IIT not only in diabetic but also in critically ill patients.

Potent Early Immune Response After Kidney Transplantation in Patients of the European Senior Transplant Program

Background. The increasing age of organ donors and the transplantation of older recipients have become clinical practice. Age-adapted immunosuppressive protocols considering these changes are currently not established. This study analyzed the age-dependent immune response after human kidney transplantation. Methods. One hundred renal allograft recipients were prospectively evaluated from 2004 to 2005. Patients older than 65 years of the European Senior Program receiving kidneys from donors older than 65 years were compared with recipients younger than 65 years receiving kidneys from donors younger than 65 years. Age-dependent modifications of the immune response were evaluated before transplantation and 7 days and 6 months after grafting by flow cytometry analysis of lymphocyte surface markers in peripheral blood. The cytokine pattern was determined by Cytometric Bead Array, T-cell alloreactivity by enzyme-linked immunospot analysis. Results. There were no differences between the groups regarding patient survival, graft survival, and function at 6 months after transplantation. Before transplantation, 7 days and 6 months thereafter recipients older than 65 years demonstrated significantly elevated numbers of memory T-cells while counts for naive T-cells were significantly reduced. Numbers of activated cytotoxic cells were elevated with increasing age before and 7 days after transplantation. T-cell alloreactivity was more pronounced in older recipients at all time points. Seven days after transplantation tumor necrosis factor-α (TNF-α) levels were significantly higher, whereas TNF-α and interleukin-10 (IL-10) concentrations were significantly reduced after 6 months in older recipients. Conclusions. Our data demonstrate an initially pronounced immune response in elderly recipients receiving grafts from elderly donors. This observation supports the concept of a donor and recipient age-adapted immunosuppression.

Accumulation of Crystal Deposits in Abdominal Organs Following Perfusion with Defrosted University of Wisconsin Solutions

Previous studies reported on both visible and invisible particles in University of Wisconsin (UW) solutions. Those particles originated from components of the bags. In recent clinical observations we noticed macroscopically visible, indissoluble particles in UW bags reaching subzero temperatures during transportation of organs and preservation solutions. In an experimental model we examined whether those particles could be detected following perfusion of abdominal organs with established perfusion solutions. UW‐, HTK‐ or physiological saline solutions reached − 3 ± 0.5 °C under conditions frequently applied during transportation. UW solutions demonstrated the accumulation of visible, indissoluble crystals and were subsequently used for the perfusion of abdominal organs in LEW rats. After perfusion with UW solutions stored at freezing temperatures, crystals were detected in all abdominal organs localized in and around vessels, bile ducts, glomeruli and in the interstitium of harvested livers, kidneys and pancreas. By spectroscopy, we were able to characterize crystals as adenosine. A 40‐μm pore‐size filter eliminated crystals from UW solutions. Crystals were absent in organs perfused with HTK‐ or saline solutions kept at subzero conditions. UW solutions can reach subzero temperatures under commonly used transportation conditions. Under these conditions, visible crystals accumulate and can be detected in abdominal organs of an experimental system.