[Conversion of sound into auditory nerve action potentials]. / Codierung von Schallsignalen in Aktionspotenziale des auditorischen Nervs.

Outer hair cells play a major role in the hearing process: they amplify the motion of the basilar membrane up to a 1000-fold and at the same time sharpen the excitation patterns. These patterns are converted by inner hair cells into action potentials of the auditory nerve. Outer hair cells are delicate structures and easily damaged, e. g., by overexposure to noise. Hearing aids can amplify the amplitude of the excitation patterns, but they cannot restore their degraded frequency selectivity. Noise overexposure also leads to delayed degeneration of auditory nerve fibers, particularly those with low a spontaneous rate, which are important for the coding of sound in noise. However, this loss cannot be diagnosed by pure-tone audiometry.

Activation of natural killer cells by hepatitis C virus particles in vitro.

Little is known about the ability of hepatitis C virus (HCV) to alter early innate immune responses in infected patients. Previous studies have shown that natural killer (NK) cells are functionally impaired after interaction of recombinant HCV glycoprotein E2 with the co-stimulatory CD81 molecule in vitro; however, the functional consequences of a prolonged contact of NK cells with HCV particles have remained unclear. We have examined the phenotypes of purified, interleukin-2-activated NK cells from healthy donors and HCV genotype 1b patients after culture for 5 days with HCV pseudoparticles (HCVpp) and serum samples containing HCV genotype 1b. NK cells from healthy donors and chronic HCV patients were found to up-regulate receptors associated with activation (NKp46, NKp44, NKp30, NKG2D), while NK receptors from the killer cell immunoglobulin-like receptor family (KIR/CD158), predominantly having an inhibitory function, were significantly down-modulated after culture in the presence of HCV particles compared with control cultures of NK cells. HCV-infected sera and HCVpp elicited significantly higher secretion of the NK effector lymphokines interferon-γ and tumour necrosis factor-α. Furthermore, HCV stimulated the cytotoxic potential of NK cells from normal donors and patients. The enhanced activation of NK cells after prolonged culture with HCVpp or HCV-containing sera for 5 days suggests that these innate effector cells may play an important role in viral control during early phases of HCV infection.

Fatal varicella in an immunocompromised adult associated with a European genotype E2 variant of varicella zoster virus.

Varicella zoster virus (VZV) seronegative patients under immunosuppressive therapy are at risk for severe life-threatening varicella. A 25-year-old male patient presented with rash and hepatitis. He had been known to suffer from Crohn's disease and received immunosuppressive treatment with azathioprine. The patient showed dyspnoea, and presented with a generalized rash with vesicular lesions typical for herpesvirus infections. He was started immediately on acyclovir therapy. Varicella infection was determined in this VZV seronegative patient in rash vesicles, blood and tracheal secretions and a high VZV viral load was detected in these specimens. The causative agent was genotyped by sequencing of several genes as a variant of the European genotype E2 containing several unique single nucleotide polymorphisms. Despite all measures, there was progressive septic shock, and the patient died due to multi-organ failure. Immunocompromised adults without varicella history are at high risk to develop life-threatening complications of varicella. Antiviral therapy with acyclovir can only be successful when administered as early as possible on suspicion of varicella infection in this group of patients. The most effective method to prevent severe varicella in immunocompromised patients is active immunization prior to immunosuppressive therapy.

[Model for end-stage liver disease. New basis of allocation for liver transplantations]. / Model for end-stage liver disease. Neue Grundlage der Allokation für die Lebertransplantation.

In December 2006 the allocation of livers from deceased donors in Germany and several other Eurotransplant countries was reset. The previous allocation system relied on CTP score to assess the need of transplantation, but it also assigned to waiting time a prominent role in prioritization. That system was replaced by the primarily urgency-oriented model of end-stage liver disease (MELD) allocation system. First experience with this classification in the U.S.A. shows that MELD scores are able to identify the urgency of liver transplantation correctly in most types of liver disease. Due to the MELD-based allocation, the growing waiting time and waiting-list mortality could be counteracted. At the same time it became evident however that MELD scores do not reflect mortality on the waiting list or thus the urgency for all types of liver diseases. Therefore the new allocation system introduced in the Eurotransplant countries contains standardized and flexible exceptions for these diseases. In addition the new allocation rules were created as a learning system. Repeated "fine tuning" of the allocation process based on continuous monitoring of daily allocation practice and clinical studies aim at just and effective distribution of the precious and limited supply of donor organs.

Extended donor criteria have no negative impact on early outcome after liver transplantation: a single-center multivariate analysis.

The organ shortage has driven many transplant centers to accept extended donor criteria and to modify graft allocation policies. This study was designed to analyze the impact of applying extended donor criteria (EDC) in orthotopic liver transplantation (OLT). Between December 2001 and December 2004, we performed 165 primary cadaveric whole OLTs. Up to three EDC, that is, ventilation >7 days; aminotransferases (ALT or AST) >3 x normal; bilirubin >3 mg/dL; anti-HBc or HBs Ag positivity; donor age >65 years; liver steatosis >40%; donor body mass index >30; cold ischemia time >14 hours; peak serum Na(+) >165 mmol/L; history of extrahepatic malignancy; or previous drug abuse were present in 55% of all grafts. Both univariate and multivariate analysis revealed that EDC status had no effect on graft or patient survival, the necessity for retransplantation, the length of intensive care/intermediate care unit stay, mechanical ventilation, complications, or posttransplant laboratory findings. Recipient age of >/=55 years was the only independent prognostic factor for survival, regardless of EDC. These findings suggested that the use of grafts from EDC donors are safe and expand the donor pool.

Severe rhabdomyolysis and renal failure triggered by a sauna visit in sickle cell trait: a case report.

Sickle cell trait (SCT) is an usually asymptomatic hemoglobinopathy. Cases of sudden excertional deaths in individuals with SCT have been described. We here report an exceptional case of excessive rhabdomyolysis and acute renal failure triggered by a sauna visit in a 29 year-old African American with SCT.

Bioreactor for a larger scale hepatocyte in vitro perfusion.

A bioreactor construction for hepatocytes and liver sinusoidal endothelial cells is described. The reactor is based on capillaries for hepatocyte immobilization. Four discrete capillary membrane systems, each serving different purposes, are woven to create a three-dimensional framework for decentralized cell perfusion with low metabolite gradients and decentralized oxygenation and CO2 removal. The biochemical performance of reactors initially seeded with 2.5 x 10(9) hepatocytes were evaluated over 3 weeks. On day 21, pig albumin synthesis was 4.7 mg/day, lidocaine metabolism was 813.7 +/- 23 micrograms/hr, galactose elimination was 210.1 +/- 3 mg/hr, and midazolam metabolism was 37.1 +/- 2 micrograms/hr. The specific construction of the reactor enables scale-up to hybrid liver support systems as extracorporeal bridging devices for liver transplantation.

Is a clinical application of hybrid liver support systems limited by an initial disorder in cellular amino acid and alpha-keto acid metabolism, rather than by later gradual loss of primary hepatocyte function?

The in-vitro amino acid (AA) and alpha-keto acid (KA) metabolism of bioreactors initially seeded with 2.5 x 10(9) pig hepatocytes was investigated with a perfusion technique. Considerable changes in the culture medium concentrations of AA and KA were measured during the first days in culture. This is indicative of dynamic cellular metabolism in the initial phase. While the concentration of pyruvate decreased starting on the first day, alpha-ketoglutarate, alpha-ketoisocaproate, alpha-ketoisovalerate, and alpha-keto-beta-methyl-n-valerate were synthesized. The long term use of hepatocyte cultures in bioreactors and thus a desirable clinical hybrid liver support therapy appears to be possible since the hepatocytes switched, after 15 days in culture, to steady-state conditions with a stable amino acid turnover featuring general AA uptake accompanied by KA release. The release of branched chain KA, in particular that of alpha-ketoisocaproate, reflects an effective transamination activity in the bioreactor system. Primary pig hepatocytes cultivated in hybrid liver support systems for therapy of acute liver failure or as devices for bridging to liver transplantation can sustain amino acid metabolism for at least 30 days in vitro. However, an initial disorder following the cell isolation that is demonstrated may limit immediate utilization of the systems prior to the reorganisation of the cells to tissue-like structures in bioreactors.

Total chemical synthesis of the 3' untranslated region of the hepatitis C virus with long oligodeoxynucleotides.

Hepatitis C Virus (HCV) is the major causative agent of chronic hepatitis. Since it has been difficult to obtain full-length cDNA clones of HCV including the 3' untranslated region (UTR) that give rise to replication competent virus, we generated the 3'UTR by a modified protocol of total chemical synthesis (TCS) with overlap-extension-PCR using four long oligodeoxynucleotides. A synthetic cDNA fragment of about 340 nucleotides (nt) in length was generated, subcloned and sequenced. This approach represents a rapid and easy alternative to RT-PCR from infectious serum and may be a highly valuable method to generate partial cDNA clones of HCV and other viruses including defined variants.

Amino acid metabolism by hepatocytes in a hybrid liver support bioreactor.

The amino acid patterns of medium perfusate in a liver cell bioreactor developed for a hybrid liver support system have been measured. There were considerable changes in the concentrations of glutamic acid, glutamine, alanine, arginine, ornithine and branched chain amino acids during the first 10 days which is indicative of dynamic cellular metabolism. From day 15, steady state conditions of nitrogen metabolism are reflected by stable amino acid turnover. Monitoring of urea, K+, and P-450 activity suggests that hepatocytes have switched to a stable protein synthesis with a general amino acid uptake and keto acid release following cell volume increase.

Alpha-keto acid metabolism by hepatocytes cultured in a hybrid liver support bioreactor.

Isolated pig liver cells cultured using a perfusion technique were analyzed over 39 days to test their ability to change the perfusate alpha-keto acid profile. While the pyruvate concentration in the culture medium decreased as of the first day, the alpha-ketoglutarate (KG), alpha-ketoisocaproate (KIC), alpha-ketoisovalerate (KIV) and alpha-keto-beta-methyl-n-valerate (KMV) were synthesized immediately and released by the liver cells. The metabolic capacity of the cell culture system increased up to day 10, decreased during the following 5 days and reached a steady state beyond day 15, which was maintained for at least 30 days. The branched chain alpha-keto acid release, in particular alpha-ketoisocaproate, reflects an effective transamination capacity of the newly developed culture system and shows an intact protein biosynthesis for at least 30 days in vitro.

An early increase in gamma glutamyltranspeptidase and low aspartate aminotransferase peak values are associated with superior outcomes after orthotopic liver transplantation.

BACKGROUND: Prediction of prognosis after liver transplantation (OLT) remains difficult. The present study determines if standard laboratory parameters measured within the first week after OLT correlate with outcome. PATIENTS AND METHODS: Laboratory parameters measured within the first weak after OLT of 328 patients were grouped either graft loss or death within 90 days after (group 1: graft loss; group 2: death; group 3: neither graft loss nor death within 90 days). RESULTS: Peak AST and ALT were significantly lower in group 3 (1867 and 1252 U/L) than in group 1 (4474 and 2077 U/L) or 2 (3121 and 1865 U/L). Bilirubin was significantly lower and gamma-GT significantly higher in group 3 compared to groups 1 and 2. In multivariate analysis, high AST peaks were independently associated with death or graft loss within 90 days. An increase in gamma-GT and low bilirubin early after transplantation were found to be independently associated with superior outcome. DISCUSSION: Unexpectedly, a disproportionate rise in gamma-GT was associated with graft and patient survival of more than 90 days. This might be explained by regeneration phenomena in the liver indicative of a well functioning graft.

[Hemoptysis and acute renal failure in a 29-year-old patient with chronic hepatitis C infection]. / Hämoptysen und akutes Nierenversagen bei einem 29-jährigen Patienten mit chronischer Hepatitis C.

A 29-year-old male patient with chronic hepatitis C infection and interferon alpha therapy in his medical history was admitted to the hospital because of the clinical manifestation of a pulmonary renal syndrome. High titers of proteinase-3-ANCA were detected, while an infectious agent was ruled out. After diagnosis of Wegener's granulomatosis the patient received prednisolone and cyclophosphamide pulse therapy and remission developed rapidly. Chronic hepatitis C infection as well as interferon therapy are frequently associated with autoimmune disorders. We assume that the interferon therapy itself has triggered autoimmune processes resulting in Wegener's granulomatosis in our patient. Thus we recommend to search specifically for autoimmune disorders in the past medical history and if necessary to consider a screening for autoantibodies before starting an interferon therapy. An autoimmune disease should also be taken into account if new symptoms develop under an ongoing interferon alpha therapy.

Development of a heterologous, multigenotype vaccine against hepatitis C virus infection.

BACKGROUND: Unquestionably viral diversity and genetic heterogeneity in hepatitis C virus (HCV) infection and other viral diseases play an essential role in viral immune escape and the development of chronicity. Despite this knowledge most vaccine approaches against HCV have excluded this important issue. Moreover the feasibility of developing an effective HCV vaccine has been questioned, mainly because prophylactic immunity against HCV cannot be achieved in chimpanzees by either vaccination or previous HCV infection, and reinfection in men has been reported, most likely due to genetic shift and immune escape. To analyse and characterize a new technique of a 'multigenotype'- and/or 'library'-vaccine, we established an envelope 1 (E1) plasmid vaccine against HCV and characterized humoral and cellular immune responses after vaccination in a mouse model. MATERIAL AND METHODS: Normally genetic information of one or two target proteins is cloned into a DNA-vaccine. In our approach we cloned a defined number of different genotypes and subtypes (defined vaccine, DV) or the genetic information from 20 patients (undefined) into a plasmid (library vaccine, LV). RESULTS: As expected, immunized animals showed both stronger humoral (ELISA) and cellular (T-cell proliferation, ELISPOT) immune responses against genotype 1, since the stimulating antigen was genotype 1 derived. However, not all genotype 1 immunized animals recognized this viral antigen leading to the assumption that some epitopes lost their immunogenicity through a change in the amino acid sequence. Interestingly, some of the genotype 4 and 5 immunized mice sera were able to react against E1 protein. CONCLUSION: Most of the assays showed immune reactivity against the DV or LV vaccine demonstrating the cross-reactive potential of such a vaccination approach. This cloning and immunization strategy based on the viral heterogeneity of the virus has in our view major implications for HCV, a virus with a broad viral genetic diversity, and may become in the future in the context of DNA- or viral-based vaccination strategies a possibility to overcome viral immune escape both in the prophylactic or therapeutic setting.

Effect of Flt3 ligand gene transfer in experimental pancreatic cancer.

BACKGROUND: Fms-like tyrosine kinase 3 receptor (Flt3) is an important receptor expressed on the cell membrane of immature antigen-presenting cells. The binding of Flt3 to its ligand (FL) activates the proliferation of dendritic cells (DCs). This mechanism is currently being evaluated in the therapy of malignant tumors. The aim of the present study was to study the effect of FL gene transfer on the immune response and tumor growth in experimental pancreatic cancer. MATERIALS AND METHODS: The rat FL was sequenced and cloned from total mRNA extract of the spleen. Transfection efficiency of subcutaneously growing rat duct-like pancreatic cancer (DSL6A) with DOTAP-/cholesterol-based liposomes was tested using a pcDNA3.1-lacZ construct. Flt3 ligand production of in vitro transfected tumor cells and in vivo transfected tumors was measured by enzyme-linked immunosorbent assay. Tumor induction was achieved in Lewis rats by a subcutaneous inoculation of syngeneic pancreatic tumor cells (DSL6A). The animals were allocated into three groups: control, mock treatment, and treatment with FL plasmid. The plasmid was injected intratumorally three times per week for 2 weeks. The total observation time was 6 weeks. RESULTS: The tumor volume was significantly lower in the FL-transfected group during the first 3 weeks. The number of responders was significantly higher in the FL group compared with control and mock treatment. The number of CD80+ DCs in the spleen was significantly higher after FL gene transfer. The responders showed a significantly higher number of splenic natural killer (NK) cells. There were no differences of infiltrating lymphocytes, proliferation, and tumor blood vessels between the groups. CONCLUSION: Intratumoral gene transfer of FL in rats activated proliferation of DCs and NK cells, which causes a moderate reduction of tumor growth. This improvement of local tumor control during the first weeks could be explained by an improved antigen presentation.

Thymoglobulin and ischemia reperfusion injury in kidney and liver transplantation.

Since the beginning of organ transplantation, graft preservation has been one of the most important concerns. Ischemia reperfusion injury (IRI), which plays an important role in the quality and function of the graft, is a major cause for increased length of hospitalization and decreased long term graft survival. Among numerous attempts which have been made to minimize graft damage associated with IRI, the use of Thymoglobulin (TG) seems to offer potential benefits. TG is a polyclonal antibody which blocks multiple antigens related to IRI, in addition to its better known T cell depleting effects. This review will focus on the use of TG in preventing IRI in kidney transplantation (KTx) and liver transplantation (LTx). Different studies in experimental and clinical transplantation have shown that TG protects renal and liver grafts from IRI. Improvement in early graft function and decreased delayed graft function (DGF) rates are some of the clinical benefits of TG. Additionally, it is used in patients with hepatorenal syndrome to support the recovery of kidney function after LTx, by allowing reduced exposure to nephrotoxic calcineurin inhibitors as well as improving liver graft function by minimizing IRI. TG can reduce acute rejection rates in kidney and liver transplant recipients, decrease the length of hospital stay, and hence reduce transplantation costs. TG can play an important role in expanding the donor pool in both KTx and LTx by improving long-term graft and patient survival rates which increases the possibility of using marginal donors. Although controversial, the development of post-transplant lymphoproliferative disorder is a potential side effect of TG. No single optimal immunosuppressive regimen has given consistent results in decreasing the graft damage following IRI; however, TG usage in KTx and LTx appears to have some benefits in reducing IRI.

Biliary complications after liver transplantation.

Biliary complications remain a substantial cause of morbidity following liver transplantation. They have been reported to occur in a rate of 10-15% of full-size transplantations and may be higher in living donor, split or reduced size liver transplantations. The most common biliary complications following liver transplantations are leaks and strictures. In both, the incidence varies with respect to type of graft and donor as well as the type of biliary anastomosis. The management of the biliary complications requires a multidisciplinary approach and has changed over the past decade, favoring endoscopic and radiological techniques. Surgical revision including retransplantation is reserved for patients in whom endoscopic and interventional modalities are unsuccessful.

The role and value of sirolimus administration in kidney and liver transplantation.

Enormous advancements in visceral transplantation have led to significant improvements in the quality of life of patients. However, despite these developments, the average graft half-life after transplantation has remained almost unchanged and chronic rejection is still considered a major problem. In this regard, more concerns have shifted to factors influencing long-term graft survival, patient survival, and quality of life. To achieve this goal, detrimental effects of immunosuppressive (IS) agents, which have deleterious influence on the quality of life and/or patient survival, should be reduced. In the course of recent years, the transplant community has worked on reducing these side effects by developing new ISs, employing new combination regimens, or finding and adjusting optimal dosages and blood level concentrations. Among the IS agents, the antifungal, antitumoral and IS activity of mammalian target of rapamycin (mTOR) inhibitors without nephrotoxicity, have received special attention regarding this new class of IS. Sirolimus (SRL), as the first member of mTOR inhibitors, has been utilized in many clinical trials with respect to its benefit-risk assessment. In our review, the clinical evolution of SRL, as well as the evidence-based clinical benefits of SRL in kidney and liver transplantation (KTx, LTx), are summarized. Various studies of SRL in KTx and LTx have shown that combination therapy with SRL will enrich the variety of IS modalities. It also can be regarded as a safe base therapy to which other necessary drugs can be added. In addition to the enhanced acute rejection prophylaxis, and in contrast to the calcineurin inhibitors (CNI) and steroids, this drug solely does not have common side effects such as nephrotoxicity, neurotoxicity, diabetes mellitus and hypertension. Moreover, this agent might diminish vasculopathic processes that mediate chronic allograft nephropathy (CAN). Therefore, by reducing the likelihood of CAN it can decrease the rate of long-term organ failure. One possibly desirable characteristic of SRL is its antiproliferative effect, which could provoke antitumoral or antiatherogenic activity following transplantation. Despite all promising impacts of SRL in organ transplantation, there are some concerns regarding the adverse effects of this drug, for instance dyslipidemia, pneumonitis and wound healing problems. However, the majority of these side effects can be reduced or ceased by careful dose adjustments and correct timing of use. In conclusion, after a decade of both in vivo and in vitro studies on SRL, it can be advocated that SRL is a promising, potent and effective IS agent as it reduces the rate of acute rejection episodes in de novo transplants. It could improve the quality of life, graft and patient survival rate, and achieve excellent outcomes with few adverse effects when wisely used in combination with other immunosuppressants.

Wound complications following kidney and liver transplantation.

Advances in surgical techniques and immunosuppression (IS) have led to an appreciable reduction in postoperative complications following transplantation. However, wound complications as probably the most common type of post-transplantation surgical complication can still limit these improved outcomes and result in prolonged hospitalization, hospital readmission, and reoperation, consequently increasing overall transplant cost. Our aim was to review the literature to delineate the evidence-based risk factors for wound complications following kidney and liver transplantation (KTx, LTx), and to present the preventive and therapeutic modalities for this bothersome morbidity. Generally, wound complications are categorized as superficial and deep wound dehiscences, perigraft fluid collections and seroma, superficial and deep wound infections, cellulitis, lymphocele and wound drainage. The results of several studies showed that the most important risk factors for wound complications are IS and obesity. Additionally, there are surgical and/or technical factors, including type of incision, reoperation, and surgeon's expertise, as well as comorbidities such as advanced age, diabetes mellitus, malnutrition, and uremia. Preventive management of wound complications necessitates defining their etiological factors so that their detrimental effects on healing processes can be addressed and reduced. IS modalities and agents, especially sirolimus (SRL), and steroids (ST) should be adjusted according to the patient's co-existing risk factors. SRL should be administered three months after transplantation and ST should be tapered as soon as possible. A body mass index (BMI) lower than 30 kg/m2 is advisable for inclusion in a transplantation program, but higher BMIs do not exclude recipients. Surgical risk factors can be prevented by applying precise surgical techniques. Therapeutic modalities must focus on the most efficient and cost-effective medications and/or interventions to facilitate and improve wound healing.