Regulation of the expression of the Cl-/anion exchanger pendrin in mouse kidney by acid-base status

Regulation of the expression of the Cl-/anion exchanger pendrin in mouse kidney by acid-base status.BackgroundPendrin belongs to a superfamily of Cl-/anion exchangers and is expressed in the inner ear, the thyroid gland, and the kidney. In humans, mutations in pendrin cause Pendred syndrome characterized by sensorineural deafness and goiter. Recently pendrin has been localized to the apical side of non-type A intercalated cells of the cortical collecting duct, and reduced bicarbonate secretion was demonstrated in a pendrin knockout mouse model. To investigate a possible role of pendrin in modulating acid-base transport in the cortical collecting duct, we examined the regulation of expression of pendrin by acid-base status in mouse kidney.MethodsMice were treated orally either with an acid or bicarbonate load (0.28 mol/L NH4Cl or NaHCO3) or received a K+-deficient diet for one week. Immunohistochemistry and Western blotting was performed.ResultsAcid-loading caused a reduction in pendrin protein expression levels within one day and decreased expression to 23% of control levels after one week. Concomitantly, pendrin protein was shifted from the apical membrane to the cytosol, and the relative abundance of pendrin positive cells declined. Similarly, in chronic K+-depletion, known to elicit a metabolic alkalosis, pendrin protein levels decreased and pendrin expression was shifted to an intracellular pool with the relative number of pendrin positive cells reduced. In contrast, following oral bicarbonate loading pendrin was found exclusively in the apical membrane and the relative number of pendrin positive cells increased.ConclusionsThese results are in agreement with a potential role of pendrin in bicarbonate secretion and regulation of acid-base transport in the cortical collecting duct

Studies on the pathophysiology of cystinuria type A: electrophysiological characterisation of the hrBAT wildtype and its mutation T216M in xenopus oocytes

Die beiden Membranproteine rBAT und b0,+AT bilden als Heteromer einen funktionellen Aminosäuretransporter. Der hierbei induzierte Transport entspricht dem zuvor beschriebenen Aminosäuretransportsystem b0,+, bei dem dibasische und neutrale Aminosäuren sowie Cystin im Verhältnis 1:1 ausgetauscht werden. Der Einstrom von dibasischen Aminosäuren und Cystin in die Zelle wird bei diesem Austausch bevorzugt. Dadurch fungiert das Heteromer rBAT-b0,+AT als Teil des Mechanismus, der die Rückresorption dieser Substrate im proximalen Tubulus der Niere und deren Absorption im Dünndarm ermöglicht. In der vorliegenden Arbeit wurden zunächst die Eigenschaften des durch Expression des humanen rBAT in Xenopus laevis Oozyten induzierten Aminosäuretransports mittels der Zwei-Elektroden-Voltage-Clamp Methode elektrophysiologisch untersucht. Alle geprüften dibasischen Aminosäuren einschließlich Cystin, sowie die meisten neutralen Aminosäuren induzierten Ströme in rBAT exprimierenden Oozyten. Die unterschiedliche Polarität dieser Ströme und die Untersuchungen zur Spannungsabhängigkeit und intrazellulären Vorbeladung zeigten für den humanen rBAT Aminosäureaustausch im Verhältnis 1:1, wie zuvor an rBAT anderer Spezies beschrieben. Die kinetischen Studien zu Substrataffinitäten ergaben hohe extrazelluläre Affinitäten für die dibasischen Aminosäuren und Cystin sowie niedrigere Affinitäten für die neutralen Aminosäuren. Bis auf die hierbei beobachtete leichte Natrium-Abhängigkeit entsprechen diese Ergebnisse den für das Transportsystem b0,+ beschriebenen Eigenschaften und bestätigen die Induktion dieses Transportsystems durch rBAT. Die Funktion des aus rBAT und b0,+AT bestehenden heteromeren Aminosäuretransporters ist essentiell für die Rückresorption von Cystin aus dem Primärharn. Daher können Mutationen in einem der beiden Proteine zum Krankheitsbild der Cystinurie führen. Wegen der schlechten Löslichkeit des Cystins treten bei Cystinurie rezidivierende Cystinsteine auf, die zu Obstruktionen, Harnwegsinfekten und bei fortschreitendem Krankheitsverlauf zur Niereninsuffizienz mit Dialysepflichtigkeit führen können. Eine der häufigsten bei Cystinuriepatienten in rBAT gefundene Mutationen führt zum Ersatz von Threonin durch Methionin an Position 216. In funktionellen Studien dieser Mutation in Xenopus laevis Oozyten zeigten sich sowohl in der Austauschfunktion als auch bei den extrazellulären Affinitäten für die verschiedenen Aminosäuren und Cystin keine Unterschiede zu den Transporteigenschaften des Wildtyp-Proteins. Die Resultate der Experimente zur Abhängigkeit der Expression von hrBAT von der Menge der cRNA sowie von der Zeitdauer nach deren Injektion zeigten jedoch für die Mutation T216M im Vergleich zum Wildtyp eine gestörte Expression. Somit führt die Mutation T216M zu einem gestörten Trafficking des Proteins zur Zellmembran, wie zuvor auch für andere Mutationen gezeigt werden konnte. An der Zellmembran ist das defekte Protein wie der Wildtyp funktionell aktiv, kann diese aber nicht im gleichen Ausmaß erreichen. Da auch die leichte Untereinheit b0,+AT nur in Anwesenheit der schweren Untereinheit rBAT an die Plasmamembran gelangen kann, verbleibt diese auch vermehrt intrazellulär. Dadurch ist in vivo der Transport von Cystin und der dibasischen Aminosäuren und damit deren Rückresorption aus dem Primärharn beeinträchtigt und erklärt bei den diese Mutation tragenden Patienten das Auftreten des Krankheitsbildes der Cystinurie.The two membrane proteins rBAT and b0,+AT form a functional heteromeric amino acid transporter. The amino acid transport induced by this proteins corresponds to the previously described amino acid transport system b0,+, which exhibits 1:1 exchange of neutral and dibasic amino acids as well as cystine favouring the influx of dibasic amino acids and cystine into the cell. The heteromer rBAT-b0,+AT acts thereby as part of the mechanism which allows reabsorption of these substrates in the kidney proximal tubule and its absorption along the small intestine. In the present work the properties of the amino acid transport induced by the expression of the human rBAT in Xenopus laevis oocytes were examined using the two-electrode-voltage-clamp technique. All studied dibasic amino acids including cystine as well as most of the neutral amino acids induced currents in oocytes expressing rBAT. The opposed polarity of these currents and the studies on voltage dependence and intracellular preload demonstrated an amino acid exchange at the rate of 1:1 for human rBAT, as it was previously shown for rBAT of other species. The kinetic studies on substrate affinities resulted in high extracellular affinities for dibasic amino acids and cystine as well as lower affinities for neutral amino acids. Apart from the observed slight sodium-dependence these results correspond to the described properties of the transport system b0,+ and confirm the induction of this transport system by rBAT. The function of the heteromeric amino acid transporter formed by rBAT and b0,+AT is essential for the reabsorption of cystine out of the primary urine. Therefore, mutations in each of these two proteins can lead to cystinuria. Due to the low solubility of cystine, recurring cystine stones are formed and cause obstructions, urinary tract infections and later renal insufficiency with necessity to dialysis treatment. One of the most frequent mutations found in patients with cystinuria leads to the replacement of threonine by methionine at position 216. Functional studies of this mutation in Xenopus laevis oocytes revealed no differences compared to the wildtype protein regarding both the exchange function and the extracellular affinities for the studied amino acids. However, the experiments examining the dependence of hrBAT expression on the amount of cRNA or the time point after its injection resulted in a reduced expression compared to the wildtype protein. In summary, the mutation T216M leads to an impaired protein trafficking to the cell membrane as it was previously shown for other mutations. At the cell membrane, the defective protein is functionally active as the wildtype protein, but it is unable to reach the surface to the same extent. Hence, the light subunit b0,+AT which reaches the plasma membrane only in presence of the heavy subunit rBAT remains likewise mostly inside the cell. Therefore, the transport of cystine and the dibasic amino acids which is in vivo necessary for their reabsorption out of primary urine is impaired and explains the presence of cystinuria in patients carrying this mutation

Association of intraoperative transfusion of blood products with mortality in lung transplant recipients

BACKGROUND The impact of intraoperative transfusion on postoperative mortality in lung transplant recipients is still elusive. METHODS Univariate and multivariate analysis were performed to investigate the influence of red blood cells (RBCs) and fresh frozen plasma (FFP) on mortality in 134 consecutive lung transplants recipients from September 2003 until December 2008. RESULTS Intraoperative transfusion of RBCs and FFP was associated with a significant increase in mortality with odds ratios (ORs) of 1.10 (1.03 to 1.16, P = 0.02) and 1.09 (1.02 to 1.15, P = 0.03), respectively. For more than four intraoperatively transfused RBCs multivariate analysis showed a hazard ratio for mortality of 3.8 (1.40 to 10.31, P = 0.003). Furthermore, non-survivors showed a significant increase in renal replacement therapy (RRT) (36.6% versus 6.9%, P <0.0001), primary graft dysfunction (PGD) (39.3% versus 5.9%, P <0.0001), postoperative need of extracorporeal membrane oxygenation (ECMO) (26.9% versus 3.1%, P = 0.0019), sepsis (24.2% versus 4.0%, P = 0.0004), multiple organ dysfunction syndrome (MODS) (26.9% versus 3.1%, P <0.0001), infections (18.1% versus 0.9%, P = 0.0004), retransplantation (12.1% versus 6.9%, P = 0.039) and readmission to the ICU (33.3% versus 12.8%, P = 0.024). CONCLUSIONS Intraoperative transfusion is associated with a strong negative influence on outcome in lung transplant recipients

Extracorporeal membrane oxygenation: beneficial strategy for lung transplant recipients

The role of extracorporeal membrane oxygenation (ECMO) as a therapeutic strategy has been very well documented for over a decade now with consistently positive remarks. The aim of the present study was analyzing the outcome of ECMO application in our lung transplant program, especially the feasibility and safety of our ECMO approach. Therefore, we retrospectively analyzed the data of 15 patients recipients requiring ECMO support. We analyzed clinical data, complications, and survival of the lung-transplanted population that needed ECMO support at our institution from 2006-2009. During that period, 19 applications of ECMO were done on 15 adult patients with the following indications: primary graft dysfunction (10 patients), "bridge to transplantation" (five), pulmonary hypertension (three), and severe acute respiratory distress syndrome (one). At 28 days, the overall survival was 93% (14 of 15 patients) and 12 of these patients (80%) survived at least 6 months. Complications included acute renal insufficiency with temporary need of renal replacement therapy (53%), bleeding (33%), critical illness polyneuropathy (66%), and reversible thrombocytopenia (73%). Based on the evaluation of the patients in this analysis, ECMO seems to be a safe therapeutic approach in lung transplant recipients with severe respiratory failure directly after transplantation

Extracorporeal Membrane Oxygenation: Beneficial Strategy for Lung Transplant Recipients

The role of extracorporeal membrane oxygenation (ECMO) as a therapeutic strategy has been very well documented for over a decade now with consistently positive remarks. The aim of the present study was analyzing the outcome of ECMO application in our lung transplant program, especially the feasibility and safety of our ECMO approach. Therefore, we retrospectively analyzed the data of 15 patients recipients requiring ECMO support. We analyzed clinical data, complications, and survival of the lung-transplanted population that needed ECMO support at our institution from 2006–2009. During that period, 19 applications of ECMO were done on 15 adult patients with the following indications: primary graft dysfunction (10 patients), “bridge to transplantation” (five), pulmonary hypertension (three), and severe acute respiratory distress syndrome (one). At 28 days, the overall survival was 93% (14 of 15 patients) and 12 of these patients (80%) survived at least 6 months. Complications included acute renal insufficiency with temporary need of renal replacement therapy (53%), bleeding (33%), critical illness polyneuropathy (66%), and reversible thrombocytopenia (73%). Based on the evaluation of the patients in this analysis, ECMO seems to be a safe therapeutic approach in lung transplant recipients with severe respiratory failure directly after transplantation

ICG-liver test versus new biomarkers as prognostic markers for prolonged length of stay in critically ill patients - a prospective study of accuracy for prediction of length of stay in the ICU

BACKGROUND: Prognostic abilities of medical parameters, which are scoring systems, measurements and biomarkers, are important for stratifying critically ill patients. Indocyanine green plasma disappearance (ICG-PDR) is an established clinical tool for the assessment of liver perfusion and function. Copeptin, MR-proANP and pro-ADM are biomarkers whose prognostic value is still unclear. The goal of this prospective study was to evaluate ICG-PDR, copeptin, MR-proANP and pro-ADM to predict prolonged length of stay (pLOS) in the ICU. METHODS: This study was conducted as a prospective single center study including 110 consecutively admitted ICU patients. Primary endpoint was prolonged length of stay (pLOS) in the ICU, defined as more than three days of stay there. RESULTS: ROC analysis showed an AUC of 0.73 for ICG-PDR, 0.70 for SAPS II, 0.65 for MR-proANP, 0.64 for pro-ADM and 0.54 for copeptin for pLOS in the ICU. CONCLUSIONS: The prediction of pLOS in the ICU might be better by means of ICG-PDR than with the new biomarkers copeptin, MR-proANP or pro-ADM. Nevertheless, there is more need for research to evaluate whether ICG-PDR is an overall prognostic marker for pLOS. TRIAL REGISTRATION: (ClinicalTrials.gov number, NCT01126554)

Influence on ICU course, outcome and costs for lung transplantation after implementation of the new Swiss transplantation law

BACKGROUND The Swiss organ allocation system for donor lungs was implemented on 1 July 2007. The effects of this implementation on patient selection, intensive care unit course, outcomes and intensive care costs are unknown. METHODS The first 37 consecutive lung transplant recipients following the implementation of the new act were compared with the previous 42 lung transplant recipients. RESULTS Following implementation of the new law, baseline characteristics and cumulative one-year patient survival were comparable in both groups (88.1% vs 83.8%, P = 0.58). The costs for each case increased by 35,000 euros after adoption of the new law. Stratifying patients after implementation of the law according to urgency status shows that urgent patients required longer mechanical ventilation (P = 0.04), a longer ICU stay (P = 0.045) and a longer hospital stay (P = 0.04) and ICU costs (median 64,050 euros) were higher compared to regular patients. CONCLUSION The new transplantation law has increased ICU costs with the implementation of the Swiss organ allocation system. Patients listed as 'urgent' contribute significantly to the increase in ICU costs

Cystinuria-specific rBAT(R365W) mutation reveals two translocation pathways in the amino acid transporter rBAT-b 0,+ AT

Apical reabsorption of dibasic amino acids and cystine in kidney is mediated by the heteromeric amino acid antiporter rBAT/b0,+AT (system b0,+). Mutations in rBAT cause cystinuria type A, whereas mutations in b0,+AT cause cystinuria type B. b0,+AT is th