Barbara Armann

EndothelinA Receptor Blockade Reduces Ischemia/Reperfusion Injury in Pig Pancreas Transplantation

Objective The effect of prophylactic administration of a selective endothelinA receptor antagonist (ETA-RA) on ischemia/reperfusion injury in an experimental model of graft pancreatitis after pancreas transplantation was evaluated. Summary Background Data It is well established that endothelin-1 (ET-1), a powerful vasoconstrictor, plays an important role in the development of pancreatitis. Recent studies have shown a beneficial effect of endothelin receptor antagonists in the therapy for experimental pancreatitis. Methods Relevant ischemia/reperfusion injury was induced in pig pancreas transplants after 6 hours hypothermic preservation in University of Wisconsin solution. The recipients were randomized into 2 groups: control pigs received isotonic saline and the treated group received the selective ETA-RA BSF 208075 at the beginning of reperfusion. On postoperative days 2 and 5, animals were relaparotomized to obtain tissue specimens. Blood monitoring included lipase, amylase, C-reactive protein, trypsinogen-activation peptide, thiobarbituric acid-reacting substances, and ET-1. Partial oxygen tension (ptiO2) was measured by a Clarke-type electrode and blood flow by laser doppler. A semiquantitative score index was used for assessment of histologic injury and for immunohistochemical analysis of ET-1 and ETA receptor expression. Tissue mRNA levels of prepro ET-1, ETA receptor, pro-interleukin (IL)-6, and pro-IL-1&bgr; were quantified using TaqMan real-time reverse transcription-polymerase chain reaction (RT-PCR). Results Prophylactic treatment with ETA-RA significantly reduced the severity of graft pancreatitis evidenced by C-reactive protein. The finding of transient capillary perfusion at the beginning of reperfusion supports the application of the ETA-RA during this period. The dramatic increase of plasma ET-1 in the therapy group is a clear evidence of effective receptor blockade. Mean trypsinogen-activation peptide levels from the portal venous effluent, but not mean systemic plasma TAP values were significantly lower in the treated group. Analysis of ptiO2 and blood flow revealed a significant improvement of capillary perfusion and blood flow in the treated group and was associated with relevant reduction of tissue injury. Intrapancreatic ET-1 and IL-6 mRNA expression and ET-1 protein levels were significantly lower in the therapy group as compared with the control group. In contrast, ETA mRNA showed a marked up-regulation by ETA receptor blockade. Conclusion Application of a ETA-RA reduces ischemia/reperfusion induced graft pancreatitis in a pig transplantation model by improving microcirculation and reducing tissue injury.

Endothelin a receptor blockade reduces hepatic ischemia/reperfusion injury after warm ischemia in a pig model

It is well established that endothelin-1 (ET-1) is a very potent mediator of vasoconstriction that leads to microcirculatory disturbances. The aim of the study was to evaluate the effect of a selective endothelin A receptor antagonist on severe ischemia/reperfusion injury in a pig model. Fourteen pigs were subjected to 120 minutes of complete vascular exclusion of the liver with a passive bypass. The animals were randomized into two groups: a control group, which was given isotonic saline solution, and a therapy group, which received the selective endothelin A receptor antagonist BSF 208075 at the beginning of reperfusion. On postoperative days 4 and 7, animals were relaparotomized to obtain tissue specimens. Blood monitoring included aspartate aminotransferase (AST), alanine aminotransferase (ALT), glutamate dehydrogenase (GLDH), alkaline phosphatase, and ET-1. Partial oxygen tension (ptiO2) was measured by a Clarke-type electrode and blood flow by laser Doppler. A semiquantitative scoring index was used for assessment of histologic injury and for immunohistochemical analysis of ET-1. Treatment with the endothelin A receptor antagonist significantly reduced the severity of the ischemia/reperfusion injury, as evidenced by lower levels of AST, ALT, and GLDH. The dramatic increase in plasma ET-1 in the therapy group is clear evidence of effective receptor blockade. Analysis of PtiO2 and blood flow revealed a significant improvement in capillary perfusion and blood flow in the treated group and was associated with relevant reduction of tissue injury. In summary, in the control group we observed serious microcirculatory disturbances and severe histologic damage in the liver after reperfusion. Treatment with a selective endothelin A receptor antagonist attenuated the ischemia/reperfusion injury in a porcine model of severe ischemia/reperfusion, as demonstrated by improved microcirculation, a reduction in histologic damage, and an decrease in liver enzymes.

Improvement of postischemic hepatic microcirculation after EndothelinA receptor blockade: Endothelin antagonism influences platelet-endothelium interactions

Endothelin (ET) contributes to disturbances of hepatic microcirculation after ischemia/reperfusion (I/ R) by causing vasoconstriction and enhancing leukocyte- and platelet-endothelium interactions. The aim of this study was to investigate a possible protective role of a selective endothelinA receptor antagonist (ETA-RA) in this setting. In a rat model, warm ischemia of the left lateral liver lobe was induced for 90 minutes under intraperitoneal anesthesia with xylazine and ketamine. Groups of rats consisted of shamoperated (SO, n = 14), untreated ischemia (n = 14), and treatment with BSF208075 (5 mg/kg body weight IV, n = 14). The effect of the ETA-RA on I/R was assessed by in vivo microscopy 20 to 90 minutes after reperfusion; by measurement of local tissue PO2, serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and glutathione S-transferase α levels, and by histologic investigation. In the untreated group, sinusoidal constriction to 69.4 ± 6.7% of diameters of SO rats was observed, leading to a significant decrease in perfusion rate (74.3 ± 2.1% of SO) and liver tissue PO2 (43.5 ± 3.2% of SO) (P < 0.05). In addition, we found an increased percentage of stagnant leukocytes (142.9 ± 11.9%) and platelets (450.1 ± 62.3%) in sinusoids and in postsinusoidal venules (P < 0.05). Hepatocellular damage (ASTandALTincrease to1330 ± 157U/Land750 ± 125U/Lrespectively; previously, 27.1 ± 3.5 U/L and 28.5 ± 3.6 U/L) was detected 6 hours after reperfusion (P < 0.05). Administration of the ETA-RA before reperfusion significantly reduced I/R injury. Sinusoidal diameters were maintained (108.5 ± 6.6%), and perfusion rate (93.1 ± 1.8%) and tissue PO2 (95.3 ± 5.7%) were significantly increased (P<0.05). According to reduced leukocyte-endothelium interactions after therapy, both platelet rolling and adhesion were significantly reduced (P < 0.05). The number of stagnant platelets in sinusoids was 199.5 ± 12.3% of 50 (P < 0.05). After treatment, hepatocellular damage was decreased (AST and ALT levels after 6 hours of reperfusion: 513 ± 106 U/L and 309 ± 84 U/L, respectively; P < 0.05), and histologic changes were reduced in the long term. Our results provide evidence that the new therapeutic approach with an ETA-RA is effective in reducing hepatic I/R injury. In addition to reduced leukocyte-endothelium interactions, the number of stagnant and rolling platelets in sinusoids and venules was significantly reduced. The reduction in microcirculatory damages is responsible for better organ outcome.

Effects of ETA Receptor Antagonism on Proinflammatory Gene Expression and Microcirculation Following Hepatic Ischemia/Reperfusion

Background: The objective of this study was to investigate the effect of a specific endothelinA receptor antagonist (ETA‐RA) on mRNA expression of genes encoding vasoactive mediators and proinflammatory cytokines and on the microhemodynamics (assessed by measurement of laser Doppler flow and tissue blood gases) following complete vascular exclusion of the porcine liver.

Paratrend sensor as a novel method for continuous monitoring of hepatic microperfusion

DESPITE THE REMARKABLE progress in liver tranplantation, 7% of graft losses occur due to vascular complications, especially of the hepatic artery. The continuous monitoring of hepatic microcirculation following liver transplantation represents a technique for early detection of vascular complications. Multiple methods have been employed to measure liver blood flow in vivo. However, most of the methods, like indocyanine green clearance, indicator dilution, or fractionation techniques, are limited by an indirect approach and the lack of repetitive sampling. Electromagnetic and Doppler flow probes and laser flowmetry allow direct quantification of whole organ blood flow but have the drawback of short-term intraoperative application, especially in the clinical setting. All of the methods mentioned above preclude continuous monitoring of hepatic perfusion in patients over several days after liver transplantation. In contrast, assessment of hepatic pCO2, pO2, and pH with the Paratrend has the potential to provide a continuous real-time quantitative monitoring of liver microcirculation with little invasiveness. Measurement of hepatic tissue oxygenation has been shown to correlate significantly with microcirculatory impairment and liver dysfunction induced by vascular problems. Additional measurement of pCO2 and pH permits assessment of tissueacid base and the metabolic state in the liver. Up to now the Paratrend technique was not inserted in parenchymal organs except for brain/spinal cord monitoring and a hemorrhagic shock study in the liver. The aim of the present study was to evaluate the Paratend technique for continuous quantitation of hepatic microcirculation in a preclinical animal model of hepatic artery and portal vein occlusion. The Paratrend technique was compared with the established methods of laser Doppler flowmetry and pO2 measurement by the Licox device.

Experiences with arachnoid cysts in children

Background Arachnoid cysts are relatively rare. True cysts are congenital. Secondary cysts may result from postinflammatory accumulation of CSF in the subarachnoid space. This examination focussed exclusively on primary cysts. There is a controversy regarding the role and the type of surgery indicated in its treatment. The distribution, clinical features and treatment modalities in different types of arachnoid cysts in our patients are analysed in this study.

Changes of vasoregulatory gene expression following hepatic ischemia/reperfusion and treatment with endothelin-A receptor blockade.

The objective of this study was to investigate the effect of a specific endothelin-A receptor antagonist on mRNA expression of genes encoding vasoactive mediators and proinflammatory cytokines following complete vascular exclusion of the porcine liver. Fourteen adult German Landrace pigs were subjected to 120 minutes of warm hepatic ischemia by total vascular exclusion. The animals were divided into two groups: the control group received saline solution and the therapy group was given the selective endothelin-A receptor antagonist BSF 208075. Liver tissue samples were collected 1 hour after reperfusion and mRNA expression for preproendothelin-1, prointerleukin-1β, prointerleukin- 6, pro-tumor necrosis factor-α and endothelial nitric oxide synthase was analyzed quantitatively using the TaqMan system. Additionally, immunohistochemical analysis using a semiquantitative score for endothelin-1 and endothelin-A receptor was performed. One hour after reperfusion, quantitative reverse transcriptase-polymerase chain reaction revealed significantly lower expression of preproendothelin-1, pro-tumor necrosis factor-α, and prointerleukin-6 in the therapy group compared to controls. Immunohistochemical analysis demonstrated significantly reduced endothelin-1 immunostaining after therapy. Treatment with the selective endothelin-A receptor antagonist exerts a protective effect on the microcirculation after liver ischemia and reperfusion. We were able to show that the endothelin-A receptor antagonist not only has effects on the expression of vasoactive genes, it also decreases gene expression of proinflammatory cytokines such as tumor necrosis factor-α and interleukin-6.

Einfluß eines selektiven Endothelin-A-Rezeptor-Antagonisten auf den Ischämie/ Reperfusionsschaden nach Pankreastransplantation im Schweinemodell

Die Transplantatpankreatitis nach Pankreastransplantation als Folge des Ischamie/Reperfusionsschadens (IRS) ist fur ca. 7% der Organverluste verantwortlich. Endothelin (ET) ist ein potenter Mediator der auftretenden Mikrozirkulationsstorungen. Fragestellung: Kann ein selektiver ET-A-Rezeptorantagonist (ET-A-RA) durch Verringerung der Mikrozirkulationsstorungen die Auspragung einer Transplantatpankreatitis reduzieren? Material und Methoden: Bei weiblichen Gottinger Minipigs (Spender: n = 14; Empfanger: n = 14) wurde eine heterotope allogene Pankreastransplantation durchgefuhrt. Das entnommene Organ wurde mit UW-Losung ex situ perfundiert und 6 h bei 4 °C in UW-Losung gelagert. Die Empfanger wurden in eine Kontroll- und Therapiegruppe randomisiert. In der Therapiegruppe erhielten die Empfanger mit Beginn der Reperfusion den ET-A-RA (BSF 208075) in einer Dosierung von 10 mg/kg i.v.und in der Kontrollgruppe NaCl. Zur Immunsuppression wurden Cyclosporin und Prednisolon verwendet. Folgende Untersuchungen zur Evaluation des IRS wurden durchgefuhrt: Sauerstoffpartialdruckmessung im Gewebe (ptiO2 Licox, GMS); Laser-Doppler-Flowmetrie (DRT4, Moor Instruments); semiquantitative Analyse histologischer Veranderungen und ET-Immunhistochemie; Bestimmung von Lipase, Amylase, Trypsinogenaktivierendem Peptid (TAP), C-reaktivem Protein (CRP) und Malondialdehyd. Das Follow-up betrug 5 Tage. Ergebnisse: Wahrend der Reperfusion kam es zu einer signifikanten Erhohung der ET-Spiegel in der Kontrollgruppe (3,4 ± 1,0 pg/ml 2 h nach Reperfusion vs 1,8 ± 0,6 pg/ ml vor Reperfusion; p < 0,05). In der Therapiegruppe zeigten sich wahrend der Reperfusion signifikant hohere ET-Spiegel als in der Kontrollgruppe als Beweis einer potenten Rezeptorblockade. Die ptiO2- und Flowmessungen ergaben zu allen Meszeitpunkten nach Reperfusion signifikant bessere Werte in der Therapiegruppe (ptiO2: 60 min nach Reperfusion 20,1 ± 3,5 vs. 35,9 dz 6,5 mmHg, p < 0,05). Histologisch zeigte sich in der Therapiegruppe 1 h und 2 d nach Reperfusion eine signifikant geringere Veranderung hinsichtlich Leukozyteninfiltration, interstitiellem odem und Hamorrhagien (p < 0.05). Die ET-Immunhistochemie, als Ausdruck der lokalen ET-Freisetzung im Transplantat, zeigte 1 h nach Reperfusion eine geringere Intensitat in der Therapiegruppe (p < 0.05). Amylase und Lipase erreichten ihren Maximalwert in beiden Gruppen nach 36 h und nach 5 d wieder Ausgangsniveau. Ein signifikanter Unterschied zugunsten der Therapiegruppe zeigte sich nur 36 h nach Reperfusion fur Amylase (Kontrolle: 457 ± 76 umol/1 vs. Therapie 346 ± 62 umol/1). CRP zeigte signifikante Unterschiede nach 12 h (10,9 ± 3,6 vs. 7,9 ± 1,3 mg/1) und 36 h (18,3 ± 2,2 vs. 13,5 ± 1,4 mg/1) zugunsten der Therapiegruppe (p < 0,05). TAP ergab 45 min nach Reperfusion im portalvenosen aber nicht im zentralvenosen Blut einen signifikant geringeren Wert in der Therapiegruppe (26,3 ± 4,3 ng/ml vs. Kontrolle: 46,7 ± 5,4 ng/ml, p < 0,05). Es bestand zwischen den Gruppen kein Unterschied im Verlauf der Malondialdehydwerte. Zusammenfassungg: Nach Reperfusion findet sich eine deutliche Beeintrachtigung der MikroZirkulation im Pankreas mit der Folge einer akuten Transplantatpankreatitis. Durch Einsatz eines selektiven ET-A-RA kann die Mikrozirkulation im Transplantat verbessert und die Auspragung einer Transplantatpankreatitis reduziert werden.