B. Vollmar

Depressed phagocytic activity of Kupffer cells after warm ischemia-reperfusion of the liver

Phagocytic activity of Kupffer cells following hepatic ischemia/reperfusion was studied in 39 livers of male Sprague-Dawley rats by in vivo fluorescence microscopy. Animals were subjected to either 20 min (group B, n = 9) and 60 min left hepatic lobar ischemia (group C, n = 9) or to 20 min of global hepatic ischemia (group D, n = 11). Sham-operated animals without ischemia served as controls (group A, n = 10). After 60 min postischemic reperfusion, fluorescent latex beads (3 x 10(8).kg body wt-1; diameter: 1.1 microns) were injected intra-arterially. The zonal distribution and kinetics of adherence of latex beads were quantified by off-line video analysis. After 20 min of left hepatic lobar ischemia, 50%, 38% and 12% of injected latex beads adhered in zones 1, 2 and 3, respectively, and did not significantly differ from control livers (group A: 57%, 32% and 11%). In contrast, after 60 min of left hepatic lobar ischemia (group C) as well as after 20 min of global hepatic ischemia (group D), a more homogeneous distribution of latex beads adherent in zones 1, 2 and 3 was observed (group C: 48%, 36% and 16%; group D: 48%, 36% and 16%). Kinetic analysis of phagocytosis (% adherence of visible latex beads 1 min and 3 min after injection) showed no significant difference between 20 min left hepatic lobar ischemia (group B: 84% and 95%) and control (group A: 81% and 95%).(ABSTRACT TRUNCATED AT 250 WORDS)

Release of Arachidonic Acid Metabolites during Acute Pancreatitis in Pigs

The pancreatic release of arachidonic acid metabolites was studied in a porcine model of acute pancreatitis. In situ isolation of the pancreatic gland enabled selective collection of pancreatic venous blood, pancreatic lymph, and ascites fluid. Three experimental groups were studied: 1) control (n = 9); 2) hemorrhagic pancreatitis induced by injection of 5% bile salt (sodium taurocholate) into the pancreatic duct (n = 10); and 3) edematous pancreatitis induced by injection of free fatty acid (FFA) into the pancreatic artery (n = 10). Determinations of cyclooxygenase metabolites were performed by radioimmunoassay; lipoxygenase metabolites (LTC4, LTD4) were measured by radioimmunoassay after purification by high-performance liquid chromatography. Prostaglandin (PG)F1 alpha, thromboxane B2, and PGF2 alpha concentrations were almost doubled in the lymph of the FFA group during pancreatitis, as were PGF1 alpha levels in pancreatic venous blood. However, concentrations of cyclooxygenase metabolites remained unchanged in the control group and in the bile salt group. Concentrations of LTC4 and LTD4 in lymph and ascites fluid of both pancreatitis groups increased from about 50 pg/ml to a mean level of 600 pg/ml at 6 h. Leukotriene concentrations in the control group were consistently below 50 pg/ml. The results of this study indicate that above all LTC4 and LTD4 are released from the organ and that these arachidonic acid metabolites may be also involved in the events following acute pancreatitis contributing to the systemic effects of the disease.

Oleic acid induced pancreatitis in pigs

An experimental model of edematous pancreatitis in pigs was established and measurement of pancreatic macro- and microcirculatory parameters and determinations of pancreatic enzymes (lipase, phospholipase A) and vasoactive mediators (prostanoids, kallikrein, kininogen) were performed. During general anesthesia the pancreas was isolated in situ. Pancreatic microcirculatory parameters were measured using videofluorescence microscopy after iv administration of FITC-Dextran. In hourly collected samples lipase and phospholipase A activities were determined enzymatically, concentrations of kallikrein, kininogen, and selected prostanoids were measured by radioimmunoassay. Two experimental groups were studied: (1) control (n = 9); (2) edematous pancreatitis induced by injection of oleic acid into the pancreatic artery (free fatty acid, ffa; n = 10). The animals were followed up for 6 hr. Systemic hemodynamic parameters remained constant in both groups. In the pancreatitis group pancreatic blood flow and O2-consumption decreased significantly (-55 and -49%), while pancreatic vascular resistance increased significantly (+50%). During baseline conditions 41% of all capillaries were perfused. In the pancreatitis group there were both areas with persistent stasis as well as areas with continuous perfusion. However, in the latter areas the portion of perfused capillaries decreased significantly to 27%. In the control group the portion of perfused capillaries remained constant. Liberation of lipase and phospholipase A especially into lymph and ascites fluid was measured during pancreatitis. Furthermore, considerable releases of kallikrein into lymph (+50%) and ascites (+800%) and a marked consumption of kininogen in lymph (+90%) and in ascites fluid (+80%) were measured. Activation of the arachidonic acid cascade and a significant release of prostacyclin and thromboxane A2 into pancreatic venous blood and lymph was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Modified Retrograde Orotracheal Intubation Technique for Airway Access in Rabbits

Anatomical features of the oral fissure and pharynx of rabbits make orotracheal intubation a difficult task with a high failure rate. We have developed a versatile technique for retrograde intubation of rabbits which consists of the following steps: (1) assessment of the trachea through a cervicotomy (2) insertion of a guidewire into its lumen, (3) delivery of the guide through the mouth, and (4) introduction of the orotracheal tube over the guide. This procedure was performed in 58 rabbits under pentobarbital anesthesia and resulted in successful orotracheal intubation in 56 animals. Awakening from anesthesia and extubation were uneventful. No complications related to this technique were observed during a follow-up period of 5-14 days. The described technique is a simple and safe method to access the airways of rabbits and can be useful in chronic experiments.

In Vivo Analysis of Hepatic NADH Fluorescence

Direct observation of intracellular events and their relationship to physiological function presents a challenging and long-standing problem. The determination of intracellular oxygen levels in tissues has been a subject of continuous discussion, but most of the techniques for measuring oxygen tension in blood and tissue fail to indicate the oxidationreduction states of the respiratory carriers. Observations of the latter, however, would be of particular importance, since they reflect the intracellular level of energy transfering components. Reduced nicotinamide adenine dinucleotide (NADH) is one of the main means of energy transfer from the tricarboxylic acid cycle to the respiratory chain in the mitochondria. Most of the energy derived from aerobic metabolism is initially generated as NADH, which is oxidized to NAD+ as electrons are passed to the electron transport chain and, ultimately, to molecular oxygen. Decades ago, Chance and coworkers pioneered the fluorescence property of NADH as an indicator of the mitochondrial redox state and, in the presence of sufficient substrate and phosphate, as an indicator of cellular oxygen (Chance et al., 1962; Chance, 1976). For example, interruption of oxidative phosphorylation is associated with a prompt increase of tissue NADH levels, and that increase can be monitored by following its optical fluorescence (Chance and Schoener, 1965; Chance et al., 1965).The optical properties of NADH and NAD+ clearly differ in that upon ultraviolet excitation (330–390 nm), NADH, unlike NAD+, fluoresces in the blue (>430 nm). Meanwhile, NADH fluorescence measurements have been applied to single cells, cell suspensions, tissue slices, ex vivo perfused organs, but also to organs in vivo (Ince et al., 1992).

Die pankreasdurchblutung bei der experimentellen biliären pankreatitis

SummaryMeasurements of pancreatic micro- and macrocirculation were performed to evaluate the pancreatitis-induced changes. Pigs were anesthetized and ventilated mechanically. Hypotension induced side-effects were avoided by adequate volume replacement. After laparatomy, splenectomy and gastroectomy the animals were enterotomized. Systemic hemodynamic parameters were monitored as well as pancreatic blood flow (Q), which was measured electromagnetically, and arterial and portal-venous blood gases. Pancreatic microcirculatory parameters were observed using fluorescence-videomicroscopy after i.v. administration of FITC dextran 150 and FITC labeled autologous erythrocytes. The pigs were randomly assigned to a control (n = 9) and a pancreatitis group (n=10), the later being induced by the retrograde infusion of sodium-taurocholate. Systemic and pancreatic macrohemodynamic parameters remained constant in both groups, except for avd02 and O2-consumption (O2-c) decreasing significantly in the pancreatitis group. At baseline 42% of all capillaries were perfused in both groups. In pancreatitis we detected focal areas with persistent stasis and areas which were continuously perfused. In these areas the portion of capillaries perfused by erythrocytes increased significantly to 67%. This was accompanied by an extravasation of FITC dextran. The finding of an unchanged Q beside reduced 02-c and avd02 during pancreatitis is explained by the changes in pancreatic microcirculation. Focal stasis was observed beside areas showing typical signs of an acute inflammation: increased macromolecular permeability and capillary recruitment, e.g. oedema and hyperaemia.ZusammenfassungEs wurde ein Modell etabliert, an dem gleichzeitig die Verdnderungen der Makro- und Mikrozirkulation des Pankreas gemessen werden können. Das Pankreas wurde dazu in situ isoliert. An diesem Modell wurde eine akute Pankreatitis Durch retrograde Infusion von Natrium-Taurocholat-Lösung (NaT) in den Pankreasgang ausgelöst. Durch ausreichende Fliissigkeitszufuhr wurde ein hypovoldmischer Schock verhindert. Die beobachteten Verdnderungen der hämodynamischen Parameter sind somit pankreatitisbedingt. Im Pankreatitismodell war intravitalmikroskopisch frühzeitig eine Permeabilitdtssteigerung fur Makromoleküle an der Pankreasoberfläche zu beobachten. Der Gefäßwiderstand des Pankreas blieb im Modell der bilidren Pankreatitis unverändert. Die Perfusionsausfälle in den Stasebereichen waren von einer Steigerung der funktionellen Kapillardurchblutung in den übrigen Bereichen begleitet. Aüßerdem war eine Abnahme der Sauerstoffausschöpfung des Blutes zu beobachten. Man kann somit von einer funktionellen Shuntdurchblutung sprechen. Die Organgesamtdurchblutung blieb in diesem Modell unverändert. Es ließen sich in diesem Pankreatitismodell somit die hdmodynamischen Zeichen einer echten Entzündung, Permeabilitätsstörung and Hyperdmie, nachweisen.Measurements of pancreatic micro- and macrocirculation were performed to evaluate the pancreatitis-induced changes. Pigs were anesthetized and ventilated mechanically. Hypotension induced side-effects were avoided by adequate volume replacement. After laparatomy, splenectomy and gastroectomy the animals were enterotomized. Systemic hemodynamic parameters were monitored as well as pancreatic blood flow (Q), which was measured electromagnetically, and arterial and portal-venous blood gases. Pancreatic microcirculatory parameters were observed using fluorescence-videomicroscopy after i.v. administration of FITC dextran 150 and FITC labeled autologous erythrocytes. The pigs were randomly assigned to a control (n = 9) and a pancreatitis group (n = 10), the later being induced by the retrograde infusion of sodium-taurocholate. Systemic and pancreatic macrohemodynamic parameters remained constant in both groups, except for avdO2 and O2-consumption (O2-c) decreasing significantly in the pancreatitis group. At baseline 42% of all capillaries were perfused in both groups. In pancreatitis we detected focal areas with persistent stasis and areas which were continuously perfused. In these areas the portion of capillaries perfused by erythrocytes increased significantly to 67%. This was accompanied by an extravasation of FITC dextran. The finding of an unchanged Q beside reduced O2-c and avdO2 during pancreatitis is explained by the changes in pancreatic microcirculation. Focal stasis was observed beside areas showing typical signs of an acute inflammation: increased macromolecular permeability and capillary recruitment, e.g. oedema and hyperaemia.

Enzymfreisetzung und Aktivierung der Kallikrein-Kinin-Systeme bei experimenteller Pankreatitis

The clinical course of acute pancreatitis is strongly influenced by secondary cardiac, pulmonary and renal damage. The aim of the present study was to gather information about the compartment promoting the systemic damage. Therefore the activity of lipase, phospholipase A and plasmaprokallikrein and the concentration of tissue kallikrein and kininogen were measured in portal venous blood, pancreatic lymph and peritoneal exudate. Anaesthetized pigs were subjected to fluid resuscitation to keep systemic haemodynamic parameters constant. The pancreas was isolated in situ. The pigs were randomly assigned to a control group (n = 9) or one of the two pancreatitis groups (n = 10 each). Pancreatitis was induced by i.a. infusion of free fatty acid (FFS) or retrograde infusion of 5 % sodium taurocholate intraductally (NaT). In both pancreatitis groups the activity of lipase and phospholipase A increased. The most pronounced changes were seen in the peritoneal exsudate (phospholipase A activity 40 min after induction: control 10.0 U/1, NaT 72.2 U/1). In both pancreatitis groups there was evidence for activation of the tissue kallikreinkinin system in the form of an increase in the kallikrein concentration and a decrease in the kininogen concentration. Again the changes were most pronounced in the peritoneal exsudate (tissue kallikrein 40 min after induction: control 14.7 ng/ml, NaT 452 ng/ml).ZusammenfassungDas klinische Bild der akuten Pankreatitis wird entscheidend durch die sekundäre Schädigung von Herz-Kreislauf-System, Lunge und Niere bestimmt. Ziel der vorliegenden Untersuchung war es, durch Messungen in venösem Pankreasblut, Pankreaslymphe und Peritonealexsudat die Kompartimente zu bestimmen, über die die systemischen Schädigungen vermittelt werden. An anästhesierten Schweinen wurden die systemischen, hämodynamischen Parameter durch gesteuerte Volumentherapie konstant gehalten. Die Schweine wurden randomisiert der Kontrollgruppe (n = 9) oder einer der Pankreatitisgruppen zugeteilt (jeweils n = 10). Die Pankreatitis wurde durch Infusion von freier Fettsäure in die Pankreasarterien (FFS) oder durch Infusion einer 5%igen Natrium-Taurocholat-Lösung retrograd in den Pankreasgang (NaT) ausgelöst. Nach Isolation des Pankreas wurde venöses Pankreasblut, Pankreaslymphe und Peritonealexsudat gewonnen und die Aktivität von Lipase, Phospholipase A und Plasmaprokallikrein sowie die Konzentration von Organkallikrein und Kininogen bestimmt. In beiden Pankreatitismodellen fand sich ein Anstieg der Enzymaktivitäten. Die höchsten Aktivitäten fanden sich im Peritonealexsudat (Phospholipase A nach 40 min: Kontrolle 10,0 U/1, NaT 72,2 U/1). In beiden Pankreatitismodellen fanden sich außerdem Hinweise für eine Aktivierung des Organkallikrein-Kinin-Systems durch den Anstieg der Organkallikreinkonzentration und den Abfall der Gesamtkininogenkonzentration. Die stärksten Veränderungen fanden sich wieder im Peritonealexsudat (Organkallikrein nach 40 min: Kontrolle 14,7 ng/ml, NaT 452 ng/ml).

Gaseous Oxygenation of the Ischemic Rat Liver

The lack of aerobic metabolism during ischemic organ preservation plays a pivotal role in the development of tissue alterations during the storage period and favours the manifestation of reper-fusion injuries, deteriorating organ viability upon postischemic resumption of nutritive blood circu-lation. Gaseous insufflation of oxygen via the venous vascular system has proven to be a useful tool to prevent anoxic tissue injury during extended time periods of ischemic preservation of kidneys (Isselhard et al., 1972) or livers (Minor and Isselhard, 1996; Minor et al., 1996) and to allow for an improved recovery of the persufflated organ after transplantation (Fischer et al., 1978; Minor et al., 1997b; Rolles et al., 1989).

Reduktion des postischämischen Reperfusionsschadens der Leber durch anti-ICAM-1

Nach leberchirurgisehen Eingriffen mit temporarer Gefasokklusion konnen protrahierte Ischamie und unzureichende Reperfusion zu postoperativer Einschrankung der Leberfunktion fuhren. Als Ursache des postischamischen Reperfusionsschadens der Leber werden fehlende Reperfusion nutritiver Sinusoide sowie Aktivierung und Adharenz von Leukozyten am mikrovaskularen Endothel mit nachfolgender Emigration ins Gewebe diskutiert. Die Interaktion von Leukozyten mit dem mikrovaskularen Endothel wird entsprechend einer Mehr-Schritt-Sequenz (Marginatum, ”rolling”, Adharenz und transendotheliale Migration) von spezifischen Adhasionsrezeptoren auf der Oberflache von Leukozyten (L-Selektin; β-Integrine) und Endothelzellen (P-, E-Selektin; Immunglobuline (ICMA-1/2)) vermittelt und reguliert [1]. Ziel der Studie war es, die Wirkung eines monoklonalen Antikorpers gegen das endotheliale Adhasionsmolekul ICMA-1 (anti-ICAM-1) auf den mikrovaskularen und funktionellen Reperfusionsschaden der Leber quantitativ zu erfassen.

Ein Modell zur Untersuchung der Vasomotorik coronarer Mikrogefäße:Coronare Dilatation durch Adenosin

Coronardilatoren wie Adenosin konnen die myokardiale Durchblutung um ein Vielfaches steigern und die Sauerstoffbilanz des Herzmuskels massiv verbessern, bewirken jedoch nicht unbedingt eine gleichsinnige Verbesserung der myokardialen Gewebeoxygenation (1). Dies konnte auf der bereits fruher beobachteten Wirkung von Adenosin vornehmlich auf die coronaren Widerstandsgefase (2) und einer daraus folgenden Storung der Blutflusvertei- lung auf der Ebene der coronaren Mikrozirkulation beruhen.