Paradoxical attenuation of leukocyte rolling in response to ischemia- reperfusion and extracorporeal blood circulation in inflamed tissue

Abstract

In contrast to acute preparations such as the exteriorized mesentery or the cremaster muscle, chronically instrumented chamber models allow one to study the microcirculation under "physiological" conditions, i.e., in the absence of trauma-induced leukocyte rolling along the venular endothelium. To underscore the importance of studying the naive microcirculation, we implanted titanium dorsal skinfold chambers in hamsters and used intravital fluorescence microscopy to study venular leukocyte rolling in response to ischemia-reperfusion injury or extracorporeal blood circulation. The experiments were performed in chambers that fulfilled all well-established criteria for a physiological microcirculation as well as in chambers that showed various extents of leukocyte rolling due to trauma, hemorrhage, or inflammation. In ideal chambers with a physiological microcirculation (Ͻ30 rolling leukocytes/mm vessel circumference in 30 s), ischemia-reperfusion injury and extracorporeal blood circulation significantly stimulated leukocyte rolling along the venular endothelium and, subsequently, firm leukocyte adhesion. In contrast, both stimuli failed to elicit leukocyte rolling in borderline chambers (30 -100 leukocytes/mm), and in blatantly inflamed chambers with yet higher numbers of rolling leukocytes at baseline (Ͼ100 leukocytes/mm), we observed a paradoxical reduction of leukocyte rolling after ischemia-reperfusion injury or extracorporeal blood circulation. A similar effect was observed when we superfused leukotriene B 4 (LTB4) onto the chamber tissue. The initial increase in leukocyte rolling in response to an LTB 4 challenge was reversed by a second superfusion 90 min later. These observations underscore 1) the benefit of studying leukocyte-endothelial cell interaction in chronically instrumented chamber models and 2) the necessity to strictly adhere to well-established criteria of a physiological microcirculation. leukotriene B 4; animal model; intravital microscopy THE MULTISTEP CONCEPT of leukocyte-endothelial cell interaction proposes early leukocyte tethering and rolling along the endothelial wall followed by firm leukocyte adhesion and transendothelial emigration; each consecutive step involves distinct adhesion molecules and distinct chemoattractant and/or adhesion-promoting mediators (5,21,23,29). It is widely established that leukocyte rolling along the endothelium constitutes a prerequisite for subsequent leukocyte adhesion and emigration (22). However, Kanwar and co-workers (21) identified an enormous redundancy in this system: even an inhibition of leukocyte rolling by Ͼ90% with fucoidin or selectin antagonists exerted no significant effect on subsequent firm leukocyte adhesion.In intact animal organisms, the study of leukocyte-endothelium interaction can be accomplished by intravital microscopy on acutely prepared tissues (e.g., hamster cheek pouch, cremaster muscle, and exteriorized mesentery) or on chronically instrumented tissues [e.g., the dorsal skinfold chamber model in hamsters (10, 28) and mice (27)...

Endogenous testosterone increases leukocyte–endothelial cell interaction in spontaneously hypertensive rats

“…Although multiple mediators work together in inflammation, LTB 4 has been implicated in the initiation of leukocyte-endothelium interaction during inflammation (Schafer et al, 2005). In our study, the enhanced LTB4-induced leukocyte adhesion observed in hypertensive rats was attenuated by castration, which suggests that endogenous testosterone exerts a proinflammatory effect contributing not only for the increased leukocyte rolling but also for the increase in leukocyte adherence in SHRs.…”

“…Leukocytes adhering to the endothelium were quantified following the superfusion of the irritant stimulus leukotriene B 4 (LTB 4 20 nM, Cayman Chemical CO, EUA) for 10 min (Schafer et al, 2005). A leukocyte was considered to be adherent to the venular endothelium if it remained stationary for more than 30 s along 100-μm length of venule.…”

Potassium restores vasorelaxation of resistance arterioles in non-hypertensive DOCA/salt fed mice

“…This number compares well to experiences made in other immunocompetent mouse species (C57BL/6, BALB/c, and ApoE KO, all showing an elimination rate of around 40%, see Li and Förstermann, 2000;Schaefer et al, 2005), but was slightly higher when compared to mice that are genetically hairless (nu/nu or nude, showing an elimination rate of around 25%; see Lehr et al, 1993). It should be acknowledged that the success rate of the chamber preparation depends largely on the quality of microsurgical technique and hence varies from experimentor to experimentor (unpublished observation), and that unexpected results are obtained in experiments in which the above described strict criteria are neglected (Schäfer et al, 2005). To eliminate the effects of anesthesia and direct surgical trauma on the microvasculature, a recovery period of 72-96 h was allowed between the implantation of the observation chambers and the intravital microscopic investigations.…”

Leukocyte-endothelial cell interaction is necessary for photodynamic therapy induced vascular permeabilization

“…The cells were counted for 30 seconds and normalized by cross‐section circumference of each microvessel. Sticking was defined as stable binding at a certain position for more than a 30 seconds period, expressed as the number of cells per endothelial surface (as calculated from the microvessel diameter and the defined length of 200 µm) 11. At least 3 different vessels were analyzed per animal, and measurements were performed before PDT, 90 and 180 minutes after PDT.…”