Interaction of neutrophils and endothelium in isolated coronary venules and arterioles YUAN YUAN, RUTH A. MIER, WILLIAM M. CHILIAN, DAVID C. ZAWIEJA, AND HARRIS J. GRANGER Microcirculation Research Institute and Department of Medical Physiology, Texas A & M University Health Science Center, College Station, Texas 77843 Yuan, Yuan, Ruth A. Mier, William M. Chilian, David C. Zawieja, and Harris J. Granger. Interaction of neutro- phils and endothelium in isolated coronary venules and arteri- oles. Am. J. Physiol. 268 (Heart Circ. Physiol. 37): H490- H498, 1995.-This study reports measurements of porcine neutrophil dynamics in isolated microvessels. Porcine coro- nary venules and arterioles were isolated, cannulated, and perfused with fluorescently labeled neutrophils at a series of flow velocities. In venules (62.50 t 5.41 km diam) under control conditions, rolling neutrophils were often observed at intraluminal flow velocities ranging from 600 to 6,000 pm/s, and the rolling fraction varied inversely as a function of flow velocity. There was no significant adherence under the control conditions at any of the various flow velocities. Pretreatment of the neutrophils with human recombinant complement 5a (C5a, lo-* M) increased adherence at low flow velocities but did not alter the rolling fraction. In contrast to venules, rolling neutrophils were not observed in arterioles (58.80 2 5.6 pm diam). Furthermore, neutrophils that were pretreated with C5a did not adhere to the arteriolar endothelium even at low flow velocities. We suggest that 1) isolated microvessels per- fused with fluorescently labeled neutrophils are suitable mod- els for the study of the interaction between neutrophils and the microvascular endothelium, 2) shear force plays an impor- tant role in neutrophil rolling in coronary venules but is not the major factor that prevents neutrophil rolling and adher- ence in arterioles, and 3) C5a causes neutrophil adherence in venules but not in arterioles, indicating that different mecha- nisms underlie the interaction between neutrophils and endo- thelium in venules and arterioles. rolling; adherence; shear rate; adhesion molecule; complement 5a INTERACTIONS between circulating neutrophils and the endothelial surface in microvessels occur under dynamic conditions. Rolling of neutrophils along the venular wall has been well characterized as a physiological phenom- enon. Altered neutrophil-endothelial interactions have been observed in many pathological processes, including acute inflammation and ischemia-reperfusion injury. The sequence of events involves neutrophil adherence to microvascular endothelium, diapedesis, and release of cytotoxic mediators, which play an active role in host defense as well as act as deleterious factors in the development of diseases. Although the importance of neutrophil-endothelium interactions has been well rec- ognized, there is a paucity of data available for neutro- phi1 dynamics in microvessels of parenchymal organs, such as the heart.. Current approaches for the study of neutrophil adher- ence and migration include in vitro assays and in vivo microscopic observations. In vitro assays involve obser- vations of neutrophil attachment to confluent endothe- lial monolayer or human amniotic membrane (4,10,30). A modified approach to measure adhesion under flow is based on the parallel-plate chamber, in which fluid shear stress can be defined for a given flow rate (9, 18). However, these in vitro studies may not reflect in vivo conditions, especially because the experiments are usu- ally carried out with cultured endothelial cells from major vessels rather than microvessels. On the other hand, investigators (2, 12, 13, 19, 21) have employed intravital microscopy to observe the behavior of neutro- phils in microvessels of living tissues. Although these in vivo approaches provide physiologically relevant informa- tion, they are unable to specify the contribution of individual factors in the interaction between neutro- phils and endothelium due to the complicating influ- ences from hemodynamic factors, blood components, and parenchymal cells. Moreover, most investigations have employed transillumination, which is limited to membranous or very thin tissues. We have developed an experimental model for the study of the interaction between neutrophils and micro- vascular endothelium in isolated coronary arterioles and venules. This approach allows quantitative measure- ments of rolling, adherence, and migration in specific microvascular segments under well-controlled intralu- minal flow velocity (shear rate). With this preparation the extrinsic factors that can influence neutrophil- endothelium interactions, including blood components and mast cells, are limited or controlled. Consequently, the technique provides precise evaluation of the direct effects of adhesion molecules and inflammatory media- tors on the neutrophil dynamics in microvessels of the heart. MATERIALS AND METHODS General Preparation Pigs weighing 9-13 kg were sedated with ketamine (2.5 mg/kg im) and xylazine (Rompun, 2.25 mg/kg im), anesthe- H490 0363-6135/95 $3.00 Copyright o 1995 the AmericanPhysiological Society