P-selectin- and CD18-mediated recruitment of canine neutrophils under conditions of shear stress

P-selectin- and CD18-mediated recruitment of canine neutrophils under conditions of shear stress

Neutrophil mobilization at sites of inflammation or thrombosis involves the participation of several adhesion molecules expressed on neutrophils and vascular endothelial cells. Local vascular damage with disruption of the endothelium results in adhesion of platelets to the exposed subendothelium, and these platelets could also participate in neutrophil recruitment. This initial phase of mobilization could be followed by heterotypic aggregation to recruit more leukocytes in the area. The present study first examined the interactions of adherent canine platelets and flowing canine neutrophils using an in vitro system that simulates vascular flow conditions. Results showed that collagen-adherent platelets express the adhesion molecule P-selectin on their surface and can support neutrophil arrest (612 +/- 43 neutrophils/mm2) at shear stresses of approximately 2. 5 dynes/cm2. Both transient adhesion (manifested by a rolling-type behavior) and complete arrest were observed. These interactions could be totally inhibited by a monoclonal antibody directed against platelet P-selectin (24 +/- 18 neutrophils/mm2) but not by a monoclonal antibody against neutrophil CD18 (625 +/- 46 neutrophils/mm2). Additionally, under shear mixing conditions (700 RPM), canine blood leukocytes exhibited aggregation (> 80% singlets recruited into aggregates after 5 minutes), and this process does not involve P-selectin but is dependent on the neutrophil integrin CD18. However, stimulation of the blood with platelet-activating factor (5-20 ng/ml) induced a rapid aggregation with a significantly greater number of aggregates when compared with stirring alone (68. 3% +/- 3. 2% versus 35. 2% +/- 6. 3% at 1 minute, P < 0. 05), and this aggregation was both P-selectin and CD18 dependent. Overall, these two mechanisms of leukocyte recruitment (neutrophil arrest on adherent platelets and aggregation) could act sequentially and in a cooperative manner to bring into close contact platelets and neutrophils at sites of inflammation and thrombosis in pathologic conditions in the dog.

                               Measuring stress in captive animals.

Stress is rarely thought to be a good thing, but is it true that all stress is necessarily bad? In the animal kingdom, such disparate events as mating, fighting, or hunting can trigger an animal's stress response.

Stress is a normal part of life, but what isn't so obvious, especially for zoologic institutions, is determining whether a captive animal is experiencing the good kind of stress that helps keep it alive or the harmful kind.

In her Oct. 11 AVMA Animal Welfare Forum presentation " Stress and distress: evaluating their impact for the well-being of zoo animals, " Nadja Wielebnowski, PhD, a behavioral endocrinologist at Chicago's Brookfield Zoo, discussed some of the studies under way to measure stress in captive animals.

" Not all stress is negative, and some stressful situations, like exploring a new element in the environment or mating, usually are seen as beneficial and can help reduce animal boredom in captivity, " Dr Wielebnowski said.

" In zoos, we are most concerned with chronic stress, when animals are repeatedly exposed to negative stressors and are not able to respond appropriately. Prolonged negative stress can become physically harmful. "

Researchers at Brookfield Zoo are currently studying stress in 18 species using techniques that may be expanded and applied to more species. The procedure includesdaily collecting, testing, and validating of fecal samples for used hormone analysis along with extensive monitoring of behavioral and physiologic changes.

Brookfield's clouded leopard collection is part of a national study started in 1999 involving 12 zoos and 74 large cats. Difficult to manage in captivity, clouded leopards are often seen pulling out their fur, pacing excessively, hiding for long periods, and acting aggressively toward exhibit mates.

Research has shown that higher-than-average concentrations corticosteroids in the feces—an indicator of stress—are positively correlated with the occurrence of self-injuring behaviors, as well as the frequency of pacing and hiding.

In addition, when husbandry factors were analyzed, it was found that enclosure height, keeper time, being able to see potential predators, and public display were all associated with hormonal changes. The higher the enclosure, for example, the lower the hormone concentrations.

The Clouded Leopard Species Survival Plan Committee of the American Zoo and Aquarium Association has already recommended increasing enclosure height to a minimum of 10 to 12 feet for this big cat species.

" Zoos have made great progress in animal welfare over the past several decades, " Dr. Wielebnowski said, " but there is still work to be done. We need to identify more accurate and reliable scientific measurements to increase our understanding of what well-being means from the individual animal's point of view. This includes species as different as the charismatic polar bear to the hissing cockroach. "