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Am J Physiol Heart Circ Physiol (December 30, 2005). doi:10.1152/ajpheart.01040.2005
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Submitted on October 3, 2005
Accepted on December 23, 2005

Priming of polymorphonuclear leukocytes - a culprit in the initiation of endothelial cell Injury

Jeanna Jacobi1, Shifra Sela2*, Hector I. Cohen3, Judith Chezar4, and Batya Kristal5

1 Eliachar Research Laboratory, Western Galilee Hospital, Nahariya, Israel
2 Eliachar Research Laboratory, Western Galilee Hospital, Nahariya, Israel; Bruce Rappoport School of Medicine, Technion, Haifa, Israel
3 Pathologic Laboratory, Western Galilee Hospital, Nahariya, Israel
4 Hematology Laboratory, Western Galilee Hospital, Nahariya, Israel
5 Nephrology and Hypertension Unit, Western Galilee Hospital, Nahariya, Israel; Bruce Rappoport School of Medicine, Technion, Haifa, Israel

* To whom correspondence should be addressed. E-mail: Shifra.Sela{at}naharia.health.gov.il.

Peripheral polymorphonuclear leukocytes (PMNL) in hemodialysis patients (HD) are primed, continually releasing and exposing the vascular endothelium to soluble factors such as reactive oxygen species (ROS) and inflammatory mediators. To mimic the close proximity between PMNL and the endothelial monolayer, and to monitor and characterize the influence of soluble mediators released from PMNL, we developed a novel co-cultivation system using primary human umbilical vein endothelial cells cultures (HUVEC) and PMNL with a sieve separating the two cell types, preventing direct adhesive effects. PMNL (106) from HD patients or from healthy normal controls (NC) were co-cultivated with HUVEC (105) for 15 min. Endothelial cell injury was assessed by HUVEC morphology, cell detachment, and apoptosis. Proinflammatory changes were estimated by the expression of HUVEC adhesion molecule P-selectin, and by endothelial IL-8 and eNOS mRNA. The levels of intracellular tissue factor reflected the procoagulant state while NADPH oxidase activity served as an indicator for pro-oxidative changes in HUVEC. Mediators released from the primed PMNL triggered activation/dysfunction of endothelial cells causing: 1) increased endothelial cell detachment and apoptosis; 2) a pro-inflammatory state manifested by increased IL-8 mRNA expression and P-selectin on the endothelial surface; 3) activation of endothelial NADPH oxidase; 4) increase in endothelial cell tissue factor, which directly correlated with PMNL priming index; 5) decreased eNOS mRNA. Our data supports a pathogenic link between PMNL priming and endothelial dysfunction, suggesting PMNL priming as a potential new non-traditional risk factor for the development of atherosclerosis.




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