Morphological and quantitative analysis of leukocytes in free-living Australian black flying foxes (Pteropus alecto).

The black flying fox (Pteropus alecto) is a natural reservoir for Hendra virus, a paramyxovirus that causes fatal infections in humans and horses in Australia. Increased excretion of Hendra virus by flying foxes has been hypothesized to be associated with physiological or energetic stress in the reservoir hosts. The objective of this study was to explore the leukocyte profiles of wild-caught P. alecto, with a focus on describing the morphology of each cell type to facilitate identification for clinical purposes and future virus spillover research. To this end, we have created an atlas of images displaying the commonly observed morphological variations across each cell type. We provide quantitative and morphological information regarding the leukocyte profiles in bats captured at two roost sites located in Redcliffe and Toowoomba, Queensland, Australia, over the course of two years. We examined the morphology of leukocytes, platelets, and erythrocytes of P. alecto using cytochemical staining and characterization of blood films through light microscopy. Leukocyte profiles were broadly consistent with previous studies of P. alecto and other Pteropus species. A small proportion of individual samples presented evidence of hemoparasitic infection or leukocyte morphological traits that are relevant for future research on bat health, including unique large granular lymphocytes. Considering hematology is done by visual inspection of blood smears, examples of the varied cell morphologies are included as a visual guide. To the best of our knowledge, this study provides the first qualitative assessment of P. alecto leukocytes, as well as the first set of published hematology reference images for this species.

Zero-fluoroscopy Radiofrequency Redo Ablation of Atrial Tachycardia Following Pulmonary Vein Isolation: A Tale of Two Systems.

Regular atrial tachycardia (AT) is one of the most important proarrhythmic complications that may occur following left atrial pulmonary vein isolation (PVI). These tachycardias that develop after atrial fibrillation ablation may lead to worse symptoms than those from the original arrhythmia existing prior to the index ablation procedure. Ablation of various types of supraventricular tachycardias without the use of fluoroscopy has been shown to be feasible in both children and adults using three-dimensional mapping systems. We describe the case of a 71-year-old woman who developed a focal AT after a successful PVI procedure. The initial ablation failed with one mapping system. Repeat electrophysiologic study despite antiarrhythmic medications revealed the same focal AT, which was successfully ablated with a different mapping system. Both ablations were performed without fluoroscopy.

12/15-Lipoxygenase mediates high-fat diet-induced endothelial tight junction disruption and monocyte transmigration: a new role for 15(S)-hydroxyeicosatetraenoic acid in endothelial cell dysfunction.

A convincing body of evidence suggests that 12/15-lipoxygenase (12/15-LO) plays a role in atherosclerosis. However, the mechanisms of its involvement in the pathogenesis of this disease are not clear. Therefore, the purpose of this study is to understand the mechanisms by which 12/15-LO mediates endothelial dysfunction. 15(S)-Hydroxyeicosatetraenoic acid (15(S)-HETE), the major 12/15-LO metabolite of arachidonic acid (AA), induced endothelial barrier permeability via Src and Pyk2-dependent zonula occluden (ZO)-2 tyrosine phosphorylation and its dissociation from the tight junction complexes. 15(S)-HETE also stimulated macrophage adhesion to the endothelial monolayer in Src and Pyk2-dependent manner. Ex vivo studies revealed that exposure of arteries from WT mice to AA or 15(S)-HETE led to Src-Pyk2-dependent ZO-2 tyrosine phosphorylation, tight junction disruption, and macrophage adhesion, whereas the arteries from 12/15-LO knock-out mice are protected from these effects of AA. Feeding WT mice with a high-fat diet induced the expression of 12/15-LO in the arteries leading to tight junction disruption and macrophage adhesion and deletion of the 12/15-LO gene disallowed these effects. Thus, the findings of this study provide the first evidence of the role of 12/15-LO and its AA metabolite, 15(S)-HETE, in high-fat diet-induced endothelial tight junction disruption and macrophage adhesion, the crucial events underlying the pathogenesis of atherosclerosis.

Both Kdr and Flt1 play a vital role in hypoxia-induced Src-PLD1-PKCγ-cPLA(2) activation and retinal neovascularization.

To understand the mechanisms of Src-PLD1-PKCγ-cPLA2 activation by vascular endothelial growth factor A (VEGFA), we studied the role of Kdr and Flt1. VEGFA, while having no effect on Flt1 phosphorylation, induced Kdr phosphorylation in human retinal microvascular endothelial cells (HRMVECs). Depletion of Kdr attenuated VEGFA-induced Src-PLD1-PKCγ-cPLA2 activation. Regardless of its phosphorylation state, downregulation of Flt1 also inhibited VEGFA-induced Src-PLD1-PKCγ-cPLA2 activation, but only modestly. In line with these findings, depletion of either Kdr or Flt1 suppressed VEGFA-induced DNA synthesis, migration, and tube formation, albeit more robustly with Kdr downregulation. Hypoxia induced tyrosine phosphorylation of Kdr and Flt1 in mouse retina, and depletion of Kdr or Flt1 blocked hypoxia-induced Src-PLD1-PKCγ-cPLA2 activation and retinal neovascularization. VEGFB induced Flt1 tyrosine phosphorylation and Src-PLD1-PKCγ-cPLA2 activation in HRMVECs. Hypoxia induced VEGFA and VEGFB expression in retina, and inhibition of their expression blocked hypoxia-induced Kdr and Flt1 activation, respectively. Furthermore, depletion of VEGFA or VEGFB attenuated hypoxia-induced Src-PLD1-PKCγ-cPLA2 activation and retinal neovascularization. These findings suggest that although VEGFA, through Kdr and Flt1, appears to be the major modulator of Src-PLD1-PKCγ-cPLA2 signaling in HRMVECs, facilitating their angiogenic events in vitro, both VEGFA and VEGFB mediate hypoxia-induced Src-PLD1-PKCγ-cPLA2 activation and retinal neovascularization via activation of Kdr and Flt1, respectively.