PMNs facilitate translocation of platelets across human and mouse epithelium and together alter fluid homeostasis via epithelial cell–expressed ecto-NTPDases
Thomas Weissmüller, … , Glenn T. Furuta, Sean P. Colgan
Thomas Weissmüller, … , Glenn T. Furuta, Sean P. Colgan
Published October 16, 2008
Citation Information: J Clin Invest. 2008;118(11):3682-3692. https://doi.org/10.1172/JCI35874.
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Research Article Inflammation

PMNs facilitate translocation of platelets across human and mouse epithelium and together alter fluid homeostasis via epithelial cell–expressed ecto-NTPDases

Abstract

Mucosal diseases are often characterized by an inflammatory infiltrate that includes polymorphonuclear leukocytes (PMNs), monocytes, lymphocytes, and platelets. A number of studies have suggested that the interaction of platelets with leukocytes has an essential proinflammatory role. Here, we examined whether platelets migrate across mucosal epithelium, as PMNs are known to do, and whether platelets influence epithelial cell function. Initial studies revealed that human platelets did not efficiently transmigrate across human epithelial cell monolayers. However, in the presence of human PMNs, platelet movement across the epithelium was proportional to the extent of PMN transmigration, and strategies that blocked PMN transmigration diminished platelet movement. Furthermore, platelet-PMN comigration was observed in intestinal tissue derived from human patients with inflammatory bowel disease (IBD). The translocated platelets were found to release large quantities of ATP, which was metabolized to adenosine via a 2-step enzymatic reaction mediated by ecto-nucleotidases, including CD73 and ecto–nucleoside triphosphate diphosphohydrolases (ecto-NTPDases), expressed on the apical membrane of the intestinal epithelial cells. In vitro studies and a mouse model of intestinal inflammation were employed to define a mechanism involving adenosine-mediated induction of electrogenic chloride secretion, with concomitant water movement into the intestinal lumen. These studies demonstrate that ecto-NTPDases are expressed on the apical membrane of epithelial cells and are involved in what we believe to be a previously unappreciated function for platelets in the inflamed intestine, which might promote bacterial clearance under inflammatory conditions.

Authors

Thomas Weissmüller, Eric L. Campbell, Peter Rosenberger, Melanie Scully, Paul L. Beck, Glenn T. Furuta, Sean P. Colgan

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Figure 1

Influence of PMNs on platelet translocation.

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Influence of PMNs on platelet translocation. Activated PMNs (106; 10–6 M...
Activated PMNs (106; 10–6 M fMLP) were added together with 6 × 106 platelets to the basolateral surface of confluent T84 cells after preincubation with antibodies directed against CD11b, CD47, P-selectin, binding control (anti–MHC class I), or vehicle (Veh). Transmigrated PMNs and platelets were then assessed by microscopic evaluation (A and B). Results are derived from 6 monolayers in each condition and are expressed as mean ± SD. *P < 0.01 compared with no-antibody control. (C) Transmigrated PMNs and platelets were incubated with 1 μg/ml BCECF for 20 minutes at 37°C. Cells were concentrated 4-fold in HBSS, and fluorescent cells were imaged by fluorescence microscopy. Shown here are examples of images after transmigration, where thick arrows indicate PMNs and thin arrows, platelets. Original magnification, ×400. (D) Platelet-PMN cotransmigration in crypt abscesses from human IBD. Depicted here are merged fluorescence immunohistochemistry images showing localization of PMNs in green (anti-MPO), platelets in red (anti-CD41), and nuclei in blue (DAPI) (left panel); secondary antibody–only control (middle panel); and an adjacent section stained with H&E demonstrating the crypt abscess (right panel). Arrows in the left panel indicate areas of platelet-PMN colocalization, shown in yellow. Scale bars: 10 μm.