Sequential Cell Signaling Events During Leukocyte Transendothelial Migration

Project: Research project

Project Details

Description

Recruitment of leukocytes during the inflammatory response involves a series of adhesive steps that allow them to tether and roll along the vessel well, firmly adhere to the apical surface of the endothelium, locomote to the endothelial borders, and finally migrate through the endothelium towards the inflamed tissue. Transendothelial migration (TEM), or diapedesis, is arguably the most critical stage in the inflammatory response.
Several signaling events have been shown to be required for TEM. 1) Platelet/endothelial cell adhesion molecule 1 (PECAM) is concentrated at the endothelial cell border and is diffusely expressed on leukocytes. Homophilic interaction between leukocyte PECAM and endothelial PECAM is critical for TEM. 2) An interconnected vesicular network located just beneath the endothelial plasma membrane at cell borders called the lateral border recycling compartment (LBRC) contains adhesion molecules, such as PECAM and CD99, that are necessary for TEM. During TEM, the LBRC is actively recruited to the adherent leukocyte in a process called targeted recycling, where it exteriorizes unligated PECAM and CD99 to promote diapedesis. Blocking the targeted movement of the LBRC membrane blocks TEM. 3) A transient increase in endothelial cytosolic free calcium ion concentration ([Ca2+]i) is also required for leukocyte TEM.
The AHA predoctoral fellowship that I have been working on for the past 1 ½ years has investigated the relationship of these critical signaling events. My results, employing complementary methodologies, show that these three events are intimately connected. PECAM homophilic interactions are upstream of endothelial [Ca2+]i in the pathway that subsequently recruits the LBRC and governs TEM. Furthermore, I have identified canonical transient potential receptor channel 6 (TRPC6) as the calcium channel responsible for specifically promoting the [Ca2+]i that is required for TEM using both knockdown and selective agonist strategies.
However, I believe that with an additional year of experimentation, these studies could have far greater impact. I have designed ways to study calcium signaling during TEM under conditions of physiological shear flow and now have a dominant negative TRPC6 construct to complement our knockdown studies. Most important, we are equipped to validate the role of endothelial TRPC6 in TEM in vivo. Therefore, I am applying for an additional year of funding to bring these studies to a higher plane of significance.
StatusFinished
Effective start/end date1/1/1412/31/14

Funding

  • American Heart Association Midwest Affiliate (14PRE18550021)

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