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The role of endothelial PI3 kinase activity and IQGAP1 in regulation of lymphocyte diapedesis

  • Author / Creator
    Nakhaei-Nejad, Maryam
  • Leukocyte extravasation is a fundamental process of the inflammatory responses. The mechanisms that control remodelling of endothelial (EC) shape and adhesive contacts during leukocyte transendothelial migration (TEM) are not completely understood. We studied the role of EC phosphatidylinositol 3-kinase (PI3K) activity in lymphocyte TEM under shear stress conditions. Inhibition of EC PI3K activity by its pan inhibitors decreased lymphocyte diapedesis in a step after VE-cadherin opening. The importance of PI3K catalytic isoforms (p110α, p110β, p110δ and p110γ) were studied in TEM. Treatment of EC with isoform inhibitors of p110β, p110δ and p110γ did not affect lymphocyte TEM. Inhibition of p110α activity or expression reduced lymphocyte diapedesis. PI3K activity was measured in EC exposed to shear stress alone or shear stress on cells where ICAM-1 or VCAM-1 were cross-linked. The most significant effect was seen in cells cross-linked with ICAM-1 and exposed to shear stress. This suggests that cooperation of shear-induced mechanotransduction and ICAM-1 during leukocyte interaction with EC facilitates leukocyte diapedesis by inducing PI3K. We hypothesized that Rho GTP proteins downstream of PI3K activity are involved in leukocyte TEM. We studied the role of IQGAP1, a Rac1/Cdc42 effector, during lymphocyte TEM. EC IQGAP1 knockdown decreases both microtubule (MT) tethered to the adherens junction (AJ) and lymphocyte TEM. Similarly, loss of AJ-associated MT induced by brief nocodazole (ND) treatment decreases lymphocyte TEM. Neither intervention affected leukocyte migration to the interendothelial junctions. These data indicate that IQGAP1 contributes to MT stability at endothelial junctions and is involved in the junction remodelling required for efficient lymphocyte diapedesis. We studied a candidate Rho guanine nucleotide exchange factor named FGD5 upstream of IQGAP1. Inhibition of FGD5 expression resulted in more sensitivity to apoptotic stimuli and a higher rate of apoptosis in resting conditions. Thus, we could not study the importance of FGD5 in lymphocyte TEM. Further characterization of FGD5 knockdown cells showed that they do not respond to VEGF signalling. These results suggest that FGD5 might play an important role in growth factor–mediated EC survival.

  • Subjects / Keywords
  • Graduation date
    2011-11
  • Type of Item
    Thesis
  • Degree
    Doctor of Philosophy
  • DOI
    https://doi.org/10.7939/R3J41K
  • License
    This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.
  • Language
    English
  • Institution
    University of Alberta
  • Degree level
    Doctoral
  • Department
    • Department of Medicine
  • Supervisor / co-supervisor and their department(s)
    • Murray, Allan G (Medicine)
  • Examining committee members and their departments
    • Tibbles, Lee Anne (Physiology and Pharmacology)
    • Touret, Nicolas (Biochemistry)
    • Vliagoftis, Harissios (Medicine)
    • Jahroudi, Nadia (Medicine)