Flowchart: Preparation: Rac1 11
 
 
 
 

 D-G

 

 BC

 

  A

 
  

Text Box: Vav                            

                                    

 


Text Box: Rac1 

 

 

 

Text Box: Mekk                                            

 

 

PURPOSE OF REVIEW: This review focuses on recent developments in understanding regulation of leukocyte transendothelial migration by small GTPase signaling. RECENT FINDINGS: New studies are refining the model for GTPase regulation of leukocyte-endothelial cell interactions that occur during leukocyte transmigration. An emerging theme is that the endothelial cell is an active participant in this process; an example of this is the identification of a novel leukocyte docking structure. The role of second messengers such as reactive oxygen species downstream and the involvement of kinases such as Pyk2 and Tec kinases upstream of GTPase activation is becoming appreciated. In the leukocyte, finer distinctions between closely related GTPases like Rac1 and Rac2 are being made, and a new role for RhoH has been characterized. Finally, the focus on Rap1 as a key regulator of leukocyte integrin-dependent adhesion is expanding to include roles in endothelial cell-cell adhesion and junctional regulation during transmigration. SUMMARY: Understanding the complex series of events involved in cell-cell interactions during leukocyte transendothelial migration is a prerequisite for designing novel therapies to treat clinical conditions in which an inappropriate inflammatory response leads to disease. A discussion is provided of recent developments in the molecular regulation of leukocyte recruitment.

 

 

 

 

The cell adhesion molecule VCAM-1 plays an important role in regulation of inflammation in atherosclerosis, asthma, inflammatory bowel disease, or transplantation. VCAM-1 activates endothelial cell NADPH oxidase and this oxidase activity is required for VCAM-1-dependent lymphocyte migration. We previously reported that a mouse microvascular endothelial cell line promotes lymphocyte migration that is dependent on VCAM-1 but not on other known adhesion molecules. Here we have investigated the signaling mechanisms underlying VCAM-1 function. Lymphocyte binding to VCAM-1 on the endothelial cell surface activated an endothelial cell calcium flux that could be inhibited with anti-alpha 4-integrin and mimicked by anti-VCAM-1-coated beads. VCAM-1 stimulation of calcium responses could be blocked by an intracellular calcium mobilization inhibitor, a calcium channel inhibitor, or a calcium chelator, resulting in the inhibition of NADPH oxidase activity. Addition of ionomycin overcame the calcium channel blocker-suppressed, VCAM-1-stimulated NADPH oxidase activity, but could not reverse the inhibitory effect imposed by intracellular calcium blockage, indicating that both intracellular and extracellular calcium mobilizations are required for VCAM-1-mediated activation of NADPH oxidase. Furthermore, VCAM-1 specifically activated the Rho family GTPase Rac1 and VCAM-1 activation of NADPH oxidase was blocked by a dominant negative Rac1. Thus, VCAM-1 stimulates the mobilization of intracellular and extracellular calcium and Rac1 activity which are required for the activation of NADPH oxidase.