Selectins Influence Thrombosis in a Mouse Model of Experimental Deep Venous Thrombosis 1 Daniel Myers, Jr., D.V.M.,* , † ,2 Diana Farris, L.V.T.,* Angela Hawley, M.S.,* Shirley Wrobleski, B.S.,* Amy Chapman, B.S.,* Lloyd Stoolman, M.D., Ph.D.,‡ Randy Knibbs, Ph.D.,‡ Robert Strieter, M.D.,§ and Thomas Wakefield, M.D.* *Jobst Vascular Laboratory, Section of Vascular Surgery, Department of Surgery, †Unit of Laboratory Animal Medicine, and ‡Department of Pathology, University of Michigan, Ann Arbor, Michigan 48109; and §Department of Medicine, University of California at Los Angeles, Los Angeles, California 90024 Submitted for publication February 25, 2002 Background. This study characterizes venous throm- bosis in the mouse and examines the important role that the adhesion molecules P-selectin and E-selectin and the anti-inflammatory cytokine interleukin-10 (IL-10) play in the thrombotic process. Materials and methods. C57BL/6 (wild-type) mice in a natural history protocol (Phase I) and gene-targeted (KO) mice for P-selectin, E-selectin, P/E-selectin, and IL-10 in a follow-up protocol (Phase II) were studied. Inferior vena caval thrombosis was produced by liga- tion just below the renal veins, and mice were sacri- ficed and evaluated at various time points up to 12 days later. Results. Phase I: A significant increase in neutro- phils on day 2 and in monocytes on day 6 postthrom- bosis was found in ligated vs sham animals. An asso- ciated significant increase in vein wall P-selectin mRNA (6 h, day 2) and an increase in protein (6 h through day 6) were found, while E-selectin mRNA was significantly increased (day 2 through day 6), with a smaller increase in E-selectin protein. IL-10 mRNA increased significantly later (day 2 through day 9), with the values increasing progressively. A positive correlation existed (r  0.77) between neutrophils and thrombosis at day 2. Phase II: The E-selectin and P/E- selectin double-KO mice showed the least thrombus at day 2 vs wild-type clotted mice, P < 0.01. Additionally, P/E-KO mice demonstrated the lowest inflammatory cell extravasation into the vein wall at day 2. Conclusions. This study demonstrates an acute to chronic inflammatory response in the vein wall asso- ciated with venous thrombosis. Inhibition of selectins decreased thrombus formation. © 2002 Elsevier Science (USA) Key Words: venous thrombosis; selectins; mouse; in- ferior vena cava. INTRODUCTION Venous thrombosis is associated with a significant inflammatory response in the vein wall and thrombus [1]. This response augments the thrombotic response and is related to the detrimental vein wall and valvular changes that lead to chronic venous insufficiency [2]. We have noted in rat [3– 6] and primate [7–10] models of venous thrombosis that adhesion molecules and the natural anti-inflammatory cytokine interleukin-10 (IL- 10) are important in the thrombotic process. Selectins appear to be important for thrombus formation and inflammatory enhancement [4, 7–10], while IL-10 ap- pears to down-regulate not only inflammation [6] but also thrombus formation [5]. In the present study, we chose to evaluate, for the first time, the inflammatory response in a mouse model of venous thrombosis and compare this to our previous results. The advantages of such a model include the ability to study large numbers of animals and to use gene targeting of important ad- hesion molecules and cytokines to determine their im- portance to the thrombotic and inflammatory pro- cesses. This is a descriptive study to document the kinetics of selectin and IL-10 expression after venous thrombosis. Currently, such a natural history baseline in a mouse model of venous thrombosis has not been established. Additionally, we then determined the role Presented at the XVIII Congress of the International Society on Thrombosis and Haemostasis, Paris, July 9, 2001 1 This work was supported by NIH RO1 HL 63148 (T.W.W.). 2 To whom reprint requests should be addressed at 1500 E. Med- ical Center Drive, TC2210/0329, Ann Arbor, MI 48109. Fax: (734) 763-7307. E-mail: ddmyers@umich.edu. Journal of Surgical Research 108, 212–221 (2002) doi:10.1006/jsre.2002.6552 212 0022-4804/02 $35.00 © 2002 Elsevier Science (USA) All rights reserved.