Adhesins of Leptospira interrogans Mediate the Interaction to Fibrinogen and Inhibit Fibrin Clot Formation In Vitro

Abstract

We report in this work that Leptospira strains, virulent L. interrogans serovar Copenhageni, attenuated L. interrogans serovar Copenhageni and saprophytic L. biflexa serovar Patoc are capable of binding fibrinogen (Fg). The interaction of leptospires with Fg inhibits thrombin- induced fibrin clot formation that may affect the haemostatic equilibrium. Additionally, we show that plasminogen (PLG)/plasmin (PLA) generation on the surface of Leptospira causes degradation of human Fg. The data suggest that PLA-coated leptospires were capable to employ their proteolytic activity to decrease one substrate of the coagulation cascade. We also present six leptospiral adhesins and PLG- interacting proteins, rLIC12238, Lsa33, Lsa30, OmpL1, rLIC11360 and rLIC11975, as novel Fg-binding proteins. The recombinant proteins interact with Fg in a dose-dependent and saturable fashion when increasing protein concentration was set to react to a fix human Fg concentration. The calculated dissociation equilibrium constants (KD) of these reactions ranged from 733.3±276.8 to 128±89.9 nM for rLIC12238 and Lsa33, respectively. The interaction of recombinant proteins with human Fg resulted in inhibition of fibrin clot by thrombin-catalyzed reaction, suggesting that these versatile proteins could mediate Fg interaction in Leptospira. Our data reveal for the first time the inhibition of fibrin clot by Leptospira spp. and presents adhesins that could mediate these interactions. Decreasing fibrin clot would cause an imbalance of the coagulation cascade that may facilitate bleeding and help bacteria dissemination

Leptospira and Leptospirosis

“…In vitro, Leptospira-bound plasmin degrades ECM components such as fibronectin (Vieira et al 2009) and human fibrinogen , and may activate host matrix metalloproteases which in turn could contribute to tissue degradation. Plasmin-coated leptospires also crossed human umbilical vein epithelial cell monolayers more efficiently than normal leptospires, although the precise mechanism was not investigated (Vieira et al 2013). Taken together these data suggest that surface-bound plasmin may facilitate crossing of ECM and tissue barriers and degradation of fibrin clots by leptospires, resulting in dissemination throughout the host.…”

“…Plasmin-coated leptospires also crossed human umbilical vein epithelial cell monolayers more efficiently than normal leptospires, although the precise mechanism was not investigated (Vieira et al 2013). As found in other pathogens (Lähteenmäki et al 2001), leptospires may up regulate activators of plasminogen in host cells (Vieira et al 2013). As found in other pathogens (Lähteenmäki et al 2001), leptospires may up regulate activators of plasminogen in host cells (Vieira et al 2013).…”

“…Transcytosis across human umbilical vein endothelial cell (HUVEC) monolayers was enhanced when leptospires were coated with plasminogen or plasmin, suggesting a proteolytic mechanism (Vieira et al 2013). Transcytosis across human umbilical vein endothelial cell (HUVEC) monolayers was enhanced when leptospires were coated with plasminogen or plasmin, suggesting a proteolytic mechanism (Vieira et al 2013).…”

Decrease in antithrombin III and prothrombin serum levels contribute to coagulation disorders during leptospirosis

“…We have recently demonstrated that leptospires bind human fibrinogen (Oliveira et al, 2013) and human thrombin (Fernandes et al, 2015), reducing fibrin clot formation and possibly interfering with the rheological properties of the clot, a characteristic that might add to the haemorrhagic symptoms in severe leptospirosis. Vieira et al (2009) have reported that pathogenic leptospires are able to bind plasminogen (PLG) and subvert the host machinery to convert it into its active form, plasmin (PLA), the main fibrinolytic enzyme.…”

“…Vieira et al (2009) have reported that pathogenic leptospires are able to bind plasminogen (PLG) and subvert the host machinery to convert it into its active form, plasmin (PLA), the main fibrinolytic enzyme. PLA-bound leptospires are able to degrade fibrinogen (Oliveira et al, 2013).…”