Salmonella-induced thrombi in mice develop asynchronously in the spleen and liver and are not effective bacterial traps

Abstract

Thrombosis is a frequent, life-threatening complication of systemic infection associated with multiple organ damage. We have previously described a novel mechanism of inflammation-driven thrombosis induced by Salmonella Typhimurium infection of mice. Thrombosis in the liver develops 7 days after infection, persisting after the infection resolves, and is monocytic cell dependent. Unexpectedly, thrombosis was not prominent in the spleen at this time, despite carrying a similar bacterial burden as the liver. In this study, we show that thrombosis does occur in the spleen but with strikingly accelerated kinetics compared with the liver, being evident by 24 hours and resolving rapidly thereafter. The distinct kinetics of thrombosis and bacterial burden provides a test of the hypothesis that thrombi form in healthy vessels to trap or remove bacteria from the circulation, often termed immunothrombosis. Remarkably, despite bacteria being detected throughout infected spleens and livers in the early days of infection, immunohistological analysis of tissue sections show that thrombi contain very low numbers of bacteria. In contrast, bacteria are present throughout platelet aggregates induced by Salmonella in vitro. Therefore, we show that thrombosis develops with organ-specific kinetics and challenge the universality of immunothrombosis as a mechanism to capture bacteria in vivo.

Platelets in Inflammation and Resolution

“…This term exemplifies the interaction of platelets with other immune cells, as well as plasmatic coagulation components, resulting in thrombus formation, possibly to protect the host organism and restrict the infection to the local environment, as has been nicely summarized by Gaertner and Massberg (35). This concept was recently challenged; in a model of Salmonella Typhimurium infection, formed thrombi contained only limited amounts of bacteria in vivo, contradicting the notion of a thrombus as a major bacterialcapturing site throughout the whole body (36,37). Rather, an organ or pathogen specificity was detected in the formation of thrombi in this study (37).…”

“…This concept was recently challenged; in a model of Salmonella Typhimurium infection, formed thrombi contained only limited amounts of bacteria in vivo, contradicting the notion of a thrombus as a major bacterialcapturing site throughout the whole body (36,37). Rather, an organ or pathogen specificity was detected in the formation of thrombi in this study (37). Mechanisms of platelet-leukocyte interaction.…”

Vascular surveillance by haptotactic blood platelets in inflammation and infection

“…In contrast, platelets recruited in inflammation employ a different strategy: they scan the mechanical properties of the substrate and follow two-dimensional haptotactic fibrin(ogen) gradients to optimize their position and to safeguard vascular microbreaches. Autonomous navigation of single platelets at sites of inflammation and infection is required to efficiently prevent inflammatory bleedings and bacterial invasion without the need of forming large platelet thrombi, which would run the risk of causing detrimental vessel occlusions 64 , 65 (Supplementary Fig. 7m ).…”

Mice Deficient in T-bet Form Inducible NO Synthase–Positive Granulomas That Fail to Constrain Salmonella

“…HRP activity was detected with SIGMA FAST 3-3′Diaminobenzidine Tablets, whereas alkaline phosphatase activity was detected using Naphtol AS-MX Phosphate and fast blue salt with levamisole (all from Sigma-Aldrich). Staining for immunofluorescence was performed as described ( 28 ). Briefly, sections were rehydrated in PBS (pH 7.4) and incubated with primary Abs in the dark at room temperature for 40 min.…”

“…At least 100 foci were counted for each section, and the average number of CD3 + cells per focus is reported. Numbers of Salmonella per unit of section was quantified by point counting as described previously (28). Median fluorescence intensity (MFI) was obtained by selecting at least 200 foci per liver section per time-point per condition across multiple mice.…”