Endothelial Exosome Plays a Functional Role during Rickettsial Infection.

Kamil Khanipov,Jiani Bei,Zhengchen Su,Jie Xiao,Angelo Gaitas,Changcheng Zhou,Bin Gong,Tais B Saito,Yuan Qiu,Alexandra Drelich,Yakun Liu,Georgiy Golovko,Yang Jin,Qing Chang

doi:10.1128/mbio.00769-21

Abstract

ABSTRACTSpotted fever group rickettsioses (SFRs) are devastating human infections. Vascular endothelial cells (ECs) are the primary targets of rickettsial infection. Edema resulting from EC barrier dysfunction occurs in the brain and lungs in most cases of lethal SFR, but the underlying mechanisms remain unclear. The aim of the study was to explore the potential role of Rickettsia-infected, EC-derived exosomes (Exos) during infection. Using size exclusion chromatography (SEC), we purified Exos from conditioned, filtered, bacterium-free media collected from Rickettsia parkeri-infected human umbilical vein ECs (HUVECs) (R-ECExos) and plasma of Rickettsia australis- or R. parkeri-infected mice (R-plsExos). We observed that rickettsial infection increased the release of heterogeneous plsExos, but endothelial exosomal size, morphology, and production were not significantly altered following infection. Compared to normal plsExos and ECExos, both R-plsExos and R-ECExos induced dysfunction of recipient normal brain microvascular ECs (BMECs). The effect of R-plsExos on mouse recipient BMEC barrier function is dose dependent. The effect of R-ECExos on human recipient BMEC barrier function is dependent on the exosomal RNA cargo. Next-generation sequencing analysis and stem-loop quantitative reverse transcription-PCR (RT-qPCR) validation revealed that rickettsial infection triggered the selective enrichment of endothelial exosomal mir-23a and mir-30b, which potentially target the endothelial barrier. To our knowledge, this is the first report on the functional role of extracellular vesicles following infection by obligately intracellular bacteria.

Highlights

Spotted fever group rickettsioses (SFRs) are devastating human infections
size exclusion chromatography (SEC), we isolated small Extracellular vesicles (EVs) (50 to 150 nm) from rickettsia-infected mouse plasma and human umbilical vein ECs (HUVECs) from culture media; both were passed through two 0.2-mm filters
Quantitative real-time PCR validated that no rickettsial DNA copies were detected in either R-plsExos isolated from R. australis- or R. parkeri-infected mice infected with 2 50% lethal doses (LD50) of bacteria [80, 84,85,86,87] on day 4 postinfection (p.i.) or the R-ECExos that were purified 72 h p.i. from R. parkeri-infected HUVECs [6] using a multiplicity of infection (MOI) of 10 (Fig. S1)

Summary

Introduction

Spotted fever group rickettsioses (SFRs) are devastating human infections. Vascular endothelial cells (ECs) are the primary targets of rickettsial infection. Next-generation sequencing analysis and stem-loop quantitative reverse transcription-PCR (RT-qPCR) validation revealed that rickettsial infection triggered the selective enrichment of endothelial exosomal mir-23a and mir-30b, which potentially target the endothelial barrier. To our knowledge, this is the first report on the functional role of extracellular vesicles following infection by obligately intracellular bacteria. Next-generation sequencing analysis revealed that rickettsial infection triggered the selective enrichment of endothelial exosomal mir-23a and mir-30b, which potentially target the endothelial barrier A growing number of reports have established that many, if not all, of the effects of EVs are mediated by microRNA [52, 55,56,57,58,59,60, 63] or tRNA fragment [61, 78] cargos, which remain functional to regulate cellular behaviors of the recipient cells [79]

Objectives

Methods

Results

Conclusion