Biology subjects

Shelite, T.

Publications and source records attributed to Shelite, T..

3 recordsLinked to original sources

bioRxiv2021

Host EPAC1 modulates rickettsial adhesion to vascular endothelial cells via regulation of ANXA2 Y23 phosphorylation

Recently we have identified that endothelial surface annexin A2 (ANAX2) functions as a receptor for spotted fever group rickettsial adhesin outer membrane protein B (OmpB), which binds to the endothelial cell (EC) surface. Moreover, we reported that intracellular cAMP receptor EPAC1 modulates ANXA2 tyrosine (Y) 23 phosphorylation, and inactivation of EPAC1 suppresses ANXA2 expression on the EC luminal surface by downregulating Y23 phosphorylation. Since we reported that EPAC1 plays a critical role in the initial step to successfully establish rickettsial infection of ECs, this work aims to answer the following: (a) What is the mechanism underlying the regulatory role of EPAC1 in ECs during the initial step of bacterial infection? (b) Is the EPAC1-ANXA2 signaling pathway involved in the regulation of rickettsial adhesion to ECs? In the present study, an established system that is anatomically-based and quantifies bacterial adhesion to ECs in vivo was combined with novel fluidic force microscopy (FluidFM) to dissect the functional role of the EPAC1-ANXA2 signaling pathway in rickettsiae-EC adhesion. We reveal that the deletion of the EPAC1 gene impedes rickettsial binding to endothelium in vivo. In addition, single living brain microvascular EC study that employs FluidFM and site-directed mutagenesis provides evidence that supports our finding that EPAC1 governs rickettsial adhesion to EC surfaces via regulation of ANXA2 Y23 phosphorylation.

pathology↗

bioRxiv2019

Annexin A2 depletion exacerbates the intracerebral microhemorrhage induced by acute rickettsia and Ebola virus infections

Intracerebral microhemorrhages (CMHs) are small foci of hemorrhages in the cerebrum. Acute infections induced by some intracellular pathogens, including rickettsia, can result in CMHs. Annexin a2 (ANXA2) has been documented to play a functional role during intracellular bacterial adhesion. Here we report that ANXA2-knockout (KO) mice are more susceptible to CMHs in response to rickettsia and Ebola virus infections, suggesting an essential role of ANXA2 in protecting vascular integrity during these intracellular pathogen infections. Proteomic analysis via mass spectrometry of whole brain lysates and brain-derived endosomes from ANXA2-KO and wild-type (WT) mice post-infection with R. australis revealed that a variety of significant proteins were differentially expressed, and the follow-up function enrichment analysis had identified several relevant cell-cell junction functions. Immunohistology study confirmed that both infected WT and infected ANXA2-KO mice were subjected to adherens junctional protein (VE-cadherin) damages. However, key blood-brain barrier (BBB) components, tight junctional proteins ZO-1 and occludin, were disorganized in the brains from R. australis-infected ANXA2-KO mice, but not those of infected WT mice. Similar ANXA2-KO dependent CMHs and fragments of ZO-1 and occludin were also observed in Ebola virus-infected ANXA2-KO mice, but not found in infected WT mice. Overall, our study revealed a novel role of ANXA2 in the formation of CMHs during R. australis and Ebola virus infections; and the underlying mechanism is relevant to the role of ANXA2-regulated tight junctions and its role in stabilizing the BBB in these deadly infections. Author SummaryTraditionally, spontaneous intracerebral microhemorrhages (CMHs) were defined as small foci of intracerebral hemorrhages. Such atraumatic CMHs are due to the rupture of a weak blood vessel wall. Infections complicating cerebrovascular accidents have been extensively investigated. However, the role of CMHs complicating infections, in particularly acute systemic infections, has been poorly explored. Population-based retrospective cohort studies suggest there are potentially more undiscovered cases of CMHs accompanying acute systemic infections. Given both the lack of an animal model and cellular/molecular pathophysiology of CMHs following acute systemic infections, there is an urgent need to increase our comprehensive understanding of acute infection-induced CMHs. Overall, our study revealed a novel role of annexin a2 (ANXA2) in the formation of CMHs during R. australis and Ebola virus infections; and the underlying mechanism is relevant to the role of ANXA2-regulated endothelial tight junctions and its role in stabilizing the blood-brain barrier in these deadly infections.

pathology↗

bioRxiv2019

Polarized Lung Inflammation and Tie2/Angiopoietin-Mediated Endothelial Dysfunction during Severe Orientia tsutsugamushi Infection

Orientia tsutsugamushi infection can cause acute lung injury and high mortality in humans; however, the underlying mechanisms are unclear. Here, we tested a hypothesis that dysregulated pulmonary inflammation and Tie2-mediated endothelial malfunction contribute to lung damage. Using a murine model of lethal O. tsutsugamushi infection, we demonstrated pathological characteristics of vascular activation and tissue damage: 1) a significant increase of ICAM-1, VEGFR2, and angiopoietin-2 (Ang2) proteins in inflamed tissues and lung-derived endothelial cells (EC), 2) a progressive loss of endothelial quiescent and junction proteins (Ang1, VE-cadherin/CD144, occuludin), and 3) a profound impairment of Tie2 receptor at the transcriptional and functional levels. In vitro infection of primary human EC cultures and serum Ang2 proteins in scrub typhus patients support our animal studies, implying endothelial dysfunction in severe scrub typhus. Flow cytometric analyses of lung-recovered cells further revealed that pulmonary macrophages (M{Phi}) were polarized toward an M1-like phenotype (CD80+CD64+CD11b+Ly6G-) during the onset of disease and prior to host death, which correlated with the significant loss of CD31+CD45- ECs and M2-like (CD206+CD64+CD11b+Ly6G-) cells. In vitro studies indicated extensive bacterial replication in M2-type, but not M1-type, M{Phi}s, implying the protective and pathogenic roles of M1-skewed responses. This is the first detailed investigation of lung cellular immune responses during acute O. tsutsugamushi infection. It uncovers specific biomarkers for vascular dysfunction and M1-skewed inflammatory responses, highlighting future therapeutic research for the control of this neglected tropical disease.\n\nAuthor SummaryScrub typhus is a life-threatening disease, infecting an estimated one million people yearly. Acute lung injury is the most common clinical observation; however, its pathogenic biomarkers and mechanisms of progression remain unknown. Here, we used a lethal infection mouse model that parallels certain aspects of severe scrub typhus, primary human endothelial cell cultures, and patient sera to define pathogenic biomarkers following Orientia tsutsugamushi infections. We found a significant increase in the levels of endothelial activation/stress markers (angiopoietins and ICAM-1) in infected mouse lungs and in patient sera, but a progressive loss of endothelium-specific Tie2 receptor and junction proteins (VE-cadherin), at severe stages of disease. These signs of vasculature disruption positively correlated with the timing and magnitude of recruitment/activation of proinflammatory M{Phi} subsets in infected lungs. Bacterial growth in vitro was favored in M2-like, but not in M1-like, M{Phi}s. This study, for the first time, reveals endothelial malfunction and dysregulated inflammatory responses, suggesting potential therapeutic targets to ameliorate tissue damage and pathogenesis.

microbiology↗