Andes Virus Regulation of Cellular MicroRNAs Contributes to Hantavirus-Induced Endothelial Cell Permeability

Abstract

Hantaviruses infect human endothelial cells (ECs) and cause two diseases marked by vascular permeability defects, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). Vascular permeability occurs in the absence of EC lysis, suggesting that hantaviruses alter normal EC fluid barrier functions. ECs infected by pathogenic hantaviruses are hyperresponsive to vascular endothelial growth factor (VEGF), and this alters the fluid barrier function of EC adherens junctions, resulting in enhanced paracellular permeability. Vascular permeability and VEGF-directed responses are determined by EC-specific microRNAs (miRNAs), which regulate cellular mRNA transcriptional responses. miRNAs mature within cytoplasmic processing bodies (P bodies), and the hantavirus nucleocapsid (N) protein binds RNA and localizes to P bodies, suggesting that hantaviruses may modify miRNA functions within infected ECs. Here we assessed changes in EC miRNAs following infection by the HPS-causing Andes hantavirus (ANDV). We analyzed 352 human miRNAs within ANDV-infected ECs using quantitative real-time (RT)-PCR arrays. Fourteen miRNAs, including six miRNAs that are associated with regulating vascular integrity, were upregulated >4-fold following infection by ANDV. Nine miRNAs were downregulated 3-to 3,400-fold following ANDV infection; these included miR-410, involved in regulating secretion, and miR-218, which is linked to the regulation of EC migration and vascular permeability. We further analyzed changes in miR-126, an EC-specific miRNA that regulates vascular integrity by suppressing SPRED1 and PIK3R2 mRNAs. While miR-126 levels were only slightly altered, we found that SPRED1 and PIK3R2 mRNA levels were increased 10-and 7-fold, respectively, in ANDV-infected ECs but were unaltered in ECs infected by the nonpathogenic Tula hantavirus (TULV). Consistent with increased SPRED1 expression, we found that the level of phospho-cofilin was decreased within ANDV-infected ECs. Moreover, small interfering RNA (siRNA) knockdown of SPRED1 dramatically decreased the permeability of ANDV-infected ECs in response to VEGF, suggesting that increased SPRED1 contributes to EC permeability following ANDV infection. These findings suggest that interference with normal miRNA functions contributes to the enhanced paracellular permeability of ANDV-infected ECs and that hantavirus regulation of miRNA functions is an additional determinant of hantavirus pathogenesis.

Andes Virus Infection of Lymphatic Endothelial Cells Causes Giant Cell and Enhanced Permeability Responses That Are Rapamycin and Vascular Endothelial Growth Factor C Sensitive

“…HPS patients are acutely hypoxic (36,39,59,62,79), suggesting a link between pulmonary edema during HPS and enhanced endothelial cell VEGF-A responses (12,15,16,38,57,64,76). In fact, both HTNV and ANDV enhance VEGF-A-directed permeability responses, and inhibitors that antagonize this pathway block the hyperpermeability of hantavirus-infected BECs (28,29,34,35,63). Collectively, these findings tie altered VEGF-A responses following hantavirus infection to edema observed in HPS and HFRS patients.…”

“…Andes virus (ANDV) causes HPS, resulting in acute pulmonary edema and respiratory insufficiency (12,14,18,22,36,47,53,59,62,86). The means by which hantaviruses cause vascular leakage and edema are likely to be multifactorial in nature, and mechanisms by which hantaviruses alter fluid barrier properties of the vasculature are still being discovered (28,29,34,35,37,45,63,70,74). Tissue and organ edema are prominent findings in hantavirus patients, and blood vessel ECs (BECs) form a primary fluid barrier that normally restricts fluid egress into tissues and permits blood and fluid recirculation (1,19,80).…”

T Cells Are Not Required for Pathogenesis in the Syrian Hamster Model of Hantavirus Pulmonary Syndrome

“…Pathogenic hantaviruses bind to inactive ␤ 3 integrin conformations on endothelial cells, causing them to remain inactive and interfering with their normal function (20,22,23,63). Recently, this has been shown to result in dysregulation of VEGFR2 signaling, rendering infected endothelial cells hypersensitive to exogenous VEGF and increasing their permeability by altering the expression of genes that regulate endothelial cell migration and permeability and causing VE cadherin internalization, resulting in adherens junction disassembly (26,27,59). Notably, nonpathogenic hantaviruses infect endothelial cells via ␣ V ␤ 1 integrin, and this binding does not result in adherens junction disassembly.…”

A Stromal Cell-Derived Factor 1α Analogue Improves Endothelial Cell Function in Lipopolysaccharide-Induced Acute Respiratory Distress Syndrome

“…Genomic ablation of miR-126 has been shown to increase vascular permeability (16,17,19,21). MicroRNA-126 targets PIK3R2 and Spred-1 leading to stabilization of vascular endothelial (VE)-cadherin via AKT and ERK 1/2 signaling cascades (22,23). CTCE injections result in significantly increased plasma miR-126 levels in CLP-induced sepsis suggesting a potential mechanism by which it reduces endothelial permeability (12).…”

“…MicroRNA-126 is the most abundant miRNA expressed during EC differentiation. It has previously been shown to have pleiotropic effects during EC differentiation and its genetic absence is known to augment vascular permeability (22,31). A potential mechanism by which miR-126 may decrease vascular permeability includes its targeted inhibition of PIK3R2 (32).…”