Highly Pathogenic PRRSV-Infected Alveolar Macrophages Impair the Function of Pulmonary Microvascular Endothelial
Weifeng Sun1, Weixin Wu1, Nan Jiang1
1Key Laboratory of Animal Epidemiology of Ministry of Agriculture and Rural Affairs, College of Veterinary Medicine, China Agricultural University, Beijing 100193, China.
Abstract:
The porcine reproductive and respiratory syndrome virus (PRRSV), especially the highly pathogenic strains, can cause serious acute lung injury (ALI), characterized by extensive hemorrhage, inflammatory cells and serous fluid infiltration in the lung vascular system. Meanwhile, the pulmonary microvascular endothelial cells (PMVECs) are essential for forming the air-blood barrier and keeping the water-salt balance to prevent leakage of circulating nutrients, solutes, and fluid into the underlying tissues. As well, they tightly regulate the influx of immune cells. To determine the possible relationship between the PMVECs' function changes and lung vascular permeability during PRRSV infection, the PMVECs were co-cultured with HP-PRRSV-inoculated primary pulmonary alveolar macrophages (PAMs) in transwell model, and then the RNA sequencing (RNA-seq) and comprehensive bioinformatics analysis were carried out to characterize the dynamic transcriptome landscapes of PMVECs. In total, 16,489 annotated genes were identified, with 275 upregulated and 270 downregulated differentially expressed genes (DEGs) were characterized at both 18 and 24 h post PRRSV inoculation. The GO terms and KEGG pathways analysis indicated that the immune response, metabolic pathways, cell death, cytokine-cytokine receptor interaction, viral responses, and apoptotic process are significantly regulated upon co-culture with PRRSV-infected PAMs. Moreover, according to the TERR and dextran flux assay results, dysregulation of TJ proteins, including CLDN1, CLDN4, CLDN8, and OCLN, is further confirmed to correlate with the increased permeability of PMVECs. These transcriptome profiles and DEGs will provide valuable clues for further exploring the roles of PMVECs in PRRSV-induced ALI in the future.
Insights
Highly pathogenic porcine reproductive and respiratory syndrome virus (PRRSV) infection disrupts pulmonary microvascular endothelial cells (PMVECs), increasing lung vascular permeability. This study reveals key gene expression changes in PMVECs during PRRSV-induced acute lung injury (ALI).
Area of Science:
- Veterinary Virology
- Pulmonary Pathology
- Molecular Biology
Background:
- Porcine reproductive and respiratory syndrome virus (PRRSV) causes significant economic losses in the swine industry, often leading to acute lung injury (ALI).
- Pulmonary microvascular endothelial cells (PMVECs) are critical for maintaining the integrity of the air-blood barrier and regulating vascular permeability.
Purpose of the Study:
- To investigate the functional changes in PMVECs during PRRSV infection.
- To identify molecular mechanisms underlying PRRSV-induced increases in lung vascular permeability.
Main Methods:
- Co-culture of PMVECs with PRRSV-infected pulmonary alveolar macrophages (PAMs) using a transwell model.
- RNA sequencing (RNA-seq) and comprehensive bioinformatics analysis to characterize PMVEC transcriptomes.
- Analysis of differentially expressed genes (DEGs), Gene Ontology (GO) terms, and KEGG pathways.
- Validation of tight junction protein dysregulation and PMVEC permeability using TERR and dextran flux assays.
Main Results:
- RNA-seq identified 16,489 annotated genes, with 275 upregulated and 270 downregulated DEGs in PMVECs at 18 and 24 hours post-PRRSV inoculation.
- Bioinformatics analysis revealed significant regulation of immune response, metabolic pathways, cell death, cytokine-cytokine receptor interaction, viral responses, and apoptotic processes.
- Dysregulation of tight junction proteins (CLDN1, CLDN4, CLDN8, OCLN) was confirmed, correlating with increased PMVEC permeability.
Conclusions:
- PRRSV infection significantly alters PMVEC gene expression, impacting crucial cellular functions.
- The identified DEGs and pathways provide insights into the molecular basis of PRRSV-induced ALI.
- PMVEC dysfunction, particularly the disruption of tight junctions, plays a key role in the increased lung vascular permeability observed during PRRSV infection.
