Investigating transcriptional responses in Parascaris-challenged equine BAL cells

Sammanfattning

Parascaris spp. is a parasitic nematode infecting the domestic horse (Equus caballus), primarily affecting foals (<1 year old). In recent years, anthelminthic drug resistance has developed against all major classes of antihelminth drugs, posing a challenge to equine health. The Parascaris spp. life cycle involves larval migration from the small intestine to the liver and lungs before returning to the small intestine to mature. Infection cannot be reliably detected until approximately 80 days post infection, when eggs are detectable in the horse feces. It is largely unknown how the Parascaris larvae interacts with the host microenvironment, and earlier studies have not identified a clearly polarized adaptive immune response. The aim of this study was to characterize the pulmonary immune response following Parascaris spp. challenge at different time points, and to identify differentially expressed genes and cytokines, with the overarching goal to identify candidate gene expression biomarkers. Differently expressed genes were identified by RNA sequencing, from which seven genes of interest (IL1B, PTX3, MIP2B, SERPINB2, RNF152, SRD5A3, and CFAP298) were selected for validation. Gene set enrichment analysis was performed to identify overexpressed biological processes and pathways, and a protein-protein interaction network was constructed to identify hub genes. Bronchoalveolar lavage (BAL) cells were collected from Standardbred horses and infected with Parascaris spp. L2/L3 larvae, and relative gene expression was quantified using reverse transcription quantitative polymerase chain reaction. In addition, cytokines associated with different T helper cell mediated immune responses (IL-4, IL-5, IL-12, and IL-23) were analyzed. This study revealed that IL1B, MIP2B, and PTX3 are significantly upregulated, while CFAP298 is significantly downregulated in Parascaris spp. challenged BAL cells. Upregulation was observed both 6 hours and 24 hours post infection, while the downregulation of CFAP298 was detected only at 24 hours. Substantial variability in the gene expression between biological replicates (individuals) was observed, limiting their suitability as robust biomarkers. None of the analyzed cytokines showed any significant differential expression, suggesting the absence of a polarized T helper cell response. The gene set enrichment analysis revealed enrichment of immune related pathways associated with responses to bacteria, virus, and the humoral immune response. Together, the data suggests a non-specific activation and inflammatory pulmonary immune response when challenged with Parascaris spp. larvae.