Abstract

Background
Pigeon fanciers are at increased risk of Hypersensitivity Pneumonitis (HP), a common interstitial lung disease (ILD). Connective tissue disease (CTD) is also associated with development of ILD, and CTD symptoms are common in pigeon fanciers without CTD. How this links to their risk of ILD is unclear. Single cell RNA sequencing (scRNAseq) can allow unbiased identification of relevant novel pathways.

Objectives
We aimed to map monocyte molecular heterogeneity among pigeon fanciers to test the hypothesis that distinct phenotypes associate with HP/ILD or CTD symptoms.

Methods
Blood monocytes from fanciers categorised as ‘no symptoms’ post pigeon exposure (n=15), previous acute HP/fibrosing ILD (HP/ILD)(n=10), symptoms of CTD (n=12) or ‘post exposure’ symptoms (n=15) were prepared for scRNA-seq. 130,000 cells (500 immune-gene panel) were analysed per subject.

Results
Following agnostic integration (SCTransform algorithm), 5 clusters in classic (CD14^hiCD16^neg), 3 in non-classic (CD14^loCD16^hi), and 4 in intermediate (CD14⁺CD16⁺) monocytes were identified across all groups. Differences in relative proportions were significant for those with CTD symptoms: an intermediate cluster (dominant gene IFITM2^pos) was higher, and a classic cluster (S100A12^hi) was lower compared to those without CTD symptoms. Among 15 differentially expressed genes GNAI2 was significantly upregulated (+2.7) in HP/ILD vs CTD.

Conclusions
GNAI2 encodes G-protein Gai2 which modulates GPCR chemokine receptor responses. Relative expression levels may determine recruitment of selective monocyte clusters to different tissue sites for local pathogenesis or resolution, and may be relevant for the evolution of disease in patients with HP.