Discovery of anovel human T cell subtype possibly modulating adaption to atherosclerotic vascular occlusion

Atherosclerosis-related diseases remain the major cause of mortality and morbidity in the entire world population, imposing a huge burden for the healthcare systems. Atherosclerotic lesion formation causes the gradual obstruction of arterial vessels by inflammatory mechanisms. While traditional CAD risk factors (age, gender, smoking, diabetes, elevated cholesterol and blood pressure), as well as genetic risk factors are highly predictive of the onset atherosclerosis, mortality and morbidity is determined by clinical events, i.e. the occurrence of ischemic tissue damage, which are not well predicted by traditional CAD risk factors. One of these determinants of tissue ischemia is the presence of collateral arteries bypassing the obstructed vessel. 

To date, this phenomenon remains incompletely understood and, as a result, attempts to impact on collateral artery formation therapeutically have not had any breakthroughs, yet.  

In order to target this open question, we have recruited a cohort of PAD patients with proven high-grade arterial stenoses of the leg or groin arteries but no ischemic walking pain. In order to find these patients, we have performed large-scale bed-side ankle-brachial-index (ABI) measurement. In the LUERPAD-Immuno-study conducted at the University Heart Centre Luebeck, we have matched these patients with respect to gender and age with patients with PAD and typical symptoms. We characterized both groups clinically in-depth and found them to be very similar except for a large difference in pain-free walking distance. We hypothesized that lack of symptoms might be due to a better capacity for collateral artery growth in asymptomatic PAD patients. 

 

In order to test the hypothesis, if a specific immune signature might be detectable in asymptomatic PAD patients, we have performed deep single cell RNA sequencing on PBMCs of these patients (6 vs 6) using the BAD Rhapsody system. This analysis revealed a T cell subtype, which occurred uniformly in asymptomatic but not in symptomatic patients. This subtype displayed strong expression of GABBR2, component of the neuronal GABA receptor B. Indeed, GABBR2 was the strongest differentially expressed gene comparing both groups overall. These GABBR2+ T cells (GENATs) displayed characteristics of naïve T cells and we were able to map them onto double-negative T cells in a difference cohort of atherosclerotic patient’s PBMCs. Apart from this, the cell type (approx.. 1% of PBMCs in asymptomatic patients) featured unique characteristics hinting at a specific compensation mechanism in asymptomatic PAD patients. We were able to identify these cells in spectral flow cytometry and describe their surface marker expression characteristics. Furthermore, we could verify these findings from single cell transcriptomics using the other half of our cohort not included in our scRNAseq study and flow cytometric cell sorting as well as droplet-based qPCR. 

 

Overall, GENATs may be a novel cell type involved in adaption to vascular occlusion. This study shows that translational approaches to clinical problems mustering state-of-the-art AI technologies can be very successful in opening up new routes for possible therapeutic interventions on the level of the immune system in cardiovascular patients.