Natural Killer (NK) cells are an innate immune cell type that has somewhat been overlooked in the context of systemic lupus erythematosus (SLE). NK cell activation is determined by the integration of inputs from a myriad of activating and inhibitory receptors. Previously, using Nanostring® gene expression technologies, we found our SLE cohort showed decreased gene expression of a number of these receptors (KLRC2, KLRC1, KLRB1, KLRF1, KLRG1, PRF1 and IL2RB), leading us to explore NK cells in SLE in more depth. We aimed to phenotype NK cells in SLE using high-dimensional flow cytometry.
To do this, we designed a 24-marker flow cytometry NK panel. Designing the panel, we accounted for antigen density and marker co-expression to reduce spillover and spread. Optimisation of the panel included titrating all the antibodies and testing them at different machine voltages to obtain optimal resolution of the populations. The panel was run 60 SLE patients and 29 healthy controls for comparison. FlowSOM clustering algorithm was used to define clusters of NK cells. Cluster proportions were compared between SLE patients and healthy controls. The median fluorescence intensity was used to compare the protein expression of the activating receptor (NKG2D), costimulatory receptors (CD244/CD226), transcription factors (Eomes/Tbet) and effector molecules (granzyme/perforin).
Analysis of this cohort was conducted using clustering algorithms such as FlowSOM. This analysis revealed several NK clusters to be significantly aberrant in the SLE patients. The protein expression of activating receptor NKG2D was reduced in the SLE cohort.
By deeply interrogating the NK cells in SLE using high-parameter flow cytometry, we found several NK subsets and markers significantly different in our SLE cohort compared to healthy controls. Future plans include investigating NK cell changes with disease course/activity, therapeutic response, and discovering potential drug targets for SLE.