Single-cell transcriptomics reveals a pivotal role of DOCK2 in Sjögren’s disease

Sjögren's disease (SjD) is an autoimmune condition characterized by the dysfunction of the salivary and lacrimal glands. The study aimed to decipher the pathogenic cell populations and their immunological pathways in the salivary glands. We further determined the therapeutic effect of inhibiting DOCK2 shared by novel clusters of CD8Cd8+ T cells in a SjD mouse model. Salivary glands of C57BL/6.NOD-Aec1/2 (51 weeks old, n=2 female, 2 male) and B6 (64 weeks old, n=2 female, 2 male) mice were explanted and digested in a buffer containing 1 mg/ml DNase (Sigma-Aldrich, St. Louis, MO) and 1 mg/ml Collagenase Type 4 (Worthington, Lakewood, NJ, USA) in RPMI (Lonza, Allendale, NJ) complete media (10% FBS, 2 mM L-glutamine, 0.05 mM β-mercaptoethanol). Tissues were placed in a MACS C tube (Miltenyi Biotec, San Diego, CA) for desiccation on GentleMACS V1.02 for a pulse of 38 seconds. After a 10-minute incubation at 37°C, the digest buffer was removed and placed into 4°C RPMI complete media. The process was repeated twice. Single-cell suspensions were centrifuged (2500 rpm, 10 min, 4°C) and resuspended in PBS for filtration through a 70-μm sterile cell strainer (Fisher, Pittsburgh, PA). After a wash with PBS, cells were resuspended again in PBS for lymphocyte isolation with Lympholyte-M cell separation media (Cedar Lane, Burlington, Canada) per the manufacturer's instructions. Single-cell suspensions were stained for DAPI, and live cells were sorted with a sorter (SH800S, Sony, San Jose, CA) into RPMI containing 10% FBS on ice for single-cell sequencing library preparation.