Abstract
Purpose :
Lymphocytic infiltration into lacrimal gland (LG) is a hallmark of Sjögren’s Syndrome (SS), which causes a severe form of dry eye. The cellular changes that drive the initiation and progression of this autoimmune response in lacrimal gland acinar cells (LGAC) are not well established. We tested the hypothesis that elevated level of IFN-γ could independently elicit some of the characteristic changes in LGAC gene and protein expression that have been linked to disease.
Methods :
Pro-inflammatory cytokine levels in LG of male non-obese diabetic (NOD) mice, and their healthy control strain, male BALB/c mice, aged 12 weeks, were measured using the MSD V-plex assay. Mouse LGAC were isolated and cultured from C57BL/6 mouse LG and recombinant mouse IFN-γ (200 U/ml) was applied for 48 hr. Gene expression levels in isolated NOD and BALB/c LGAC and in cultured C57BL/6 LGAC with/without IFN-γ were determined by RT- and q-PCR. Isolation of LGAC from NOD and BALB/c mouse LG utilized laser capture microdissection. Indirect immunofluorescence was used to investigate changes in expression and distribution of proteins of interest.
Results :
Pro-inflammatory cytokine levels were significantly increased in NOD mouse LG relative to BALB/c mouse LG. Among these, IFN-γ level was dramatically increased (fold change=45, p=0.0205). Gene expression levels of CTSS (Relative Quantity (RQ)=9.7, p=0.0006), MHC II (RQ=5.428, p=0.0119), Ii (RQ=93.02, p=0.0366) and ICAM-1 (RQ=13.9, p<0.0001) were increased in NOD mouse LGAC. Increased protein expression of HLA-DMα was likewise revealed by indirect immunofluorescence. Gene expression levels of CTSS (RQ=2.872, p=0.2202), MHC II (RQ=21.86, p=0.014), Ii (RQ=75.48, p=0.0003) and ICAM-1 (RQ=8.417, p=0.0003) were similarly increased in cultured LGAC treated with IFN-γ. In contrast, gene expression of Rab3D was deceased in both NOD mouse LGAC (RQ=0.6994, p=0.0006) and cultured LGAC (RQ=0.498, p=0.006) treated with IFN-γ.
Conclusions :
These findings support our hypothesis that increased IFN-γ level can promote increased expression and activity of CTSS and other proteins involved in antigen presentation, and reduce Rab3D expression, changes seen in murine models of SS and/or in SS patients. Further studies will determine whether the expressed antigen-presentation machinery enables LGAC to function as antigen-presenting cells in disease.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.