672. The Effect of Serine Protease Inhibitor 6 (SPI-6) Gene Transfer on a Mouse Model of Sjögren's Syndrome

Abstract

Top of pageAbstract Sj|[ouml]|gren's syndrome (SS) is an autoimmune disease characterized by a focal and diffuse lymphoid cell infiltration into the salivary and lacrimal glands. This chronic immune cell activation leads to reduced secretory function with symptoms of xerostomia (dry mouth) and keratoconjunctivitis sicca (dry eyes). It has been hypothesized that these sequelae are due to apoptosis of the exocrine epithelial cells. Apoptosis can be induced by cytotoxic T-lymphocytes (CTLs), which contain the enzyme Granzyme B (GrB) in cytoplasmic granules. GrB initiates a caspase-independent apoptosis pathway and is inhibited by serine proteinase inhibitor 6 (SPI-6) in the mouse. In this study, we evaluated the effect of transgenic SPI-6 produced in salivary glands after gene transfer to non-obese diabetic (NOD) mice, which develop exocrine gland infiltrates and are a commonly used disease model for SS. We constructed two serotype 2 rAAV vectors, rAAVSPI-6 and rAAVLacZ (encoding |[beta]|-galactosidase; vector control) using the CMV promoter. rAAVs (1010 particles/gland) were administered directly into both submandibular glands of 8-week old female NOD mice (n=8) via retroductal delivery. Salivary flow rates were determined before vector delivery and at time of sacrifice (16-weeks old). Serum glucose levels and weight were measured weekly and diabetic mice (serum glucose|[ge]|400 mg/dL) were treated with insulin. After sacrifice, submandibular glands were harvested and analyzed for inflammatory infiltrates (focus scores), as well as SPI-6 expression and cytokine profile [mouse interleukins (IL)-2, 4, 6, 12, 18, interferon (IFN)-|[gamma]|, tumor necrosis factor-|[alpha]| and RANTES] in aqueous gland extracts. At 8 weeks, salivary flow rates were not different between mice assigned to the rAAVSPI-6 and rAAVLacZ groups. In contrast, at 16 weeks flow rates were significantly higher in the rAAVSPI-6-treated mice (p=0.018). Submandibular gland IL-10 levels were significantly increased (p=0.009), while IFN-|[gamma]| levels were significantly decreased (p=0.029), in the rAAVSPI-6 mice compared to the rAAVLacZ group. There were no differences in focus scores, serum glucose levels or animal weights between the two treatment groups at 16 weeks. Overall, these results suggest that SPI-6 gene transfer may be beneficial in limiting the autoimmune exocrinopathy of NOD mice.