<p>Sjögren’s syndrome (SS) is an autoimmune disease characterized by dysfunction of exocrine glands. Pilocarpine is used as a symptomatic treatment for xerostomia, and chronic administration has been shown to increase salivary secretion; however, the underlying molecular mechanisms remain unclear. This study examined the effects of pilocarpine administration twice on salivary gland function in non-obese diabetic (NOD) mice, an animal model of SS. Pilocarpine was administered to 16-week-old NOD and wild-type (ICR) mice once a week for 2 weeks, and saliva was collected. Salivary volume, amylase activity, and the expression of aquaporin-5 (AQP5), connective tissue growth factor (Ctgf), and serum-and glucocorticoid-regulated kinase-1 (Sgk1) were examined. The salivary volume in ICR mice was significantly increased following the second administration, but not in NOD mice. No significant differences were observed in the total salivary amylase activity in either ICR or NOD mice. Western blot analysis demonstrated higher AQP5 expression in the membrane fractions and significantly lower expression in the cytosolic fractions of the submandibular glands following pilocarpine administration twice in both ICR and NOD mice, although the total AQP5 protein expression levels did not differ between the two types of mice. Gene expression analysis showed that Ctgf and Sgk1 were significantly upregulated in ICR mice following pilocarpine administration twice, but not in NOD mice. These findings suggest that the mechanism underlying the enhanced gene expression of Ctgf and Sgk1 following pilocarpine administration twice is impaired in NOD mice and that the expression of these genes may be associated with pilocarpine-induced salivary secretion.</p>