A total of 24 Wistar rats were randomly divided into control and irradiation groups. The irradiation groups were further divided into 1, 4, and 12 weeks groups after irradiation. One-time 20 Gy irradiation was given to the SMG area on the experimental side of the irradiation group. At 1, 4, and 12 weeks after irradiation, the secretion of SMGs was measured using the Schirmer's test. The pathological changes in the gland tissues were observed under light microscopy after hematoxylin?eosin (HE) staining. The changes in the TJ ultrastructure were observed under transmission electron microscopy. The immunofluorescence staining and Western blot were used to detect the expression levels of muscarinic acetylcholine M3 receptor, aquaporin 5 (AQP5), and claudin-4 protein.

At 1, 4, and 12 weeks after irradiation, the secretion of SMGs in the irradiation group was significantly decreased and lower than that in the control group (P<0.01). At 1 week, the interstitial edema was observed in SMG tissues. Nuclear pyknosis, decreased number of acinar cells, and small focal necrosis with inflammatory infiltration were also observed over time. However, these changes were most evident at 12 weeks after irradiation. In the irradiation group, the TJ ultrastructure of glands at different times appeared to be fuzzy, collapsed, and had decreased electron density. Moreover, the width of TJs was remarkably decreased (P<0.01). The expression levels of M3 and AQP5 were decreased in a time-dependent manner, and the fluorescence intensity was significantly reduced after irradiation. However, the expression levels and fluorescence intensity of claudin-4 were enhanced in different degrees.

The changes in the TJ structure, the upregulation of the claudin-4 expression, and the damage in the paracellular pathway were involved in the hyposecretion of SMGs after irradiation.