Acute salivary gland hypofunction in the duct ligation model in the absence of inflammation

Abstract

Objective: The commonly associated aetiology of salivary gland inflammation and salivary hypofunction has led to the widely held belief that inflammation causes salivary gland hypofunction. Indeed, our own recent study seemed to support this contention. Here, we tested the hypothesis that, in an acute duct ligation model, eliminating inflammation the submandibular gland would recover normal function.

Materials and methods: Ligation of the rat submandibular gland excretory duct for 24 h was used to induce inflammation and salivary gland hypofunction. A group of duct ligated rats was compared with a second group given dexamethasone, on the day of duct ligation. Twenty-four hours later salivary gland function was assessed and salivary glands were collected.

Results: Histology and myeloperoxidase activity assay revealed a profound decrease in inflammatory cell infiltration of ligated glands from rats given dexamethasone, compared with ligated glands in the absence of dexamethasone. Salivary flow rate evoked by methacholine was decreased (P < 0.01) by approximately 56% (ligated vs control, 79 +/- 9 microl min(-1) g(-1)vs 177 +/- 11 microl min(-1) g(-1)) and salivary flow from ligated dexamethasone-treated and ligated glands was similar.

Conclusion: Despite eliminating the inflammatory reaction in the ligated gland, salivary hypofunction was not reversed, suggesting that other mechanisms must be at work in the ligation-induced salivary hypofunction.

Figure 1

Effects of 24-h duct ligation with and without the presence of dexamethasone on submandibular gland weight and acinar cell size, stored acinar cell peroxidase and glandular protein content. Results were compared using paired and unpaired Student's t test and P < 0.05 considered significant. (a) Mean submandibular gland weight of 24-h ligated gland is increased comparing with control glands (n = 7). In contrast, dexamethasone-treated ligated glands are smaller than respective contralateral control (n = 7) and control and ligated untreated submandibular glands. (b) The mean area of acini from 24-h duct ligated glands and dexamethasone-treated ligated glands shows a reduction in the size of acini in the ligated glands (n = 100), and dexamethasone-treated ligated glands (n = 100), when compared with contralateral controls, (n = 100). Acinar size of control dexamethasone-treated glands was bigger than acini of control glands (unpaired Student's t test). (c) Secretory peroxidase activity from submandibular gland homogenates. In the dexamethasone-treated ligated group, after 24 h, the activity of peroxidase was significantly decreased, n = 5. In the 24-h duct ligation group, the difference was not significant, n = 6. (d) Total protein from submandibular gland homogenates. In the dexamethasone-treated ligated glands (n = 7), total protein content decreased compared with control dexamethasone-treated and untreated glands

Figure 2

Morphology of stimulated submandibular gland. In the top row, H&E staining for general morphology and in the lower row, AB/PAS reagent staining showing salivary glycoprotein containing secretory granules. (a) and (d) Dexamethasone-treated control submandibular gland. (b) and (e) Dexamethasone-treated ligated submandibular gland. (c) and (f) Ligated submandibular gland. (a) and (d) Illustrate a normal submandibular gland histological architecture with abundant closely packed acini (a) and large spaced, different sized ducts (d). (b) Contains a condensed stroma with some atrophic acini (arrowheads) and shows a marked reduction in inflammatory cells compared with (c); arrows on (c) indicate inflammatory cells. (e) and (f) Contain fewer acinar cell secretory granules than (d). All micrographs are ×125 magnification. Scale bars = 20 μm

Figure 3

Effect of 24-h duct ligation with and without the presence of dexamethasone on myeloperoxidase activity. Myeloperoxidase activity after 24-h duct ligation is greatly increased, compared with control, n = 6. Dexamethasone dramatically reduced myeloperoxidase activity in duct ligated glands, n = 5. Myeloperoxidase was decreased in the control dexamethasone-treated group compared with the control group (unpaired Student's t test)

Figure 4

Effects of 24-h duct ligation alone and in the presence of dexamethasone on submandibular gland function. (a) The mean flow rate of methacholine-evoked saliva (in units of microlitres per minute per gram gland wet weight) from the 24-h ligated glands was reduced by 78% compared with control glands (n = 7). In the dexamethasone-treated ligated glands, flow rate was reduced by 56% compared with contralateral control submandibular glands (n = 5) and by 55% compared with control untreated submandibular glands. Following duct ligation, salivary gland ducts were cannulated posterior to the site of clip ligation. (b) Salivary protein output induced by isoprenaline with methacholine is significantly reduced in both 24-h duct ligated (n = 4) and dexamethasone-treated ligated glands (n = 5). (c) Mean chloride concentration of methacholine-evoked saliva was increased in the 24-h ligated (n = 5) and dexamethasone-treated ligated glands (n = 6), when compared with the control untreated glands