Changes in the Submandibular Salivary Gland Epithelial Cell Subpopulations During Progression of Sjögren's Syndrome‐Like Disease in the NOD/ShiLtJ Mouse Model

Abstract

Sjögren’s syndrome (SS), an autoimmune exocrinopathy, is associated with dysfunction of the secretory salivary gland epithelium, leading to xerostomia. The etiology of SS disease progression is poorly understood as it is typically not diagnosed until late stage. Since mouse models allow the study of disease progression, we investigated the NOD/ShiLtJ mouse to explore temporal changes to the salivary epithelium. In the NOD/ShiLtJ model, SS presents secondary to autoimmune diabetes, and SS disease is reportedly fully established by 20 weeks. We compared epithelial morphology in the submandibular salivary glands (SMG) of NOD/ShiLtJ mice with SMGs from the parental strain at 12, 18, and 22 weeks of age and used immunofluorescence to detect epithelial proteins, including the acinar marker, aquaporin 5, ductal cell marker, cytokeratin 7, myoepithelial cell marker, smooth muscle α-actin, and the basal cell marker, cytokeratin 5, while confirming immune infiltrates with CD45R. We also compared these proteins in the labial salivary glands of human SS patients with control tissues. In the NOD/ShiLtJ SMG, regions of lymphocytic infiltrates were not associated with widespread epithelial tissue degradation; however, there was a decrease in the area of the gland occupied by secretory epithelial cells in favor of ductal epithelial cells. We observed an expansion of cells expressing cytokeratin 5 within the ducts and within the smooth muscle α-actin+ basal myoepithelial population. The altered acinar/ductal ratio within the NOD/ShiLtJ SMG likely contributes to salivary hypofunction, while the expansion of cytokeratin 5 positive-basal cells may reflect loss of function or indicate a regenerative response.

FGF2-dependent mesenchyme and laminin-111 are niche factors in salivary gland organoids

“…If EpCAM + spheroids showed more than 30% (by area) of the proacinar marker AQP5 expression, they were considered to be an AQP5 + organoid. Positively stained areas (μm 2 ) for EpCAM, AQP5 or vimentin markers were quantified by using calibrated and thresholded images using ImageJ software (Abràmofff et al, 2005), as we performed previously (Gervais et al, 2015). Organoid diameter was measured from calibrated images with ImageJ software.…”

Par-1b is required for morphogenesis and differentiation of myoepithelial cells during salivary gland development

“…Immunohistochemistry was performed as previously described. 48 Briefly specimens were fixed in 10% neutral buffered saline and then moved to 70% ethanol until paraffin embedding. 0.5 mm sections were mounted on slides, deparaffinized, and immunostained with Smooth muscle a-actin, ECAD and DAPI.…”

“…Images were captured using an Olympus IX-81 microscope with a 20X Plan Apo 0.75 NA objective and processed as previously described. 48 Images were manually overlaid in Photoshop CS6 (13.0.1£64).…”

RARα and RARγ reciprocally control K5+ progenitor cell expansion in developing salivary glands

“…One SMG epithelial progenitor cell type is characterized by expression of cytokeratin 5 (K5), the K5 C population. Initially present in the majority of the cells in the placode, the K5 C population becomes ductally restricted, forming a basal layer in the large and intercalated ducts as development proceeds and in the adult 22,23 . K5 is a marker of basal epithelial progenitor cells in several organs, including the mammary 24 and the salivary 25 glands.…”