Objective. Disorganization of acinar cell apical microvilli and the presence of stromal collagen in the acinar lumen suggest that the labial salivary gland (LSG) barrier function is impaired in patients with Sjögren's syndrome. Tight junctions define cell polarity and regulate the paracellular flow of ions and water, crucial functions of acinar cells. This study was undertaken to evaluate the expression and localization of tight junction proteins in LSGs from patients with SS and to determine in vitro the effects of tumor necrosis factor ␣ (TNF␣) and interferon-␥ (IFN␥) on tight junction integrity of isolated acini from control subjects.Methods. Twenty-two patients and 15 controls were studied. The messenger RNA and protein levels of tight junction components (claudin-1, claudin-3, claudin-4, occludin, and ZO-1) were determined by semiquantitative reverse transcriptase-polymerase chain reaction and Western blotting. Tight junction protein localization was determined by immunohistochemistry. Tight junction ultrastructure was examined by transmission electron microscopy. Isolated acini from control subjects were treated with TNF␣ and IFN␥.Results. Significant differences in tight junction protein levels were detected in patients with SS. ZO-1 and occludin were strongly down-regulated, while claudin-1 and claudin-4 were overexpressed. Tight junction proteins localized exclusively to apical domains in acini and ducts of LSGs from controls. In SS patients, the ZO-1 and occludin the apical domain presence of decreased, while claudin-3 and claudin-4 was redistributed to the basolateral plasma membrane. Exposure of isolated control acini to TNF␣ and IFN␥ reproduced these alterations in vitro. Ultrastructural analysis associated tight junction disorganization with the presence of endocytic vesicles containing electron-dense material that may represent tight junction components.Conclusion. Our findings indicate that local cytokine production in LSGs from SS patients may contribute to the secretory gland dysfunction observed in SS patients by altering tight junction integrity of epithelial cells, thereby decreasing the quality and quantity of saliva.

Our observation that the proteolytic action of MMPs toward ECM macromolecules is increased in SS patients provides a rationale for understanding the dramatic changes in the structural organization observed in the basal lamina and apical surface of acini in these patients. The results provide new evidence that acinar and ductal cells from the LSGs of SS patients display a molecular potential, with increased capacity to markedly disorganize their ECM environment and, thus, damage their architecture and functionality.

Objective To determine the enzymatic activity and cellular localization of matrix metalloproteinases (MMPs) 2, 3, and 9 in labial salivary glands from patients with different degrees of severity of primary Sjögren's syndrome (primary SS). Methods Gelatinase activity was determined by zymography and quantified by densitometry. The specificity of MMPs was determined using protease inhibitors and chelators, as well as activators of the latent forms of these enzymes. The cellular localization of MMPs was carried out using monoclonal antibodies that recognize their latent and active forms. Results Labial glands from control subjects and patients showed gelatinase activity for MMP‐2 and MMP‐9. Activation studies revealed that both enzymes were predominantly present in their latent forms. The highest levels of MMP‐9 activity were detected in patients with severe, active, primary SS (except for patients with severe clinical symptoms for extended periods) and correlated with structural and functional glandular changes. MMP‐2 activity was almost the same in patients and controls. MMPs were detected by immunolocalization only in acinar and ductal cells and were homogeneously distributed throughout patients' glands. MMP‐2 and MMP‐9 expression paralleled their gelatinase activity. MMP‐3, detectable only with immunologic methods, was absent in control subjects but abundantly expressed in patients. Importantly, MMP protein levels in acinar and ductal cells were independent of either the presence or the proximity of mononuclear infiltrate cells. Conclusion MMP‐3 and MMP‐9 expression, as well as MMP‐9 catalytic activity, were increased in tissue samples from SS patients in a manner that correlated with the severity of the disease. Most important, increased MMP activity stemmed from exocrine epithelial cells and was not due to infiltrating lymphocytes. Thus, changes in salivary glands as a consequence of proteolysis may lead to severe glandular destruction.

Reduced sulfation of muc5b is linked to xerostomia in patients with Sjögren syndrome Alliende, C.; Kwon, Y.-J.; Brito, M.; Molina, C.; Aguilera, S.; Pérez, P.; Leyton, L.; Quest, A.F.G.; Mandel, U.; Veerman, E.C.I.; Espinosa, M.; Clausen, H.; Leyton, C.; Romo, R.; González, S.

Objective. To investigate remodeling of the basal lamina of labial salivary glands (LSGs) from patients with Sjögren's syndrome (SS) by analyzing the expression of specific components that participate in its assembly and attachment to acinar and ductal cells.Methods. Two groups of SS patients with similar levels of remnant glandular tissue but with low and high levels of interacinar fibrosis, respectively, were studied. The expression of laminin ␣1, ␣4, and ␥2 chains and nidogens was examined at the messenger RNA (mRNA) and protein levels. Nidogens 1 and 2 were also studied in situ by immunohistochemistry.Results. Increases in the amount of mRNA and protein for both the processed and unprocessed laminin ␥2-chain were more pronounced in patients with low interacinar fibrosis. Increases in the protein levels of laminin ␣1 and ␣4 chains were observed in patients with low interacinar fibrosis, but not in those with high interacinar fibrosis. Nidogen mRNA and protein levels were similar in SS patients and controls. Interestingly, high levels of nidogen degradation were observed in patients with low interacinar fibrosis. Nidogens were readily detected by immunofluorescence in the basal lamina of the capillaries and stroma in SS patients, but were less apparent in the basal lamina of the acini and ducts.Conclusion. These results suggest that the basal lamina of LSGs from patients with SS is undergoing active remodeling, such that alterations are less evident in patients who have advanced morphologic signs of the disease (high interacinar fibrosis). Nidogen proteolysis might account for the disorganization of the basal lamina that is typically observed in LSGs from SS patients, assuming that cleavage impairs their ability to crosslink type IV collagen and laminin networks.Sjögren's syndrome (SS) is a chronic disorder that leads to insufficient production of saliva and tears (1,2). Labial salivary glands (LSGs) from patients with SS show dramatic changes in the basal lamina of the acini and ducts, including loss of individual components and structural disorganization, as detected by morphologic analysis. These events favor invasion of the acini and ducts by inflammatory cells that trigger cytotoxic responses (3). However, acinar cell death is also promoted by detachment of the cells from their extracellular matrix (ECM), a process known as anoikis (4). Importantly, the latter process does not require the presence of mononuclear cells. Instead, matrix metalloproteinases (MMPs) secreted by the acinar and ductal cells themselves have been implicated; in particular, imbalanced ratios of MMP-9 to tissue inhibitor of meMs Pérez and Ms Brito