Sofia V. Andersson

Cytoskeletal participation in stimulated secretion and compensatory apical plasma membrane retrieval in lacrimal gland acinar cells.

One of the main functions of the lacrimal gland is the regulated secretion of tear fluid, rich in proteins and electrolytes, in response to signals provided through the sympathetic and parasympathetic nervous system. Since proper tear-fluid secretion is essential for maintenance of ocular health, a long-term focus of our laboratory has been to understand the molecular mechanisms governing regulated secretion in lacrimal acini. In particular, we have focused on the role of microtubules (MTs), actin-based microfilaments (MFs), and motor proteins associated with either filament system in the stimulated lacrimal acinar secretory response. MTs and MFs constitute two of the three major cytoskeletal filament systems in mammalian cells, the third system being the intermediate filaments.

Presence of α-and β-Integrin Subunits in Rabbit Lacrimal Gland Acinar Cells Cultured on a Laminin-Rich Matrix

Integrins play critical roles in the interactions between the extracellular matrix and the cell by directing a number of functions, e.g. cell adhesion, cytoskeletal organization, and signal transduction. It is our hypothesis that they are also involved in modulating the regulation of secretion by lacrimal gland acinar cells, either by direct interaction with intracellular signaling cascades, or through modulation of cytoskeletal organization. Relatively little is known about the integrins in the lacrimal gland, but our laboratory is presently surveying for potential candidates capable of modulating secretion by lacrimal acinar cell. We have previously reported the presence of α2 αv, β1 and β4 integrin subunits in cells obtained by a cytospin technique.1, 2 The present study is an expanded survey for integrin subunits using rabbit lacrimal gland acinar cells in primary culture on laminin coated multi-well slides.

Regulation of Lacrimal Gland Cell Secretion by Integrin ClELL Adhesion

Purpose. We have previously shown that acinar cells cultured on exogenously added laminin responds stronger to carbachol stimulation, compared with other ECM proteins evaluated. Expression of the integrin subunits 6 and 1, functioning as laminin receptors, has been confirmed in primary cultured acinar cells and lacrimal gland tissue by immunofluorescence and sequence analysis. The purpose of this study was to explore integrin regulation of secretion in rabbit lacrimal gland acinar cells. Methods. Single acinar cells were isolated from rabbit lacrimal glands and cultured without serum in Matrigel coated 48-well plates for 40 hrs. For secretion assays, cultured cells were incubated 30 min with functional 6 or 1 integrin antibodies, followed by 1 hr carbachol (Cch) (0.1 mM) stimulation. Integrin antibody induced secretory response was also monitored over time. Pre-incubation (30 min) with PKC, IP3-receptor, tyrosine kinase or phosphatase inhibitors were utilized to investigate intracellular signaling from integrin 6 and 1. Cell media was collected and analyzed for -hexosaminidase activity as a measure of total protein secretion. Results. A rapid secretory response was observed within 5 min after addition of antibodies against integrin subunits 6 or 1, with no change during residual 1.5 hrs. Cch induced an elevated secretion after pretreatment with the 1 integrin antibody, which could not be detected after 6 integrin antibody incubation. Inhibition of selected signaling molecules reduced the Cch response but only orthovanadate blocked the 1 integrin antibody induced secretion, not affecting the 6. Conclusions. These results demonstrate that cell adhesion through integrins regulates secretion from acinar cells. The fact that 6 integrin antibody incubation, but not 1, blocked the carbachol response suggests that each subunit utilizes separate signaling pathways to induce secretion. The results also indicate that secretion triggered by the 1 integrin antibody is generated through dephosphorylation events. Support: Kalmar Faculty Research Grant, STINT and the Swedish Knowledge Foundation EFFECT OF OPC-12759 ON EXPRESSION OF MUCIN GENES IN HUMAN CORNEAL EPITHELIAL CELLS. Akihiro Aoki, Hiroki Urashima, Kazuhiko Fujita, Tamotsu Takizawa, Satoshi Oshima. Ako Research Institute, Otsuka Pharmaceutical Co. Ltd., Hyogo, Japan