Intermediate filament, laminin and integrin expression in lacrimal gland acinar cells: Comparison of an immortalized cell line to primary cells, and their response to retinoic acid

Purpose. The goal of this study was to characterize intermediate filament, integrin and laminin expression by rabbit lacrimal gland acinar cells in cu lture, to determine whether retinoic acid (RA) alters expression of these p roteins and to compare primary cells to an immortalized rabbit lacrimal gla nd acinar cell line using flow cytometric analysis. Methods. Primary cells, maintained in serum free medium, were exposed to 10 (-6)M retinoic acid for 24 hours. Immortalized cells were grown in defined medium with Nu-Serum and exposed to retinoic acid. Cells were labeled with monoclonal antibodies to cytokeratins (AE1, AE2, AE3, AE5, CK10/13, CK18), integrins (alpha(3), alpha(6), alpha(v), beta(1), beta(2), beta(2) and beta (4)), laminin, or vimentin and with FITC-conjugated secondary antibodies. C ells were analyzed for antigen expression by flow cytometry and immunocytoc hemistry. Results. Primary and immortalized cells expressed type I and type II epithe lial cytokeratins (AE1 and AE3), cytokeratin 18, and cytokeratin 3 (AE5) Bo th cell types were negative to AE2 and CK10/13. Primary and immortalized ce lls expressed vimentin in culture, with immortalized cells expressing this protein at higher levels. Lacrimal acinar cells appear to synthesize lamini n which was detected intracellularly in both cells types. Integrins alpha(6 ) (CD49f) and alpha(v) (CD51) were expressed by primary and immortalized ce lls. Expression of integrin alpha(6) was 10-fold higher in immortalized cel ls compared to primary cells. Retinoic acid increased integrin alpha(v) exp ression by primary and immortalized cells 1.3-fold and 3-fold, respectively , and caused a slight increase in integrin alpha(6) expression by primary c ells. Both cell types also expressed integrins beta(1),beta(2) and beta(3), but beta(4) was detected only in immortalized cells. Lacrimal acinar cells do not express integrin alpha(3). Conclusions. Expression of cytokeratins, laminin and integrins by primary a nd immortalized cells was similar, suggesting that the immortalized cell li ne is a good model for the study of lacrimal structure and function. Since retinoic acid up-regulated only integrin alpha(v), but not cytokeratins, th ese cells appear to be highly differentiated. Flow cytometry is a useful me thod for analysis of protein expression by lacrimal acinar cells.