Objective: To improve cell survival and differentiation of freshly isolated salivary cells using mechanical dispersion.
Methods: PG and SMG were extracted from 5 week old C57/BL6 mice. PG and SMG cells were freshly isolated using mechanical dispersion then cultured on GFR-MG. Immunofluorescence studies were performed to visualize actin ring formation and cell polarization. Western blot analyses were used to detect survival and proliferation rates (active caspase-3 and PCNA) as well as secretory protein expression (amylase and VAMP-2). Intracellular calcium measurements were used to determine carbachol-induced intracellular calcium release. Electron microscopy was performed to examine the functionality of secretory granules.
Results: Cell survival rates for PG cells reached up to three days, whereas SMG cells were able to survive beyond two weeks. PG cells formed single lumens while SMG cells formed multiple interconnected lumens. Intracellular calcium signaling was detected in PG up to three days while SMG was detected up to two weeks. Expression of amylase and VAMP-2 was observed in both cell types
Conclusions: These results indicate that using mechanical dispersion for cell isolation allows for higher survival rates and acinar formation. SMG cells exhibited higher cell survival and differentiation rates than PG. Future studies characterizing the different cell types present in these cultures will be necessary to better understand why SMG survive for longer periods of time.