Novel adaptation of brush cytology technique for short-term primary culture of squamous and Barrett's esophageal cells

Background: Esophageal disease research is impeded by a paucity of nonmalig nant cell lines. Furthermore, culture primary esophageal cells is difficult because of a lack of cell adhesion and contamination. The aim of this stud y was to develop a short-term culture method to facilitate cell physiologic studies by using primary esophageal cells. Methods: By using a cytology brush, squamous and Barrett's epithelial cells were obtained from the esophagus of patients undergoing upper endoscopy. C ells were brushed onto chamber slides and allowed to adhere to the surface. Primary culture media was then added and cells were maintained at 37 degre esC for up to 72 hours. Cell yield and viability were calculated after tryp an blue staining. For cell physiologic studies, cells were loaded with pH-s ensitive dye BCECF-AM and intracellular pH measured on a dual excitation fl uorescence microscope after an ammonium chloride prepulse. At the end of th e physiologic experiments, cells were fixed in methanol and acetone and the cell type was verified with cytokeratin immunocytochemistry. Results: Viable human esophageal cells were maintained in culture for up to 72 hours. The cells extruded trypan blue, and BCECF-AM was cleaved to BCEC F by an intact cell membrane and permitted intracellular pH measurements. T he epithelial cell origin of the cells was confirmed by cytokeratin stainin g. There was no contamination over the culture period and no overgrowth by lymphocytes or fibroblasts. Conclusions: This novel adaptation of brush cytology technique enables shor t-term culture of esophageal cells for in vitro studies. This rapid, simple primary culture technique is suitable for endoscopy and does not require t he immediate, time-consuming laboratory techniques.