Ciliated Epithelial Cell Differentiation at Air-Liquid Interface Using Commercially Available Culture Media

The human nasal epithelium contains basal stem/progenitor cells that produce differentiated multiciliated and mucosecretory progeny. Basal epithelial cells can be expanded in cell culture and instructed to differentiate at an air-liquid interface using transwell membranes and differentiation media. For basal cell expansion, we have used 3T3-J2 co-culture in epithelial culture medium containing EGF, insulin, and a RHO-associated protein kinase (ROCK) inhibitor, Y-27632 (3T3 + Y). Here we describe our protocols for ciliated differentiation of these cultures at air-liquid interface and compare four commercially available differentiation media, across nine donor cell cultures (six healthy, two patients with chronic obstructive pulmonary disease (COPD), and one with primary ciliary dyskinesia (PCD)). Bright-field and immunofluorescence imaging suggested broad similarity between differentiation protocols. Subtle differences were seen in transepithelial electrical resistance (TEER), ciliary beat frequency, mucus production, and the extent to which basal cells are retained in differentiated cultures. Overall, the specific differentiation medium used in our air-liquid interface culture protocol was not a major determinant of ciliation, and our data suggest that the differentiation potential of basal cells at the outset is a more critical factor in air-liquid interface culture outcome. Detailed information on the constituents of the differentiation media was only available from one of the four manufacturers, a factor that may have profound implications in the interpretation of some research studies

Poor adherence to antibiotic prescribing guidelines in acute otitis media—obstacles, implications, and possible solutions

Many countries now have guidelines on the clinical management of acute otitis media. In almost all, the public health goal of containing acquired resistance in bacteria through reduced antibiotic prescribing is the main aim and basis for recommendations. Despite some partial short-term successes, clinical activity databases and opinion surveys suggest that such restrictive guidelines are not followed closely, so this aim is not achieved. Radical new solutions are needed to tackle irrationalities in healthcare systems which set the short-term physician–patient relationship against long-term public health. Resolving this opposition will require comprehensive policy appraisal and co-ordinated actions at many levels, not just dissemination of evidence and promotion of guidelines. The inappropriate clinical rationales that underpin non-compliance with guidelines can be questioned by evidence, but also need specific developments promoting alternative solutions, within a framework of whole-system thinking. Promising developments would be (a) physician training modules on age-appropriate analgesia and on detection plus referral of rare complications like mastoiditis, and (b) vaccination against the most common and serious bacterial pathogens