Changing with the times: Pandemic spread of a virus in naïe populations can select for mutations that alter pathogenesis, virulence, and/or transmissibility. The ancestral form of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that emerged from China has now been largely replaced by strains containing the mutation D614G (Asp ⁶¹⁴ -to-Gly) in the viral spike protein. Hou et al. compared the characteristics of the new variant against the ancestral form in a series of experiments in human cells and animal models. The variant is better at infecting upper-airway epithelial cells and replicates in greater numbers than the ancestral virus. Evidence indicates modest, if any, significant changes to virulence in animal models. Therefore, the virus appears to have evolved for greater transmissibility in humans rather than for greater pathogenicity. The mutation renders the new virus variant more susceptible to neutralizing antisera without altering the efficacy of vaccine candidates currently under development. Science , this issue p. 1464

Well-differentiated primary human bronchial epithelial (HBE) cell cultures are vital for cystic fibrosis (CF) research, particularly for the development of cystic fibrosis transmembrane conductance regulator (CFTR) modulator drugs. Culturing of epithelial cells with irradiated 3T3 fibroblast feeder cells plus the RhoA kinase inhibitor Y-27632 (Y), termed conditionally reprogrammed cell (CRC) technology, enhances cell growth and lifespan while preserving cell-of-origin functionality. We initially determined the electrophysiological and morphological characteristics of conventional versus CRC-expanded non-CF HBE cells. On the basis of these findings, we then created six CF cell CRC populations, three from sequentially obtained CF lungs and three from F508 del homozygous donors previously obtained and cryopreserved using conventional culture methods. Growth curves were plotted, and cells were subcultured, without irradiated feeders plus Y, into air–liquid interface conditions in nonproprietary and proprietary Ultroser G–containing media and were allowed to differentiate. Ussing chamber studies were performed after treatment of F508 del homozygous CF cells with the CFTR modulator VX-809. Bronchial epithelial cells grew exponentially in feeders plus Y, dramatically surpassing the numbers of conventionally grown cells. Passage 5 and 10 CRC HBE cells formed confluent mucociliary air–liquid interface cultures. There were differences in cell morphology and current magnitude as a function of extended passage, but the effect of VX-809 in increasing CFTR function was significant in CRC-expanded F508 del HBE cells. Thus, CRC technology expands the supply of functional primary CF HBE cells for testing CFTR modulators in Ussing chambers.

Airway epithelial stem cells are not well characterized. To examine clonal growth potential, we diluted single, viable B6.129S7-Gtrosa26 (Rosa26) mouse tracheal epithelial cells that constitutively express β-galactosidase into non-Rosa26 cells in an air-liquid interface cell culture model; 1.7% of the cells formed colonies of varying size, and, on average, 0.1% of the cells formed large colonies. Thus only a small subset of cells displayed progenitorial capacity suggestive of stem or early transient amplifying cells. Prior studies identified cells with high keratin 5 (K5) promoter activity in specific niches in the mouse trachea and these cells corresponded to the location of bromodeoxyuridine label-retaining cells, thought to be stem cells (Borthwick DW, Shahbazian M, Todd KQ, Dorin JR, and Randell SH, Am J Respir Cell Mol Biol: 24: 662–670, 2001). To explore the hypothesis that stem cells were present in the K5-expressing compartment, we created transgenic mice in which enhanced green fluorescent protein (EGFP) was driven by the K5 promoter. These mice expressed EGFP in most basal cells of the body including a subset of tracheal basal cells apparently located in positions similar to previously identified stem cell niches. Flow cytometrically purified EGFP-positive cells had an overall colony-forming efficiency 4.5-fold greater than EGFP-negative cells, but the ability to generate large colonies was 12-fold greater. Thus adult mouse tracheal epithelial cells with progenitorial capacity sufficient to generate large colonies reside in the basal cell compartment. These studies are a first step toward purification and characterization of airway epithelial stem cells.