Everyone knows that cystic fibrosis produces its most life-threatening problems in the lungs, but no one fully understands how the loss of CFTR, a chloride channel that the CF gene normally produces, causes lung disease. Dr. Walter Finkbeiner and his colleagues at the University of California at San Francisco (UCSF) have now discovered a new immortal cell line from among a large collection of human lung tumor cell lines. This cell line is starting to provide answers to previous mysteries.
A major problem in understanding lung disease is the lack of an adequate animal model or an adequate cell model. The CF mouse, which is proving to be of great help in understanding many aspects of the disease, has not yet shown signs of significant lung disease. The lungs of mice differ from those of humans in several respects, including the presence of an alternate chloride channel. With regard to a model cell line, most cell lines from mouse lungs either do not express CFTR or do not perform like epithelial cells, or both, making them essentially useless for further studies of cystic fibrosis. These technical limitations are compounded by a paradox encountered when studying normal lungs: when large pieces of lung are assayed for CFTR protein, little is found. If there is usually so little CFTR in the lung, how can its loss be significant?
This last problem has recently been solved by investigators in Philadelphia, and their answer highlights the importance of Finkbeiner's new cell line. The low average levels of CFTR in the lung come about because most lung cells have little or no CFTR, while some cells have abundant amounts. The lung cells that are rich in CFTR are called serous cells and are usually located deep within glands that lie beneath the surface of the airways. These cells secrete a watery fluid rich in antibiotics, and have been considered by Carol Basbaum, Ph.D., and her colleagues at UCSF to be the first line of defense against lung infections. Serous cells secrete a watery, antibacterial-rich fluid that mixes with sticky mucin molecules secreted by other gland cells. The mucin-secreting cells have been shown by several investigators, including Jonathan Widdicombe, Ph.D., and his colleagues at UCSF, to lack CFTR, and so they are presumably normal in CF lungs.
Serous cells are difficult to isolate and grow in culture. That is why there is excitement about the recently discovered cell line mentioned above, which turns out to have many properties of serous cells, including the production of antibiotics and the expression of high levels of CFTR protein. Experiments by Jonathan Widdicombe, Jeffrey Wine and their colleagues at UCSF and Stanford have shown that the serous cell model, called Calu-3 cells (for cancer, lung), have preserved all of the important features of epithelial transport and, hence, are outstanding cells for understanding CFTR. Unlike most immortal cells, Calu-3 cells form sheets of cells that are welded to each other by tight junctions. These sheets form a fully functional epithelium that can transport large quantities of ions and fluid. In addition, Calu-3 cells have the highest level of natural CFTR expression of any known immortalized cell, even higher than some intestinal cell lines that once held the record.
Although still hypothetical, a picture is emerging for a direct role of CFTR chloride channels in lung mucosal defense. If serous cells stopped secreting fluid, the airway surface fluid of the lung should lose critical components needed for defense.
In fact, Jonathan Widdicombe and colleagues have direct evidence that in cystic fibrosis, serous cells no longer secrete to any kind of stimulus. Thus, they differ from other glands (for example, the sweat glands), which in CF subjects retain their ability to secrete to agents that elevate calcium.
Patch-clamp experiments, which can reveal the activity of single CFTR channels, have helped to explain the key role that CFTR plays in Calu-3 cells (and presumably in serous cells as well). CFTR turns out to be the only significant chloride channel in the membrane of these cells. Other kinds of chloride channels are sometimes seen under abnormal conditions, but so far CFTR appears to be the dominant channel of healthy Calu-3 cells, which explains why they lose their ability to respond to all agents in CF.
Unfortunately, this suggests that attempts to bypass CFTR by activating calcium-dependent chloride channels will not work in this critical gland within the lung. An interesting hypothesis is that some people have small amounts of calcium-activated chloride channels in these glands that help to maintain more healthy lungs. This would not necessarily be correlated with their CFTR alleles.
The investigation of the role of serous cells in preventing lung infections, and the role of CFTR within serous cells, gets at the heart of the most important problem in cystic fibrosis, and should provide information that will be essential in designing effective therapies.All of the investigators involved in studying Calu-3 cells have received funding from CFRI.
Return to Fall 1994 Index Page