Development of an Improved Tracheal Explant Bioassay for the Detection of the Ciliary Dyskinesia Factor in Cystic Fibrosis Serum

Abstract

SummaryA tracheal ring explant system, when used with 25% cystic fibrosis (CF) serum, displayed obvious ciliostasis. Hamster, rabbit, and guinea pig explants all had measurable decreases in ciliary activity after 24 hr of incubation in the serum. The differential response to CF serum (relative to normal serum) was greatly increased by using explants which were maintained 24-72 hr in i*dnimal essential medium (MEM) with 10% horse serum and which were selected on the basis of optimal ciliary activity and vigor. With such a bioassay system of guinea pig tracheal explants, incubation with 25% normal serum would produce essentially no change in relative ciliary activity (score of 242 of a possible 300), whereas CF serum resulted in an 86% decrease (score of 33). Scanning electron microscopic observation indicated that the explants displaying the CF-ciliostatic effect had significant accumulations of mucous over the ciliated epithelial surface. A biochemical viability assay (dehydrogenase activity) showed no cytonecrosis when CF serum-treated tissues were compared to standard explants (10% horse serum in MEM) or control explants (25% normal human serum). SpeculationCystic fibrosis serum, when tested at 25% concentrations on preselected guinea pig tracheal ring cultures, showed an obvious and easily measurable ciliostatic response. This new bioassay may serve as an improved model to detect and analyze the CF dyskinesia factor.

Secretory activity of hamster tracheal explants and isolated tracheal epithelial cells and the effects of cystic fibrosis serum

“…In agreement with the recent work of others (Blitzer and Shapira, 1982;Gabridge et al, 1979) we have demonstrated that CF serum stimulates mucus droplet formation from the cells lining tracheal explants, as observed by scanning electron microscopy. The advent of methods for culturing HTE cells (Goldman and Baseman, 1980a,b) prompted us to attempt similar experiments with the isolated cells of the tracheal lining in the expectation that sensitivity of CF factor would be a property of the individual cells and not just of the intact explant.…”

“…Various sera were added to a final concentration of 25%, and, after incubation at 37 "C for increasing times, the rings were removed and processed for SEM. This protocol is essentially that which Gabridge et al (1979) used to observe the discharge of mucus droplets by tracheal goblet cells. Resulting micrographs show that CF serum (Fig.…”

“…Unfortunately, these proteins have been obtained on the basis of their ability to cause ciliary dyskinesis or ciliotoxicity in tracheal explants (Blitzer and Shapira, 1982;Carson et al, 1976) or oyster gills (Bowman et al, 1980) but the relationship between this inhibitory activity to CF symptoms is not clear. In a few recent cases it has been shown microscopically that a serum component stimu-0 1984 ALAN R. LISS, INC. lates the release of material, presumably mucus, from the luminal surface of the rabbit trachea (Blitzer and Shapira, 1982;Gabridge et al, 1979). Furthermore, Boat et al (1982) have developed a system for quantitating release of gl coproteins labeled to equilibrium with plants and have demonstrated that 50% CF serum induces the release of radiolabeled glycoproteins over that of control serum.…”

The effects of serum immunoglobulins on the metachronal coordination of the lateral cilia of Mytilus edulis

“…The changes in ciliary activity observed in this study are particularly relevant to bioassay studies used for the detection of the disease cystic fibrosis (CF). These assays rely on a differential effect of normal and CF (homo-or heterozygote) serum on ciliary activity (Spock et al, 1967;Conover et al, 1973;Bowman et al, 1977;Gabridge et al, 1979), usually ascribed to the presence of a "CF factor." The inhibition or changes in ciliary activity (ciliary dyskinesia) observed by these authors could result from a reaction between serum IgM and the ciliated tissue used.…”

Effects of cystic fibrosis serum on calcium influx and secretion using isolated tracheal epithelial cells

“…In the difficult search for a molecular component which might be the CF causative agent, perhaps the most consistent finding has been that serum from CF patients contains a factor which may be responsible for the etiology of the disease (Gabridge et al, 1979;Nagy et al, 1979;Kurlandsky et al, 1980;Impero et al, 1981;McNeely et al, 1982;Carson and Bowman, 1982; Mc-Pherson et al, 1983). Such factors have been isolated by 0 1984 ALAN R. LISS, INC several groups on the basis of their ability to inhibit ciliary activity in tracheal explants or oyster gills (Conover and Conod, 1978;McNeely et al, 1982; Carson and Bowman, 1982), and have proved to be low molecular weight (5-10 kdaltons), cationic glycoproteins reportedly associated with IgG (Danes, 1973;Blitzer and Shapira, 1982).…”