P048 Cigarette smoke impairs intestinal epithelial cell polarity development and morphogenesis
L. J. Klunder*1, T. Blokzijl2, R. Ferreira1, K.-N. Faber2, G. Dijkstra2, S. C. D. van IJzendoorn1
1University Medical Centre Groningen, Department of Cell Biology, Groningen, Netherlands, 2University Medical Centre Groningen, Department of Gastroenterology and Hepatology, Groningen, Netherlands
Cigarette smoking is emerging as an important life-style factor that affects intestinal function. Epidemiological studies indicated that cigarette smoking influences the course of inflammatory bowel disease and is a risk factor for colorectal cancer and intestinal polyp development. The mechanism via which cigarette smoke exerts its effect on the intestinal epithelium is unclear. These may involve indirect effects via alterations in the immune system or direct effects of cigarette smoke on the intestinal epithelium. Little is known about these direct effects of cigarette smoke on intestinal epithelial cells. Here we present an in-vitro study of these direct effects
Human intestinal epithelial Caco-2 cell-matrigel-embedded spheriods were grown for 7 days in the presence of, or pretreated with, cigarette smoke extract (CSE). For pretreatment cell were incubated with CSE for 24 hours, and subsequently plated out in fresh in 3D culture. Intestinal epithelial barrier function was measured with a FITC-dextran leakage assay. Cell polarity and morphogenesis were assessed with fluorescently labelled polarity markers
We found that CSE did not affect the polarity and barrier function of preestablished lumen-forming Caco-2 spheroids. However, when exposed to CSE during the process of 3D morphogenesis, cells showed impaired growth and impaired de novo development of apical-basal cell polarity and tight junctions, resulting in defective self-assembly into single lumen-forming spheroids. This effect was dose-dependent and this phenotype could be induced with preincubation with CSE as well. However, the administration of the glutathione precursor N-acetyl-L-cysteine (NAC) completely neutralised the inhibitory effects of CSE. The mechanisms by which NAC rescued the phenotype were independent of its capacity to induce glutathione production
Our data demonstrate that CSE has direct and persistent inhibitory effects on intestinal epithelial cell polarity development and morphogenesis, and provide insight into the cell biological mechanisms. NAC supplementation was identified as a protective strategy for smoke-induced epithelial dysfunction.