N-glycosylation augmentation of the cystic fibrosis epithelium improves Pseudomonas aeruginosa clearance.
Chronic lung colonization with Pseudomonas aeruginosa is anticipated in cystic fibrosis (CF). Abnormal terminal glycosylation has been implicated as a candidate for this condition. We previously reported a down-regulation of mannose-6-phosphate isomerase (MPI) for core N-glycan production in the CFTR-defective human cell line (IB3). We found a 40% decrease in N-glycosylation of IB3 cells compared with CFTR-corrected human cell line (S9), along with a threefold-lower surface attachment of P. aeruginosa strain, PAO1. There was a twofold increase in intracellular bacteria in S9 cells compared with IB3 cells. After a 4-hour clearance period, intracellular bacteria in IB3 cells increased twofold. Comparatively, a twofold decrease in intracellular bacteria occurred in S9 cells. Gene augmentation in IB3 cells with hMPI or hCFTR reversed these IB3 deficiencies. Mannose-6-phosphate can be produced from external mannose independent of MPI, and correction in the IB3 clearance deficiencies was observed when cultured in mannose-rich medium. An in vivo model for P. aeruginosa colonization in the upper airways revealed an increased bacterial burden in the trachea and oropharynx of nontherapeutic CF mice compared with mice treated either with an intratracheal delivery adeno-associated viral vector 5 expressing murine MPI, or a hypermannose water diet. Finally, a modest lung inflammatory response was observed in CF mice, and was partially corrected by both treatments. Augmenting N-glycosylation to attenuate colonization of P. aeruginosa in CF airways reveals a new therapeutic avenue for a hallmark disease condition in CF.
Martino
Ashley T
AT
Department of Molecular Genetics and Microbiology, College of Medicine, University of Florida, Gainesville, Florida, USA.
Mueller
Christian
C
Braag
Sofia
S
Cruz
Pedro E
PE
Campbell-Thompson
Martha
M
Jin
Shouguang
S
Flotte
Terence R
TR
D000818:Animals; D002460:Cell Line; D002469:Cell Separation; D003550:Cystic Fibrosis / Q000378:metabolism* / Q000382:microbiology*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics; D004848:Epithelium / Q000378:metabolism*; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D015316:Genetic Therapy / Q000379:methods; D006031:Glycosylation; D006801:Humans; D007249:Inflammation; D008358:Mannose / Q000737:chemistry; D008359:Mannose-6-Phosphate Isomerase / Q000378:metabolism; D051379:Mice; D011550:Pseudomonas aeruginosa / Q000378:metabolism*
D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D008359:Mannose-6-Phosphate Isomerase; D008358:Mannose