Der p 1 facilitates transepithelial allergen delivery by disruption of tight junctions

Wan, Hong; Winton, Helen L; Soeller, Christian; Tovey, Euan R.; Gruenert, Dieter C.; Thompson, Philip J.; Stewart, Geoffrey A.; Taylor, Graham W.; Garrod, David R.; Cannell, Mark B.; Robinson, Clive

doi:10.1172/jci5844

Cited by 665 publications

(542 citation statements)

References 52 publications

(44 reference statements)

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“…16HBE14o-cells possess tight junctions, cystic fibrosis transmembrane conductance regulator and functional chloride conductances [19], they express functional b-adrenoceptors [39], and the cytokines interleukin-2 [40], and interleukin-8 [39], utilize the transcription factor nuclear factor-kB [41] and contain type-III phosphodiesterase [42]. Taken together with the evidence from other investigations [25], these data collectively suggest that the 16HBE14o-and Calu-3 epithelial cell lines may prove to be useful paradigms of the permeability functions of human airway epithelium, especially in studies of tight junction assembly and disassembly.…”

Section: Discussionsupporting

confidence: 55%

“…The human airway cell lines had additional weak bands detectable by immunoblotting. It is likely that these weak bands arise by proteolysis since the authors have detected similar bands in cells explicitly treated with proteinases prior to immunoblotting [25]. Whilst the possibility that the proteinase inhibitors failed to provide adequate inhibition of TJ protein proteolysis during the preparation of cell extracts cannot be excluded, the high prevalence of breakdown products in Calu-3 cells may be indicative of a continuous turnover of TJ proteins in this cell line.…”

Section: Discussionmentioning

confidence: 96%

“…Alterations in the plasticity of TJs, or their outright cleavage, have been suggested to be significant potentially pathophysiological events in the airways epithelium [1,25]. Progress towards understanding the mechanisms involved have been impeded by the absence of wellcharacterized cellular models.…”

Section: Discussionmentioning

confidence: 99%

“…The technique of 2PMEM limits specimen-bleaching outside the focal plane and thus enables quantitative analysis of fluorescent antibody labelling to be performed. Quantitative digital image analysis of the breaks within the TJ staining pattern was performed as fully described elsewhere [25].…”

Section: Cellular Staining By Fluorescent Probes and Indirect Immunocmentioning

confidence: 99%

“…Whole cell protein extracts for immunoblotting were prepared exactly as described elsewhere [25]. Nonspecific protein binding was blocked with 5% weight (w)/v nonfat milk and 0.1% v/v Tween-20 in Tris-buffered saline (TBS) followed by incubation with primary antibodies diluted in TBS containing 2% w/v bovine serum albumin and 0.1% v/v Tween-20.…”

Section: Immunoblottingmentioning

confidence: 99%

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Tight junction properties of the immortalized human bronchial epithelial cell lines Calu‐3 and 16HBE14o‐

Wan¹,

Winton²,

Soeller³

et al. 2000

Eur Respir J

139

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Tight junctions (TJs) make a vital contribution to the barrier properties of the airway lining. Opening of TJs, or their frank cleavage, is suspected as a pathophysiological event in the lung, but research into the cellular and molecular mechanisms involved has been impeded by technical limitations of available experimental models. The authors have compared the properties of two epithelial cell lines derived from bronchial epithelium to explore whether these cell lines could constitute appropriate tools for the study of TJ regulation in bronchial epithelium.Investigations of TJs in 16HBE14o-cells and Calu-3 cells were made by fluorescent antibody labelling in conjunction with wide-field, confocal or 2-photon molecular excitation microscopy (2PMEM). The presence of TJ proteins was confirmed by immunoblotting and functional properties of the monolayers were studied by measurements of transepithelial electrical resistance and mannitol permeability.Cells of both lines formed confluent monolayers in which the cells expressed the TJ proteins occludin and ZO-1 in continuous circumferential patterns suggestive of functional TJs. This interpretation was supported by the development of transepithelial electrical resistances and of low paracellular permeability to solutes. Within the limits of resolution offered by 2PMEM, occludin and ZO-1 appeared to colocalize at TJs.These studies suggest that the 16HBE14o-cells and Calu-3 cell lines are potentially useful in vitro models to study how tight junction opening or cleavage changes the functional barrier properties of bronchial epithelium. Eur Respir J 2000; 15: 1058±1068.

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