Faculty Opinions recommendation of Epithelial fluid transport: protruding macromolecules and space charges can bring about electro-osmotic coupling at the tight junctions.

Mammary epithelial 31EG4 cells (MEC) were grown as monolayers on filters to analyze the apical membrane mechanisms that help mediate ion and fluid transport across the epithelium. RT-PCR showed the presence of cystic fibrosis transmembrane conductance regulator (CFTR) and epithelial Na+ channel (ENaC) message, and immunomicroscopy showed apical membrane staining for both proteins. CFTR was also localized to the apical membrane of native human mammary duct epithelium. In control conditions, mean values of transepithelial potential (apical-side negative) and resistance ( R T) are −5.9 mV and 829 Ω · cm2, respectively. The apical membrane potential ( V A) is −40.7 mV, and the mean ratio of apical to basolateral membrane resistance ( R A/ R B) is 2.8. Apical amiloride hyperpolarized V A by 19.7 mV and tripled R A/ R B. A cAMP-elevating cocktail depolarized V A by 17.6 mV, decreased R A/ R B by 60%, increased short-circuit current by 6 μA/cm2, decreased R T by 155 Ω · cm2, and largely eliminated responses to amiloride. Whole cell patch-clamp measurements demonstrated amiloride-inhibited Na+ currents [linear current-voltage ( I-V) relation] and forskolin-stimulated Cl−currents (linear I-V relation). A capacitance probe method showed that in the control state, MEC monolayers either absorbed or secreted fluid (2–4 μl · cm−2 · h−1). Fluid secretion was stimulated either by activating CFTR (cAMP) or blocking ENaC (amiloride). These data plus equivalent circuit analysis showed that 1) fluid absorption across MEC is mediated by Na+ transport via apical membrane ENaC, and fluid secretion is mediated, in part, by Cl− transport via apical CFTR; 2) in both cases, appropriate counterions move through tight junctions to maintain electroneutrality; and 3) interactions among CFTR, ENaC, and tight junctions allow MEC to either absorb or secrete fluid and, in situ, may help control luminal [Na+] and [Cl−].

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Campylobacter jejuni inhibits the absorptive transport functions of Caco-2 cells and disrupts cellular tight junctions

Microbiology ◽

10.1099/mic.0.27950-0 ◽

2005 ◽

Vol 151 (7) ◽

pp. 2451-2458 ◽

Cited By ~ 61

Author(s):

Amanda MacCallum ◽

Simon P. Hardy ◽

Paul H. Everest

Keyword(s):

Tight Junction ◽

Tight Junctions ◽

Campylobacter Jejuni ◽

Fluid Transport ◽

Cell Function ◽

Short Circuit ◽

Short Circuit Current ◽

Ussing Chambers ◽

Cell Monolayers ◽

Junction Protein

Caco-2 cells are models of absorptive enterocytes. The net transport of fluid from apical to basolateral surfaces results in ‘domes' forming in differentiated monolayers. Here, the effect of Campylobacter jejuni on this process has been examined. C. jejuni caused no changes in short-circuit current upon infection of Caco-2 cell monolayers in Ussing chambers. Thus, no active secretory events could be demonstrated using this model. It was therefore hypothesized that C. jejuni could inhibit the absorptive function of enterocytes and that this may contribute to diarrhoeal disease. C. jejuni infection of fluid-transporting (‘doming’) Caco-2 cells resulted in a significant reduction in dome number, which correlated with a decrease in tight junction integrity in infected monolayers, when measured as transepithelial electrical resistance. Defined mutants of C. jejuni also reduced dome numbers in infected monolayers. C. jejuni also altered the distribution of the tight junction protein occludin within cell monolayers. The addition to monolayers of extracellular gentamicin prevented these changes, indicating the contribution of extracellular bacteria to this process. Thus, tight junction integrity is required for fluid transport in Caco-2 cell monolayers as leaky tight junctions cannot maintain support of transported fluid at the basolateral surface of infected cell monolayers. Inhibition of absorptive cell function, changes in epithelial resistance and rearrangement of tight junctional proteins such as occludin represent a potential diarrhoeal mechanism of C. jejuni.

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PERMEABLE JUNCTIONAL COMPLEXES

The Journal of Cell Biology ◽

10.1083/jcb.54.2.302 ◽

1972 ◽

Vol 54 (2) ◽

pp. 302-312 ◽

Cited By ~ 130

Author(s):

Terry E. Machen ◽

David Erlij ◽

F. B. P. Wooding

Keyword(s):

Tight Junctions ◽

Fluid Transport ◽

Electrical Potential ◽

Junctional Complex ◽

Bathing Solution ◽

Ion Permeation ◽

Electron Microscopic ◽

Resistance Pathway ◽

Junctional Complexes

Ionic lanthanum has been used to study transepithelial ion permeation in in vitro rabbit gallbladder and intestine (ileum) by adding 1 mM La3+ to only the mucosal bathing solution. Transepithelial fluid transport electrical potential differences (p.d.), and resistances were measured. During La3+ treatment the gallbladder's rate of active solute-coupled fluid transport remained constant, the resistance increased, and the 2:1 NaCl diffusion p.d. decreased. Mucosa-to-serosa fluxes of 140La3+ were measured and indicate a finite permeability of the gallbladder to La3+. La3+ also increased the transepithelial resistance and p d. of ileum. Electron microscopic examination of La3+-treated gallbladder showed: (a) good preservation of the fine structure, (b) electron-opaque lanthanum precipitates in almost every lateral intercellular space, most frequently near the apical end of the lateral spaces close to or within the junctional complex, (c) lanthanum among the subjacent muscle and connective tissue layers, and (d) lanthanum filling almost the entire length of so-called "tight" junctions. No observations were made which unequivocally showed the penetration of lanthanum into the gallbladder cells. Electron micrographs of similar La3+-treated ilea showed lanthanum deposits penetrating the junctional complexes. These results coupled with other physiological studies indicate that the low resistance pathway for transepithelial ion permeation in gallbladder and ileum is through the tight junctions A division of salt-transporting epithelia into two main groups, those with "leaky" junctional complexes and those with tight junctional complexes, has been proposed.

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Freeze-etch observations on the tight junctions of sertoli cells grown in organ culture with FSH

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010008256x ◽

1978 ◽

Vol 36 (2) ◽

pp. 340-341

Author(s):

Rita Meyer ◽

Zoltan Posalaky ◽

Dennis Mcginley

Keyword(s):

Organ Culture ◽

Tight Junctions ◽

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Barrier Function ◽

Cell Junction ◽

Intramembranous Particles ◽

Junctional Complexes ◽

Oriented Parallel

The Sertoli cell tight junctional complexes have been shown to be the most important structural counterpart of the physiological blood-testis barrier. In freeze etch replicas they consist of extensive rows of intramembranous particles which are not only oriented parallel to one another, but to the myoid layer as well. Thus the occluding complex has both an internal and an overall orientation. However, this overall orientation to the myoid layer does not seem to be necessary to its barrier function. The 20 day old rat has extensive parallel tight junctions which are not oriented with respect to the myoid layer, and yet they are inpenetrable by lanthanum. The mechanism(s) for the control of Sertoli cell junction development and orientation has not been established, although such factors as the presence or absence of germ cells, and/or hormones, especially FSH have been implicated.

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