Paracellular barrier and junctional protein distribution depend on basolateral extracellular Ca2+ in cultured epithelia.
The polarised nature of the increase in paracellular permeability induced by Ca(2+)-chelation with EGTA was investigated in several cultured epithelial cell lines. In strain I MDCK cells (canine kidney cells), a marked decrease (> 90%) in transepithelial electrical resistance (RT) and increase in mannitol and inulin permeabilities were only observed after addition of EGTA (for 4 h) to either basolateral (basal) or both (apical+basal) bathing solutions; apical Ca(2+)-chelation resulted in significant smaller changes (approximately 30%) in these variables. The increase in paracellular permeability upon basal EGTA addition was significantly lower than that produced by simultaneous apical and basal addition of 2 mM EGTA. A higher concentration of EGTA (20 mM) did not significantly eliminate this difference in potency between basal and apical+basal Ca(2+)-chelation. The polarised Ca(2+)-dependence of the paracellular barrier was associated with polarised effects on the junctional/cytoskeletal protein distribution. Basal or apical+basal EGTA addition induced substantial internalisation of uvomorulin with some cellular redistribution of the perijunctional actin ring and desmosomes and gaps in ZO-1 location between adjacent cells. In addition, polarised Ca(2+)-dependence of the paracellular barrier (assessed by measuring RT) was observed also in strain II MDCK and two human adenocarcinoma intestinal cell lines, Caco-2 and HCT-8, demonstrating generality of the phenomenon. Therefore, the data show a polarity in the ability of EGTA to enhance epithelial permeability and induce cellular redistribution of cytoskeletal/junctional proteins in several epithelia. The basolateral membrane sensitivity to Ca(2+)-chelation might be explained by the polarised distribution of uvomorulin.
Collares-Buzato
C B
CB
Gastrointestinal Drug Delivery Research Centre, University of Newcastle upon Tyne, Medical School, UK.
McEwan
G T
GT
Jepson
M A
MA
Simmons
N L
NL
Hirst
B H
BH
D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002118:Calcium / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D002463:Cell Membrane Permeability; D003598:Cytoskeletal Proteins / Q000378:metabolism; D004285:Dogs; D004533:Egtazic Acid / Q000737:chemistry / Q000494:pharmacology; D004553:Electric Conductivity; D017097:Electric Impedance; D007444:Inulin / Q000378:metabolism; D008353:Mannitol / Q000378:metabolism; D011188:Potassium / Q000378:metabolism
D015820:Cadherins; D003598:Cytoskeletal Proteins; D008353:Mannitol; D004533:Egtazic Acid; D007444:Inulin; D011188:Potassium; D002118:Calcium