Rat lung alveolar type II cell line maintains sodium transport characteristics of primary culture.
Culture of primary alveolar type II cells has been widely used to investigate the Na+ transport characteristics of alveolar epithelium. However, this model was restricted by early morphological and physiological dedifferentiation in culture. Recently, a cell line has been obtained by transfection of neonatal type II cells with the simian virus SV40 large T antigen gene (SV40-T2). SV40-T2 cells have retained proliferative characteristics of the primary type II cells (Clement et al., 1991, Exp. Cell Res., 196:198-205.) In the present study, we have characterized Na+ transport pathways in SV40-T2 cells. SV40-T2 cells retained most cardinal properties of the original alveolar epithelial cells. Na+ entry occurred, as in primary cultures, through both Na(+)-cotransporters and amiloride-sensitive Na+ channels. SV40-T2 cells expressed Na(+)-phosphate. Na(+)-amino acid and Na(+)-K(+)-Cl cotransports which are quantitatively similar to that of primary cultures. The existence of amiloride-sensitive Na+ channels was supported by molecular and functional data. SV40-T2 expressed the cloned alpha- and gamma-mRNAs for the rat epithelial Na+ channel (rENaC), whereas beta subunit was not detected, and 22Na+ influx was significantly inhibited by 10 microM amiloride. Na+, which enters SV40-T2 cells, is extruded through a Na+, K(+)-ATPase: mRNA for alpha 1 and beta 1 isoforms of Na+, K(+)-ATPase were present and Na+, K(+)-ATPase activity was evidenced either on intact cells by the presence of a ouabain-sensitive component of 86Rb+ influx or on cell homogenates by the measurement of ouabain-inhibitable ATP hydrolysis. These results indicate that SV40-T2 cell line displays most of the Na+ transport characteristics of well-differentiated primary cells in the first days of culture. We conclude that the SV40-T2 cell line provides a model of differentiated alveolar type II cells and may be a powerful tool to study, in vitro, the modulation of Na+ transport in pathophysiological conditions.
Michaut
P
P
Department of Physiology, Faculté de Médecine Xavier Bichat, Université Denis Diderot, Paris, France.
Planes
C
C
Escoubet
B
B
Clement
A
A
Amiel
C
C
Clerici
C
C
D000584:Amiloride / Q000494:pharmacology; D026905:Amino Acid Transport Systems; D026921:Amino Acid Transport Systems, Neutral; D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D003470:Culture Media / Q000494:pharmacology; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D011650:Pulmonary Alveoli / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012964:Sodium / Q000378:metabolism*; D015222:Sodium Channels / Q000187:drug effects / Q000378:metabolism; D051269:Sodium-Glucose Transporter 1; D050608:Sodium-Phosphate Cotransporter Proteins; D050610:Sodium-Phosphate Cotransporter Proteins, Type II; D028021:Sodium-Potassium-Chloride Symporters; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D027981:Symporters
D026905:Amino Acid Transport Systems; D026921:Amino Acid Transport Systems, Neutral; D002352:Carrier Proteins; D003470:Culture Media; D008562:Membrane Glycoproteins; D009004:Monosaccharide Transport Proteins; D015222:Sodium Channels; D051269:Sodium-Glucose Transporter 1; D050608:Sodium-Phosphate Cotransporter Proteins; D050610:Sodium-Phosphate Cotransporter Proteins, Type II; D028021:Sodium-Potassium-Chloride Symporters; D027981:Symporters; C052168:sodium-alanine cotransporter; D000584:Amiloride; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase