Expression and subcellular localization of aquaporin water channels in the polarized hepatocyte cell line, WIF-B.
BACKGROUND
Recent data suggest that canalicular bile secretion involves selective expression and coordinated regulation of aquaporins (AQPs), a family of water channels proteins. In order to further characterize the role of AQPs in this process, an in vitro cell system with retained polarity and expression of AQPs and relevant solute transporters involved in bile formation is highly desirable. The WIF-B cell line is a highly differentiated and polarized rat hepatoma/human fibroblast hybrid, which forms abundant bile canalicular structures. This cell line has been reported to be a good in vitro model for studying hepatocyte polarity.


RESULTS
Using RT-PCR, immunoblotting and confocal immunofluorescence, we showed that WIF-B cells express the aquaporin water channels that facilitate the osmotically driven water movements in the liver, i.e. AQP8, AQP9, and AQP0; as well as the key solute transporters involved in the generation of canalicular osmotic gradients, i.e., the bile salt export pump Bsep, the organic anion transporter Mrp2 and the chloride bicarbonate exchanger AE2. The subcellular localization of the AQPs and the solute transporters in WIF-B cells was similar to that in freshly isolated rat hepatocytes and in intact liver. Immunofluorescent costaining studies showed intracellular colocalization of AQP8 and AE2, suggesting the possibility that these transporters are expressed in the same population of pericanalicular vesicles.


CONCLUSIONS
The hepatocyte cell line WIF-B retains the expression and subcellular localization of aquaporin water channels as well as key solute transporters for canalicular bile secretion. Thus, these cells can work as a valuable tool for regulatory and mechanistic studies of the biology of bile formation.
Gradilone
Sergio A
SA
Instituto de Fisiología Experimental, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, 2000 Rosario, Santa Fe, Argentina. sgradilo@fbioyf.unr.edu.ar
Tietz
Pamela S
PS
Splinter
Patrick L
PL
Marinelli
Raúl A
RA
LaRusso
Nicholas F
NF
D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000818:Animals; D027321:Anion Transport Proteins / Q000378:metabolism; D017920:Antiporters / Q000378:metabolism; D020346:Aquaporins / Q000378:metabolism*; D001648:Bile Canaliculi / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D016764:Cell Polarity; D005136:Eye Proteins / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D022781:Hepatocytes / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D006822:Hybrid Cells; D007473:Ion Channels / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D064606:SLC4A Proteins; D013347:Subcellular Fractions / Q000378:metabolism*; D014018:Tissue Distribution
D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters; C490548:Abcb11 protein, rat; C423013:Abcc2 protein, rat; D027321:Anion Transport Proteins; D017920:Antiporters; C491637:Aqp9 protein, rat; D020346:Aquaporins; D002352:Carrier Proteins; D005136:Eye Proteins; D007473:Ion Channels; D008562:Membrane Glycoproteins; D064606:SLC4A Proteins; C030328:aquaporin 0; C108330:aquaporin 8