Abstract
GRB-PAP1 is a continuous line of epithelial cells derived from a rabbit renal inner medulla. Elevation of the NaCl concentration in the medium bathing these cells strongly induced the expression of a soluble protein with an apparent molecular mass of 39 kDa. The protein, purified by affinity chromatography with Amicon Matrex Gel Orange A, had enzyme activity characteristic of aldose reductase (alditol:NADPH+ oxidoreductase, EC 1.1.1.21). Goat antiserum against this purified aldose reductase selected the 39-kDa band from immunoblots of cells grown in a medium containing high NaCl. When the osmolality of the medium was increased by adding NaCl, the amount of aldose reductase protein and the aldose reductase activity increased together from very low to sustained high levels over several days. The aldose reductase protein was more than 10% of the soluble cell protein when cells were propagated in medium made hyperosmotic by adding NaCl to increase medium osmolality to 600 mosm.kg-1.
Highlights
CeZLs-GRB-PAP1 is a line of epithelial cells, derived from the of the NaCl concentration in thme edium bathing these surface of the rabbit renalpapilla and grown in an isosmotic medium cells strongly induced theexpression of a soluble pro- (300 mosmkg-') with a normal NaCl concentration past 7 years and tein with an apparent molecular mass of 39 kDa
A conspicuous band corresponding to a molecular mass of about 39 kDa was present in the preparation from hyperosmotic medium (Fig. 1, left lane),but thisband appeared much reduced in the cells grown in isosmotic medium (Fig. 1,right lane).This 39-kDa protein band was only observed in thesoluble protein fraction
When PAP-HT25 cellsg,rown in an isosmotic mediumwere switched to a hyperosmotic medium, the 39-kDa band was observed to increase between 1and 5 days (Fig. 2)
Summary
CeZLs-GRB-PAP1 is a line of epithelial cells, derived from the of the NaCl concentration in thme edium bathing these surface of the rabbit renalpapilla and grown in an isosmotic medium cells strongly induced theexpression of a soluble pro- (300 mosmkg-') with a normal NaCl concentration past 7 years and tein with an apparent molecular mass of 39 kDa. Both protein concentration and aldose reductase activity (using 100-pl samples) were monitored in all the fractions.
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