Functional properties of proximal tubule cell lines derived from transgenic mice harboring L-pyruvate kinase-SV40 (T) antigen hybrid gene

This study describes the functional characterization of two cell lines derived from the proximal convoluted (PKSV-PCT cells) and proximal straight (PKSV-PR) tubules microdissected out from kidneys of transgenic mice harboring the simian virus 40 (SV40) large T and small t antigens placed under the control of the rat Ltype pyruvate kinase (L-PK) 5 regulatory sequence. Both cell lines exhibited cellular cyclic AMP stimulated by parathormone (PTH) and calcitonin (CT) and a sodium-dependent glucose transporter. Uptake of the fluid-phase marker [ 3 H]inulin showed that both cell lines grown on filters exhibited biphasic apical and basolateral endocytic rates. Results from Northern blot analysis indicate that the expression of the T antigen gene (Tag) is dependent on the concentration of D-glu cose in the medium and show that the L-PK construct has maintained its capacity for up- or down-regulati on by carbohydrates. Replacement of D-glucose by neoglucogenic substrates (lactate, oxaloacetate) blunted the expression of Tag transcripts and induced arrest of cell growth. Compared to cell grown in D-glucoseenriched medium, the hormonal sensitivities to PTH and CT and the sodium-dependent glucose uptake were unchanged whereas quiescent cells exhibited increased hydrolase content. Thus the proximal function has been preserved in these cultured cells derived from tissuespecific targeted oncogenesis in transgenic mice. As the expression of Tag transcripts is controlled by D-glucose, the structural and physiological characteristics of these cell lines can be studied in either quiescent or active growth conditions. SUMMARY