Adh resistance of llc-pk1 cells caused by overexpression of camp-phosphodiesterase type-iv
Yamaki, M Mcintyre, S Murphy, Jm Swinnen, Johan Conti, M Dousa, Tp # ×
Blackwell science inc
Kidney international vol:43 issue:6 pages:1286-1297
The studies of animal models of nephrogenic diabetes insipidus (NDI) suggest that abnormally high activity of cAMP phosphodiesterase (cAMP-PDE) may cause unresponsiveness to the diuretic effect of AVP. We explored whether overexpression of one of the cAMP-PDE type isozymes, PDE-IV, in [8-Arg]-vasopressin (AVP) sensitive renal epithelial LLC-PK1 cells can prevent the hormone-elicited cAMP increase. LLC-PK1 cells were stably transfected with ratPDE3.1 cDNA (which encodes for rolipram-sensitive PDE-IV), inserted in plasmid pCMV5 and then were compared with sham-transfected LLC-PK1 cells and wild LLC-PK1 cells. In the stably transfected clone (LLC-PK1-S#16), the rolipram-sensitive PDE-IV activity was about five times higher than in controls, whereas activities of other types of PDEs were not different. The presence of cognate mRNA for PDE-IV was confirmed by Northern blot. Whereas in the control cells (wild LLC-PK, cells and sham-transfected LLC-PK1 cells), the incubation with 10(-7) m AVP increased cAMP more than tenfold, the LLC-PK1-S#16 cells with overexpressed cAMP-PDE were resistant to cAMP-increasing effects of AVP and forskolin. However, in the same LLC-PK1-S#16 cells the cGMP increases in response to nitroprusside were not diminished. The AVP-dependent cAMP accumulation in LLC-PK1-S#16 cells with overexpressed PDE-IV was restored by addition of roliprams which decreased cAMP-PDE activity to the levels similar to those in wild LLC-PK1 cells and sham-transfected LLC-PK1-#A1 cells. In contrast, inhibitors of other PDE isozymes (PDE-I or PDE-III) had little or no effect. Our findings show that excessive activity of cAMP-PDE, in this case of isozyme PDE-IV, can cause resistance to AVP which is analogous to that observed in collecting ducts of mice with hereditary nephrogenic diabetes insipidus.