BS and GS are characterized by an abnormal cell signaling (reduced stimulated [Ca++]i and [IP3]i), pointing toward a reduced PKC and cell reactivity. NO system is up regulated (increased ecNOS mRNA, NO2−/NO3− and cGMP excretion) which further contribute to the vascular hyporeactivity. Since PKC regulates many signaling processes and ecNOS gene expression, we evaluated basal and fMLP-stimulated (10 min, 300 nM) PKC activity PKCα translocation rate (Western blot) in the cytosol and membrane of neutrophyls from 3 BS, 6 GS and 10 controls (C). We also studied the effect of PKC stimulation and inhibition on monocyte ecNOS gene expression. Finally, we evaluated monocyte Gαq gene expression, since Gq protein transduces signals to PLCβ, to generate IP3. Cytosol and membrane basal PKC activity were similar in BS-GS and C (70±3 vs 80±2; 37±3 vs 46±2 pmol/min/mg prot respectively), while fMLP-stimulated membrane PKC activity increased to a lower extent in BS-GS (from 43±2 to 53±3 vs 38±2 to 66±3 pmol/min/mg prot, p<0.05). Membrane PKCα expression was similar (8.5±1.5 vs 12.4±4.0 d.u.), while its Δ increase after fMLP was reduced in BS-GS vs C (4.5±1.4 vs 9.5±2.1 p<0.01). PMA (100 nM) reduced ecNOS gene expression in BS-GS (from 0.80±0.05 to 0.55±0.07 d.u. p<0.001), while incubation of control cells with GF109203X, (a PKC inhibitor, 1.2 mM), increased ecNOS mRNA (from 0.41±0.04 to 0.62±0.05, d.u. p<0.001). Gαq mRNA was reduced in BS-GS vs C (0.87±0.013 vs 0.98±0.005 d.u., p<0.0004). Our data establish that in BS-GS Gαq is reduced and this may cause the altered cell signaling responsible of reduced vascular contractility through reduced PKC. Reduced PKC-dependent NO system up regulation may further contribute to BS and GS vascular hyporeactivity.

Gαq, PKC, ecNOS and vascular reactivity in Bartter's (BS) and Gitelman's syndrome (GS)

MARESCA, ANDREA MARIA;
2000-01-01

Abstract

BS and GS are characterized by an abnormal cell signaling (reduced stimulated [Ca++]i and [IP3]i), pointing toward a reduced PKC and cell reactivity. NO system is up regulated (increased ecNOS mRNA, NO2−/NO3− and cGMP excretion) which further contribute to the vascular hyporeactivity. Since PKC regulates many signaling processes and ecNOS gene expression, we evaluated basal and fMLP-stimulated (10 min, 300 nM) PKC activity PKCα translocation rate (Western blot) in the cytosol and membrane of neutrophyls from 3 BS, 6 GS and 10 controls (C). We also studied the effect of PKC stimulation and inhibition on monocyte ecNOS gene expression. Finally, we evaluated monocyte Gαq gene expression, since Gq protein transduces signals to PLCβ, to generate IP3. Cytosol and membrane basal PKC activity were similar in BS-GS and C (70±3 vs 80±2; 37±3 vs 46±2 pmol/min/mg prot respectively), while fMLP-stimulated membrane PKC activity increased to a lower extent in BS-GS (from 43±2 to 53±3 vs 38±2 to 66±3 pmol/min/mg prot, p<0.05). Membrane PKCα expression was similar (8.5±1.5 vs 12.4±4.0 d.u.), while its Δ increase after fMLP was reduced in BS-GS vs C (4.5±1.4 vs 9.5±2.1 p<0.01). PMA (100 nM) reduced ecNOS gene expression in BS-GS (from 0.80±0.05 to 0.55±0.07 d.u. p<0.001), while incubation of control cells with GF109203X, (a PKC inhibitor, 1.2 mM), increased ecNOS mRNA (from 0.41±0.04 to 0.62±0.05, d.u. p<0.001). Gαq mRNA was reduced in BS-GS vs C (0.87±0.013 vs 0.98±0.005 d.u., p<0.0004). Our data establish that in BS-GS Gαq is reduced and this may cause the altered cell signaling responsible of reduced vascular contractility through reduced PKC. Reduced PKC-dependent NO system up regulation may further contribute to BS and GS vascular hyporeactivity.
2000
L., Calò; G., Ceolotto; E., Pagnin; E., Baritono; M., Milani; M., Sartori; Maresca, ANDREA MARIA; A., Semplicini
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11383/1715241
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