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Phorbol and calcium decreased atriopeptin response in a human renal cell line
T Iwata1, J Vaughn, E D Frohlich
1Alton Ochsner Medical Foundation, New Orleans, LA 70121.
Abstract:
We investigated regulation of atrial natriuretic factor (ANF)-stimulated cellular cGMP accumulation (ANF-s-cGMP) in an ANF-responsive human renal cell line, SK-NEP-1. Dose-response data indicated that the EC50 for ANF(99-126) was 1.1 x 10(-9) M. Brain natriuretic peptide (10(-6) M) increased cGMP to a level indistinguishable from that of ANF (10(-6) M). [Met-(O)]ANF was only half as potent as ANF, and atriopeptin I (10(-6) M) did not increase cGMP over basal levels. Preincubation of SK-NEP-1 cells with ANF, but not atriopeptin I (API), for two hours or longer, caused a concentration-dependent down-regulation of ANF-s-cGMP. Phorbol 12-myristate 13-acetate (PMA), a protein kinase C (PKC) activator, and A23187 and its 4-bromo derivative, calcium ionophores, inhibited ANF-s-cGMP in a dose-dependent manner. A23187 inhibition was calcium dependent and promoted net cGMP degradation. Thirty-six hour preincubation with PMA, a procedure used to down-regulate PKC, abolished acute PMA inhibition of ANF-s-cGMP without having an effect on ANF-s-cGMP or on 4-bromo-A23187 inhibition thereof. These data indicate that PKC activation specifically inhibited ANF-s-cGMP but that PKC was not required for ANF-s-cGMP in SK-NEP-1 cells. Thus structurally related ANF peptides, protein kinase C (PKC) activators, calcium ionophores are potential modulators of ANF-s-cGMP in cells from this human renal cell line.
Insights
This study shows that protein kinase C (PKC) activation and calcium ionophores can inhibit atrial natriuretic factor (ANF)-stimulated cGMP accumulation in human renal cells. PKC activation specifically inhibits ANF-s-cGMP but is not required for it.
Area of Science:
- Biochemistry
- Cell Biology
- Renal Physiology
Background:
- Atrial natriuretic factor (ANF) plays a crucial role in regulating cellular processes, including cGMP accumulation.
- Understanding the modulators of ANF-stimulated cGMP (ANF-s-cGMP) is vital for comprehending renal cell function.
Purpose of the Study:
- To investigate the regulation of ANF-s-cGMP in the human renal cell line SK-NEP-1.
- To identify specific signaling pathways and molecules that modulate ANF-s-cGMP.
Main Methods:
- Dose-response studies with various natriuretic peptides and modulators.
- Preincubation experiments to assess down-regulation effects.
- Use of protein kinase C (PKC) activators (PMA) and calcium ionophores (A23187).
Main Results:
- ANF(99-126) demonstrated an EC50 of 1.1 x 10(-9) M for ANF-s-cGMP.
- Brain natriuretic peptide mimicked ANF's effect, while [Met-(O)]ANF and atriopeptin I showed reduced or no activity.
- ANF preincubation led to down-regulation of ANF-s-cGMP.
- PKC activators and calcium ionophores inhibited ANF-s-cGMP in a dose-dependent manner.
- A23187-mediated inhibition was calcium-dependent and promoted cGMP degradation.
- PKC activation specifically inhibited ANF-s-cGMP but was not essential for its basal accumulation.
Conclusions:
- Protein kinase C (PKC) activation specifically inhibits ANF-s-cGMP in SK-NEP-1 cells.
- Calcium ionophores also modulate ANF-s-cGMP, with A23187 promoting cGMP degradation.
- Structurally related ANF peptides, PKC activators, and calcium ionophores are key regulators of ANF-s-cGMP in this human renal cell line.