Activation of protein kinase C reduces thromboxane receptors in glomeruli and mesangial cells.
The potential role of protein kinase C (PKC) in the modulation of thromboxane A2 (TX) receptor density was evaluated in intact glomeruli and cultured renal mesangial cells (MC) from the rat. Incubation of glomeruli with 0.1 microM phorbol dibutyrate (PDBu) or 30 mM glucose for four hours activated PKC as reflected by increased in situ phosphorylation of the 80 kDa MARCKS protein, a specific endogenous substrate for PKC. High affinity binding to TX receptors, as assessed from the binding of the stable TX antagonist [3H]-Sq-29548 (Sq), was decreased 30% in glomeruli exposed to PDBu and 28% in glomeruli incubated in 30 mM D-glucose for four hours. Concurrent incubation with 0.05 microM of the PKC inhibitor staurosporine blocked both MARCKS protein phosphorylation and the decrease in TX receptor sites in response to either PDBu or 30 mM glucose. Neither 30 mM L-glucose nor 30 mM mannitol altered glomerular PKC activity or TX receptor density, thus excluding an osmotic effect of D-glucose, and implicating cellular metabolism of glucose in the expression of these actions. Inhibition of endogenous production of TX with indomethacin during exposure of glomeruli to 30 mM glucose did not prevent the decrease in TX binding. Homologous down-regulation of TX receptors mediated by endogenous TX was therefore not implicated in this action of glucose. The affinity of the glomerular receptor sites for [3H]-Sq was not affected by PKC activation. MC in passages 3 to 7 also demonstrated high affinity sites for [3H]-Sq (Kd, 2.8 nM). Culture of MC with PDBu (0.05 or 0.1 microM) for four hours decreased TX receptor density.(ABSTRACT TRUNCATED AT 250 WORDS)
Studer
R K
RK
Department of Medicine, VA Medical Center, Pittsburgh, Pennsylvania.
Craven
P A
PA
DeRubertis
F R
FR
D000818:Animals; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D005231:Fatty Acids, Unsaturated; D005260:Female; D005920:Glomerular Mesangium / Q000187:drug effects / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D006834:Hydrazines / Q000378:metabolism; D066298:In Vitro Techniques; D047908:Intracellular Signaling Peptides and Proteins; D007678:Kidney Glomerulus / Q000187:drug effects / Q000378:metabolism*; D008565:Membrane Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017482:Receptors, Thromboxane / Q000378:metabolism*
D019086:Bridged Bicyclo Compounds, Heterocyclic; D005231:Fatty Acids, Unsaturated; D006834:Hydrazines; D047908:Intracellular Signaling Peptides and Proteins; C000619712:Marcks protein, rat; D008565:Membrane Proteins; D011506:Proteins; D017482:Receptors, Thromboxane; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D015240:Phorbol 12,13-Dibutyrate; C045749:SQ 29548; D011493:Protein Kinase C; D005947:Glucose