Oxidized low-density lipoprotein stimulates protein kinase C (PKC) and induces expression of PKC-isotypes via prostaglandin-H-synthase in P388D(1) macrophage-like cells

Treatment of cells with LPS-free oxLDL significantly enhanced protein kinase C (PKC) activity in cell extracts from P383D(1) macrophage-like cells as determined by phosphorylation of histone H1 or Ac-MBP[4-14] substrate peptide. This effect was abolished by the PKC inhibitors H-7 and bisindolylmaleimide I while pertussis toxin failed to block stimulation. The phosphotransferase activity was also increased by acetylated LDL (acLDL) and maleylated albumin (malBSA), the oxLDL effect was inhibited by chloroquine which also blocked oxLDL-induced stimulation of tyrosine kinase activity. Marginal stimulation of PKC activity was observed when lipid extracts from oxLDL were used, indicating that uptake via scavenger receptors (SR) is mandatory. Polyinosinic acid (poly I) exhibited a concentration-dependent inhibition of the oxLDL-induced effect suggesting that SR II/I but not CD36 interactions are critical to PKC activation. Modified (lipo)proteins increased the concentration of diacylglycerol and differentially affected the levels of individual PKC isoenzymes predominantly in the cytosolic fraction. Changes of activity induced by oxLDL could be primarily assigned to alterations of the activities and levels of the isoenzymes beta and delta. Treatment with oxLDL, acLDL, and malBSA was also accompanied by increased production of prostaglandins as well as by an enhanced level of cyclooxygenase 2 (COX 2) as determined by Western blot analysis. Effecs of oxLDL on PKC activity/expression was supressed by the cyclooxygenase inhibitors indomethacin, by pre-exposure to the inhibitor of both lipoxygenase and cyclooxygenase, 2,2-dimethyl-6-(4-chlorophenyl)-7-phenyl-2,2-dihydro-1H-pyrrolizine-5-ylacetic acid (ML 3000), and by treatment with the specific COX 2-inhibitor N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide (NS-398). These results indicate that oxLDL, acLDL, and malBSA exhibit a COX 2-dependent and isotype specific effect on PKC in P388D(1) cells following uptake via SR II/I and subsequent lysosomal degradation.