Receptor‐Mediated Regulation of Leukotriene C₄ Synthase Activity in Human Platelets

Abstract

Human platelets possess a specific membrane-bound leukotricnc (LT) C., synlhase, which catalyzes the conversion of LTA, to LTC,. Sti tnulation of the rcccptoi-s for thronibin, collagen or thromboxane A, provoked inhibition of this cnzyme, as judged by suppressed trmsformation of exogenous LTA, to LTC,.Similarly, direct activation of protein kinase (PK) C with riiltiomolur concentrations of 4/1-phorhol 12-myristate 13-acelate (PMA) inhibited the production of I,TC:,. Kinetic studies demonstrated that the i nhibition induced by thrombin and PMA was nori-compctitive. Elevation of inlracellular CAMP levels with carbacyclin did not affect basal LTC, formation, hut abolished the attenuation d' platelet LTC, synthasc activity induced by the thromboxane receptor agonisl U-46619. The unselective prolein kinasc inhibitor lstaurosporine prevented both receptor-mediated and PMA-induced suppression of LTC, formation. In contrast, two selective PKC inhibitors, Ro 31 4 2 2 0 and GF 100203X, reversed the inhibitory effect provoked by PMA, but failed to prevent thrombin-induced inhibition, Furthermore, the protein tyrosine phosphatase inhibitor, sodium orthovanadate, induced dose-dependent inhibition of LTC, production in platelet sonicates. In conclusion, receptor-mediated activation u f human platclets leads to decreased I,TC:., synthase activity via phosphoregulation. Although the present results demonstrate that platelet LIC, synthase can be regulated via PKC-dependent events, alternative mechanisms appears to be involved in the physiological regulation of this enzyme. The findings suggcst the possible importance of protein tyrosine phophorylations in this process.

Phorbol ester‐induced suppression of leukotriene C4 synthase activity in human granulocytes

“…Nevertheless, the amounts of LTC 4 produced from LTA4 in the presence of catalase were almost 50% of LTB4 levels in intact cells and similar to those of LTB 4 in disrupted granulocytes, demonstrating considerable LTC4-producing capacity within the granulocyte preparation. The possibility that contaminating platelets contributed substantially to this capacity can be excluded, since the conversion of LTA4 to LTC4 in granulocyte suspensions was unaffected by the thromboxane analogue U-46619 (Sj61inder et al, unpublished observation), which induce down-regulation of LTC4 synthase activity in platelets [23]. Furthermore, the number of remaining platelets in the granulocyte suspensions was always low (see section 2).…”

“…However, although direct activation of PKC induced attenuation of platelet LTC 4 synthase activity, receptormediated regulation of the enzyme appeared to be primarily dependent on the function of another protein kinase, possibly a tyrosine specific kinase [23]. Therefore, further investigation of possible receptor-mediated LTC 4 synthase regulation in myeloid cells is warrented.…”

“…Lymphoprep was from Nycomed Pharma AS (Oslo, Norway). Leukotriene Aa-methyl ester was a generous gift from Dr. Robert Zipkin, Biomol Research Laboratories, and was saponified as described [23].…”

“…Thus, phorbol 12-myristate 13-acetate (PMA) provoked suppression of ionophore A23187-or LTA4-induced LTC4 formation in established human pre-leukemic cell lines [21,22]. Similarly, LTC4 synthase activity in human platelets has been demonstrated to be phosphoregulated via both PKC-dependent and receptor-mediated, PKC-independent mechanisms [23]. It was therefore of interest to investigate whether LTC 4 formation in normal human granulocytes is governed via similar mechanisms.…”

TBXA2R gene polymorphism and responsiveness to leukotriene receptor antagonist in children with asthma

“…Recent studies have shown that TBXA2 can be considered a negative regulator of LTC 4 synthase activity, which is responsible for leukotriene production, and have also suggested that when TBXA2 formation is prevented, LTC 4 synthase activity is elevated, leading to increased biosynthesis of leukotriene and clinical symptoms [11][12][13][14].…”