Effects of 1-chloro-2,4,6-trinitrobenzene on 5-lipoxygenase activity and cellular leukotriene synthesis.
5-Lipoxygenase (EC 1.13.11.34) is the key enzyme in the regulation of leukotriene synthesis. Here, the effects of various substituted nitrobenzene compounds on 5-lipoxygenase activity and the formation of leukotriene B4 (LTB4) were studied in polymorphonuclear leukocytes (PMNL), B lymphocytes, and human whole blood. 1-Chloro-2,4,6-trinitrobenzene (TNCB) was found to inhibit calcium ionophore A23187-induced leukotriene synthesis in PMNL in a biphasic manner. Thus, 1.0 microM TNCB caused 50% inhibition of LTB4 formation, but only 16% inhibition was found at 10 times higher concentration. In contrast, this higher concentration of TNCB activated the synthesis of LTB4 when PMNL were stimulated with arachidonic acid alone, demonstrating that TNCB can exert both stimulatory and inhibitory effects on leukotriene synthesis depending on the experimental conditions. The inhibitory effect of 1.0 microM TNCB on ionophore A23187-induced leukotriene synthesis could be circumvented by addition of exogenous arachidonic acid. At high concentrations of TNCB (25-100 microM), the drug blocked ionophore A23187-induced leukotriene synthesis. TNCB also inhibited LTB4 formation in B lymphocytes, as well as in human whole blood. The activity of recombinant 5-lipoxygenase was inhibited by TNCB, and reduced glutathione or beta-mercaptoethanol counteracted this inhibition. This suggests that TNCB might inhibit 5-lipoxygenase by alkylating thiol groups. TNCB possessed a high specificity for 5-lipoxygenase with only modest inhibitory effects on 12-lipoxygenase (EC 1.13.11.31), 15-lipoxygenase (EC 1.13.11.12), and phospholipase A2 (EC 3.1.1.4) activities. Taken together, these results show that TNCB can both specifically inhibit and stimulate leukotriene formation and might be useful in further studies on the regulation of 5-lipoxygenase.
Larsson
P K
PK
Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden.
Hammarberg
T
T
Johnsson
T
T
Lindgren
J A
JA
Rådmark
O
O
Claesson
H E
HE
D001092:Arachidonate 12-Lipoxygenase / Q000187:drug effects; D001093:Arachidonate 15-Lipoxygenase / Q000187:drug effects; D001094:Arachidonate 5-Lipoxygenase / Q000187:drug effects*; D001402:B-Lymphocytes / Q000187:drug effects*; D000001:Calcimycin / Q000494:pharmacology; D002460:Cell Line; D006098:Granulocytes / Q000187:drug effects*; D006801:Humans; D007975:Leukotriene B4 / Q000096:biosynthesis; D015289:Leukotrienes / Q000096:biosynthesis*; D009578:Nitrobenzenes / Q000494:pharmacology; D010741:Phospholipases A / Q000187:drug effects; D054467:Phospholipases A2; D010853:Picryl Chloride / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000187:drug effects
D015289:Leukotrienes; D009578:Nitrobenzenes; D011994:Recombinant Proteins; D007975:Leukotriene B4; D000001:Calcimycin; D001092:Arachidonate 12-Lipoxygenase; D001093:Arachidonate 15-Lipoxygenase; D001094:Arachidonate 5-Lipoxygenase; D010741:Phospholipases A; D054467:Phospholipases A2; D010853:Picryl Chloride