Renal leukotriene C4 synthase: characterization, partial purification and alterations in experimental glomerulonephritis.
Cysteinyl leukotrienes (LT) play an important role in the development of experimental glomerulonephritis (GN). We have partially purified and characterized LTC4 synthase, the enzyme responsible for cysteinyl LT formation, from rat renal microsomes and have investigated this enzyme activity in nephritic rats. LTC4 formation, measured in vitro, was linear for > 10 min at 25 degrees C in the presence of 50 mM serine borate (an inhibitor of gamma-glutamyl transpeptidase), with Km values for LTA4 and GSH of 56 microM and 8.5 mM, respectively. Detergent solubilization and anion-exchange chromatography of microsomal proteins resulted in a 7-fold increase in enzyme specific activity. Enzymatic and immunoblot analysis demonstrated that cytosolic and microsomal glutathione S-transferase (GST) activities were distinct from LTC4 synthase activity. Comparison of LTC4 synthase activity in nephritic rats over 21 days revealed an initial increase over the first 24 h following injection of nephrotoxic sera, followed by a subsequent decline until day 7 and a gradual recovery by day 21. Inhibition of LT biosynthesis with MK-0591 (10 mg kg-1 d-1) reduced GN-associated proteinuria by 72% (P < 0.05). These results suggest a potential mechanism for enhanced cysteinyl LT formation in the development of experimental GN and further support their causal role in the etiology of this disease.
Petric
R
R
Department of Pharmacology, Merck Frosst Centre for Therapeutic Research, Pointe Claire-Dorval, QC, Canada.
Nicholson
D W
DW
Ford-Hutchinson
A W
AW
D000818:Animals; D003600:Cytosol / Q000201:enzymology; D005921:Glomerulonephritis / Q000201:enzymology*; D005978:Glutathione / Q000378:metabolism; D005982:Glutathione Transferase / Q000302:isolation & purification / Q000378:metabolism*; D007211:Indoles / Q000494:pharmacology; D007672:Kidney Cortex / Q000201:enzymology*; D007700:Kinetics; D017572:Leukotriene A4 / Q000378:metabolism; D017997:Leukotriene C4 / Q000096:biosynthesis; D008297:Male; D008861:Microsomes / Q000201:enzymology; D011804:Quinolines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley
D007211:Indoles; D017572:Leukotriene A4; D011804:Quinolines; C078266:MK 0591; D017997:Leukotriene C4; D005982:Glutathione Transferase; C041116:leukotriene-C4 synthase; D005978:Glutathione