Cathepsin L activity in alveolar macrophages of rats: response to cigarette smoke.
Cathepsin L activity was quantitated in alveolar macrophages (AMs) and bronchoalveolar lavage (BAL) fluid from Sprague-Dawley rats exposed through the nose only to fresh mainstream smoke from University of Kentucky reference cigarettes (2Rl), and in AMs and BAL fluid of room control (RC) and sham control (SH) animals. Activity was determined with the methylcoumarylamide substrate, benzyloxycarbonyl-Phe-Arg-4-methyl-7-coumarylamide (Z-Phe-Arg-NMec). Total activity of cysteine proteinases in AM homogenates towards Z-Phe-Arg-NMec was measured by determining inhibition with E-64 (0.5 microM), a specific inhibitor of this class of enzymes. The activities of cathepsin B and L towards the substrate were differentiated by use of the diazomethane inhibitor, Z-Phe-Phe-CHN2 (0.01 microM), which selectively inhibits cathepsin L at this concentration. We found that cathepsin L activity was significantly elevated in AMs of animals exposed to 10 puffs of cigarette smoke, twice a day, 7 days/wk for 22 wk (1,620 +/- 870 units/mg protein) as compared with cells from RC (420 +/- 150) and SH (550 +/- 160) animals. Cathepsin B activity was also increased in AMs from animals exposed to cigarette smoke (2,360 +/- 890) as compared to RC (930 +/- 170) and SH (1,020 +/- 70) animals. Cathepsin L and B activity was also present in unconcentrated BAL fluid, but levels did not differ significantly among the three groups. The results demonstrate that AMs contain significant levels of cathepsin L activity as measured with methylcoumarylamide substrates, and that activity increases in cells from rats exposed to cigarette smoke in amounts comparable to those inhaled by a healthy 70-kg human smoker using 20 cigarettes a day.(ABSTRACT TRUNCATED AT 250 WORDS)
Lesser
M
M
Veterans Administration Medical Center, Bronx, New York 10468.
Galicki
N
N
Cardozo
C
C
Gariola
C G
CG
D000818:Animals; D015415:Biomarkers; D001835:Body Weight; D001992:Bronchoalveolar Lavage Fluid; D056668:Cathepsin L; D002403:Cathepsins / Q000378:metabolism*; D003374:Coumarins; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004151:Dipeptides; D010450:Endopeptidases; D005456:Fluorescent Dyes; D008264:Macrophages / Q000201:enzymology*; D008297:Male; D011650:Pulmonary Alveoli / Q000201:enzymology*; D051381:Rats; D011919:Rats, Inbred Strains; D012907:Smoking
D015415:Biomarkers; D003374:Coumarins; D004151:Dipeptides; D005456:Fluorescent Dyes; C021159:benzyloxycarbonyl-phenylalanylarginine-4-methylcoumaryl-7-amide; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C534281:CTSL protein, human; D056668:Cathepsin L; C534280:Ctsl protein, rat