FEMALE SEX STEROID HORMONES 313 the hormones were used as a control. In a number of samples, cells were stimulated with zymosan opsonized with a pool of sera. After incubation of neutrophils for 1 h in the pres ence of the hormones, the enzyme activity was deter mined spectrophotometrically in cell culture superna tants. We assessed both the spontaneous activity and the activity induced by the opsonized zymosan (30 µL, Sigma, United States), which was added to the sample simultaneously with the hormones. The activity of sMPO was determined using the substrate mixture of 0.04% orthophenylenediamine and 0.014% Н2О2 in phosphate citrate buffer (pH 5.0)[7]. The optical density was recorded with an Anthos HMTL III mul tichannel spectrophotometer (Labtec Instruments, Austria) at a wavelength of 492 nm. The results were expressed in units of optical density (U). The activity of elastase and cathepsin G was evaluated at 340 nm using the specific synthetic substrates BAEE (Sigma, United States) dissolved in acetonitrile (for elastase [6]) and BTEE (Sigma, United States) dissolved in methanol (for cathepsin G [7]). The enzyme activity was expressed in units, each of which corresponded to 1 nM of the substrate hydrolyzed in 1 min (nM/mL/min) in the case of elastase and 1 micromole of the substrate hydrolyzed in 1 min per 1.0 mL of the supernatant (µmol/mL/min) in the case of cathepsin G. Statistical analysis was performed using paired Stu dent t test.

Among the oxygen dependent mechanisms of bac tericidal activity of neutrophils, an important role is played by sMPO, which, similarly to elastase and cathepsin G, is contained in the azurophilic granules. Secreted MPO catalyzes the formation of hypochlor ous acid and other bactericidal products [8]. In addi tion, MPO is involved in the inactivation of granule contents and reduces the binding of chemoattractants to specific receptors, which hampers the process of