Study of Possible Mechanisms Involved in the Inhibitory Effects of Coumarin Derivatives on Neutrophil Activity

Table 2

Doses of the compounds tested producing 50% inhibition of control extracellular CL and intracellular CL of human neutrophils and cell-free CL system.

Chemiluminescence	HHC	HHDC
	EC50 (μmol/L)	EC50 (μmol/L)
Extracellular	1.04±0.20	1.01±0.13
Intracellular	5.24±0.57	>100
Cell free	0.59±0.01	2.47±0.04

Human isolated neutrophils (5 × 10⁵/sample) were stimulated with PMA (0.05 μmol/L) at 37°C, in the presence of HHC or HHDC (0.01–100 μmol/L). Extra- and intracellular ROS production were measured by isoluminol-/luminol-enhanced CL. In the cell-free system (luminol, horseradish peroxidase, and hydrogen peroxide), CL responses were measured in the presence of HHC or HHDC (1, 10, or 100 μmol/L) at 37°C. Percentage inhibition was calculated on the basis of integrated values of CL over 1800 s (extra- and intracellular CL) and of cell-free CL over 10 minutes. Mean ± SEM, 𝑛=3–8. Control values given in RLU × seconds were 5.8 × 10⁶ ± 1.4 × 10⁶ for extracellular CL, 5 × 10⁵ ± 0.5 × 10⁴ for intracellular CL, and 3.8 × 10⁵ ± 0.18 × 10⁴ for cell-free CL.