Studies on flow cytometric analysis of neutrophil functions of systemic lupus erythematosus
Watanabe, I.
Hokkaido Journal of Medical Science 67(2): 200-215
1992
ISSN/ISBN: 0367-6102 PMID: 1597301 Document Number: 398112
Flow cytometric quantitative methods of (1) phagocytosis and (2) oxidative product formation by neutrophils were investigated by using whole blood. The neutrophil function of systemic lupus erythematosus (SLE) patients was analyzed by these methods. Kinetics of phagocytosis were investigated by using 1.97 microns fluorescent carboxylated microspheres (Polyscience, USA). The rate of phagocytosis (P) was increased logarithmically according to the T/C (target particle-to-cell) ratio (P = K1 x log (T/C ratio) +K2 K1, K2: constant). The value of Ph 50 were defined as the value of T/C ratio at which rate of phagocytosis was 50%. The PH 50 in SLE patients were statistically higher than in normal controls, which indicated the decreased phagocytic activity in SLE. Active SLE serum increased Ph 50 more than inactive SLE serum, and heat inactivated normal serum (30 min at 56 degrees C) increased Ph 50. Amount of oxidative products of neutrophils was measured by using dichlorofluorescein (DCF), which was converted from dichlorofluorescin diacetate (DCFH-DA) in the cells. After pre-incubation in room temperature from sampling to assay, the DCF fluorescent intensity of neutrophil of a normal control was low and revealed one-peak histogram, but that of SLE neutrophils revealed two peaks in histogram. The 2nd peak of high fluorescent intensity was thought to be composed of the subpopulation of neutrophil. The rate of the cells with oxidative products in unstimulated neutrophils was higher in SLE patients than in normal control and correlated with some clinical markers of disease activities. These two flow cytometric methods were rapid and accurate, and the whole blood could be used without separation of neutrophils by these methods. These methods might be useful technique to investigate the neutrophil functions.