Immune complexes isolated from patients with pulmonary tuberculosis modulate the activation and function of normal granulocytes

Abstract

Circulating immune complexes (ICs) are associated with the pathogenesis of several diseases. Very little is known about the effect of ICs on the host immune response in patients with tuberculosis (TB). The effects of ICs isolated from patients with TB in modulating the release of calcium, cytokines, and granular proteins were studied in normal granulocytes, as were their chemotactic, phagocytic, and oxidative burst processes. ICs from TB patients induced decreased production of cytokines and platelet-activating factor (PAF) from normal granulocytes. ICs from TB patients also induced enhanced chemotaxis and phagocytosis but caused diminished oxidative burst. This was accompanied by an increased release in intracellular calcium. On the other hand, ICs from TB patients induced increased release of the granular proteins human neutrophil peptides 1 to 3 (HNP1-3). Thus, ICs from patients with TB exhibit a profound effect on granulocyte function with activation of certain effector mechanisms and dampening of others.

Fig 1

Levels of ICs in serum samples from patients with TB and healthy volunteers. Serum samples from UNINF and INF subjects (10 subjects in each group) were treated with PEG 6000, and the absorbance of the precipitate was read at 280 nm using a spectrophotometer. CIC levels were determined by interpolation from a standard curve plotted using aggregated human gamma globulins as a standard. Each symbol represents the value for one individual. The horizontal line shows the geometric mean. The P values were calculated using the Mann-Whitney U test. UNINF, uninfected individuals; INF, infected patients (patients with TB).

Fig 2

PEG 6000-precipitated plasma ICs from patients with TB induce diminished production of cytokines from granulocytes. Granulocytes from healthy volunteers (n = 10) were stimulated with PEG 6000 precipitates of plasma ICs from UNINF and INF subjects (10 subjects in each group) for 18 h, and cytokines were measured by multiplex ELISA. The P values were calculated using the Mann-Whitney U test. UNINF, uninfected individuals; INF, infected patients (patients with TB). The geometric means (bars) and 95% confidence interval (error bars) are shown.

Fig 3

PEG 6000-precipitated plasma ICs from patients with TB induce increased release of HNP1–3 and decreased release of PAF from granulocytes. Whole-blood samples from healthy volunteers (n = 10) were stimulated with PEG 6000 precipitates of plasma ICs from UNINF and INF subjects (10 subjects in each group) for 4 h, and the granular proteins (HNP1–3) (A) and PAF (B) in the culture supernatants were measured using ELISA. The P values were calculated using the Mann-Whitney U test. UNINF, uninfected individuals; INF, infected patients (patients with TB).

Fig 4

PEG 6000-precipitated plasma ICs from patients with TB induce enhanced chemotaxis (A) and phagocytosis (B) of normal granulocytes. Whole-blood samples from healthy volunteers (n = 10) were stimulated with PEG 6000 precipitates of plasma ICs from UNINF and INF (10 subjects in each group) for 10 min, and chemotaxis and phagocytosis of granulocytes were measured using flow cytometry. The P values were calculated using the Mann-Whitney U test. UNINF, uninfected individuals; INF, infected patients (patients with TB).

Fig 5

PEG 6000-precipitated plasma ICs from patients with TB induce decreased oxidative burst of granulocytes. Whole-blood samples from healthy volunteers (n = 10) were stimulated with PEG 6000 precipitates of plasma ICs from UNINF and INF subjects (10 subjects in each group) for 10 min, and oxidative burst of granulocytes was measured using flow cytometry in response to PMA (A) or E. coli (B). The P values were calculated using the Mann-Whitney U test. UNINF, uninfected individuals; INF, infected patients (patients with TB).

Fig 6

PEG 6000-precipitated plasma ICs from patients with TB induce increased release of calcium from granulocytes. Granulocytes from healthy volunteers (n = 5) were stimulated with PEG 6000-precipitates of plasma ICs from UNINF and INF subjects (five subjects per group), and calcium release was assessed using a fluorimeter. Data are presented as a representative histogram (A) showing calcium release in response to ICs from one INF individual and one UNINF individual and as percentages of nontreated control (B) for which calcium-associated fluorescence was measured in parallel cultures. The P values were calculated using the Mann-Whitney U test. UNINF, uninfected individuals; INF, infected patients (patients with TB).