Serum-Dependent Phagocytosis of Paraffin Oil Emulsified With Bacterial Lipopolysaccharide

Abstract

The ability of fresh serum to opsonize microorganisms was recognized nearly 70 yr ago (1). Subsequent studies identified synergistic effects of both antibody and complement on opsonizatiou (2) and demonstrated that particles to which C3 was affixed by the sequential participation of antibody, C1, C4, and C2 became opsonized (3-6). In 1968, a patient was found with marked susceptibility to infection who spontaneously inactivated C3 in vivo. His serum had normal functional levels of antibody, CI, C4, and C2, but failed to opsonize bacteria even when fortified with purified C3 (7). Further studies of his serum revealed a defect in his properdin pathway (8, 9). This patient, and the good health of humans deficient in C2 (10) and of guinea pigs deficient in C4 (~ 1), have emphasized the functional significance of this alternate pathway of C3 activation, first recognized by Pillemer and his associates (12). The properdin system has been shown to participate in bacteriolysis, protozoal inactivation, virus neutralization, and lysis of erythrocytes in paroxysmal nocturnal hemoglobinuria (13-16). Unlike the classical complement pathway, the properdin system can be activated by polysaccharides in the absence of antibody (17).This report describes quantitative studies of the phagocytosis of paraffin oil droplets coated with bacterial lipopolysaccharide. Opsonization of these particles occurs in the absence of C4 and C2, but is dependent upon C3 and the properdin pathway. Measurement of the opsonization of these particles thus constitutes a quantitative functional assay for this system. Materials and MethodsPreparation of Phagocytes.--Suspensions containing over 95% polymorphonuclear leukocytes were obtained from guinea pig peritoneal exudates. The exudates were elicited with sodium caseinate, and the cells were collected and washed as previously described (18). Human peripheral blood from normal laboratory personnel or patients with infections and leukocytosis was collected with acid-citrate dextrose, National Institutes of Health formula A; the erythrocytes were removed by dextran sedimentation and lysis with ammonium chloride; and leukocytes were washed as described previously (19). The celts were suspended in KrebsRinger phosphate medium, pH 7.4 (medium), at a concentration of 2-5 mg of guinea pig

Pathogenesis of Foreign Body Infection: Description and Characteristics of an Animal Model

“…S. aureus strain Wood 46 was used in all experiments to avoid interference of staphylococcal protein A with IgG; this strain has been repeatedly shown to contain negligible amounts of protein A [26,34] and to require complement only for "optimal opsonization [13,24,26]. Although several biochemical determinations in tissue cage fluid differed from values in serum (data not shown), their variations were not sufficient to inhibit phagocytosis [35][36][37][38][39]. Such an effect, as well as a deleterious effect of tissue cage fluid on PMNLs, was further ruled out by appropriate mixing experiments involving preincubation of tissue cage fluid with competent blood PMNLs.…”

Phagocytic activity and hyperpolarizing responses in L‐strain mouse fibroblasts.

“…External high Ca2+ markedly enhanced the uptake of U.V.-irradiated beads (Table 1). Such an enhancing effect of Ca2+ on ingestion was also found in leukocytes (Greendyke, Brierty & Swisher, 1963;Stossel, Alper & Rosen, 1973) and in macrophages (Rabinovitch, 1967;Mason, Stossel & Vaughan, 1973;Stossel, 1973). Contrary to these 'professional' phagocytes, however, external Mg2+ at a high concentration significantly inhibited the uptake of beads in L cells (Table 1).…”

Influence of a deficiency of the second component of complement on the bactericidal activity of neutrophils in vitro.

“…The basis for the conflicting evidence has usually been attributed to methodological differences. Published studies have employed a wide variety of phagocytic cells, test particles, and assay systems (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). A comparison of the findings from these studies indicates that these modifications of experimental design have influenced the results.…”

“…The specific requirements for these agents in providing optimal phagocytosis and killing of bacteria are not completely understood (1)(2)(3)(4)(5)(6)(7)(8)(9). Considerable controversy exists regarding the necessity for early components of complement for the opsonization of bacteria (11)(12)(13)(14)(15)(16)(17)(18). The basis for the conflicting evidence has usually been attributed to methodological differences.…”