Comprehensive analysis of the factor h binding capabilities of borrelia species associated with lyme disease: delineation of two distinct classes of factor h binding proteins

Some Lyme disease spirochete isolates can bind complement regulatory protein factor H (fH), a process that may allow evasion of complement-mediated killing. Here we demonstrate significant differences in the fH binding capabilities of species of the Borrelia burgdorferi sensu lato complex. The percentages of B. burgdorferi, B. afzelii, and B. garinii bacteria that bound fH in either enzyme-linked immunosorbent assays or affinity ligand binding immunoblot assays were 100, 83, and 29%, respectively. The fH binding protein profiles were examined and found to exhibit variability among isolates and to form two distinct classes. Differences in fH binding ability may contribute to the differences in pathogenesis and clinical course observed upon infection with different species of the B. burgdorferi sensu lato complex.     

A new function of the activated third component of complement: binding to C5, an essential step for C5 activation

Incubation of factor B, factor [unk]D, (properdin) and C3b—or of C1[unk]s, C4b and C2—with C3 and/or C5 in a fluid system leads to the generation of C3 cleaving activity while C5 remains unaffected. C5 can be cleaved—activated—when in addition to the enzyme-generating components of either pathway C3b is present which has been fixed to a solid surface immediately after its release from C3 by a surface-fixed enzyme. This has been demonstrated with three C3b-carrying solids: agarose to which C3b was fixed after cleavage of C3 by agarose-coupled trypsin (Ag-C3b), zymosan incubated with human serum (Z-C3b) and sheep red cells in the state EAC43. C3b fixed to these surfaces after release in the fluid phase by soluble enzymes does in general not support C5 cleavage. C3b species active in C5 cleaving processes differ from those inactive by having a newly discovered property: active C3b is capable of reversibly binding C5. Fluid C3 convertase C3b[unk]B cleaves C5 in the presence of surface-fixed active C3b also under conditions under which it cannot interact with the latter. This indicates that two C3b molecules having different functions are involved in this system, one which is incorporated in the enzyme complex and another which binds and thereby prepares C5 for cleavage. The binding requires a special configuration of C3b which is preserved only by fixation immediately after its generation from C3. Efficient binding is possible only when C3b is free of other ligands such as factor B or properdin; these components interfere with binding and cleavage of C5 when having access to active C3b. C5 cleavage by the convertase of the classical complement pathway, C42, appears to proceed by the same mechanism, i.e. free C42 per se attacks C5 when this is bound to active C3b. It is concluded from the results that C5-cleaving complement enzymes do not differ in composition from C3 convertases. The active C3b which is essential for C5 cleavage, and existent only on surfaces, serves to modulate the substrate configuration to make it accessible to the enzyme.     

Requirement for an additional serum factor essential for the antibody-independent activation of the classical complement sequence by Gram-negative bacteria.

Killing of Salmonella minnesota and Salmonella typhimurium S and R strains in serum of nonimmune humans and guinea pigs was drastically reduced in the selective absence of C1q, C1r, Ca2+, C4, or C2, the components of the classical complement pathway. Binding of C1 and C1q to the S form and six different core-deficient R mutant strains became stronger the shorter the lipopolysaccharide molecule. C1 and C1q had, under physiological conditions, no affinity to the serum-resistant S forms, whereas these components were bound by the serum-sensitive R forms with high affinity. However, a mixture of the individual complement components C1-C9, which rapidly lysed sensitized erythrocytes, did not kill the serum-sensitive bacteria. Isolated C1 bound to these bacteria cleaved fluid-phase C4 but did not convert C2. C2 turnover could be detected only when serum was used as a source of C1 or C4, indicating that an additional serum component is necessary for the antibody-independent bactericidal effect. Functional tests indicated that this factor is a euglobulin which mediates binding of C4 to the bacteria even in the absence of C1 or after treatment with EDTA. Binding of C4 followed by the generation of C4b sites as acceptors for C2 was a prerequisite for the killing of the bacteria. The factor could not be replaced by immunoglobulin G or immunoglobulin M, nor was it blocked by preincubation with anti-immunoglobulin G or anti-immunoglobulin M.     

Expression of an activation antigen, Mo3e, associated with the cellular response to migration inhibitory factor by HL-60 promyelocytes undergoing monocyte-macrophage differentiation

HL-60 promyelocytic cells acquire the surface expression of the Mo3e antigenic determinant after exposure to PMA or compounds that raise intracellular concentrations of cyclic AMP (dibutyryl cyclic AMP or a combination of cholera toxin and IBMX). The expression of Mo3e by these stimulated HL-60 cells coincides with the development of features of monocyte-macrophage differentiation (characteristic morphology, nonspecific esterase activity, and respiratory burst activity). During in vitro monocyte-macrophage differentiation, HL-60 cells become responsive to migration inhibitory factor (MIF); the MIF responsiveness of differentiated HL-60 cells is blocked by anti-Mo3e monoclonal antibody. These findings further support the relationship between the expression of Mo3e and the cellular response to MIF.