Loss of autoantibody activity by alteration in autoantigen.

C3 nephritic factor (C3NeF) is an autoantibody produced by patients with membranoproliferative glomerulonephritis (MPGN); it is thought to be responsible for the continued breakdown of C3 in these patients. We have studied three patients with MPGN for periods of 7 to 17 years who were noted to develop a normal C3 and factor B level in the face of persistent circulating C3NeF. In addition, C3NeF obtained from other sera were inactive when added to these patients' sera. When C3NeF was isolated from these patients' sera, however, it was completely active when added to normal human serum as a source of C3 and factor B. Removal of the C3 and factor B from these patients' sera by passage over anti-C3 and anti-factor B columns followed by reconstitution with C3 and factor B isolated from normal serum allowed C3NeF to be active in these sera. Replacement of C3 alone did not restore activity to these sera but replacement with factor B alone restored full C3NeF activity. These data suggest some alteration in autoantigen (factor B) as a mechanism for reduction in autoantibody (C3NeF) activity.

Nucleotide sequence of a human autoantibody to the alternative pathway C3/C5 convertase (C3NeF).

The production of autoantibodies to the alternative pathway C3/C5 convertase or C3 Nephritic Factor (C3NeF) is one characteristic of membranoproliferative glomerulonephritis. The complete nucleotide sequences of the heavy and light chain variable regions of an IgG C3NeF produced by an EBV transformed B cell line derived from a patient with membranoproliferative glomerulonephritis were determined. The VH and VL gene segments used by this C3NeF are extensively mutated suggesting that antigenic selection and affinity maturation may occur during the generation of these autoantibodies.

On the origin of C3 nephritic factor (antibody to the alternative pathway C3 convertase): evidence for the Adam and Eve concept of autoantibody production.

The antibody to the alternative pathway C3 convertase, designated C3 nephritic factor or C3NeF, is an autoantibody that is produced in everyone from the time of birth. The elaboration of C3NeF utilizes germline V-region genes which undergo antigen-driven affinity maturation, resulting in an autoantibody that is produced in large amounts with high affinity and narrow specificity. Our data also suggest that under normal conditions, the idiotypic network may play an important part in the control of this autoantibody. Further, a defect in the network with loss of control or inappropriate stimulation may be an underlying mechanism in the unrestricted production of C3NeF in patients with membranoproliferative glomerulonephritis.

Long-term follow-up of non-systemic lupus erythematosus glomerulonephritis in patients with hereditary angioedema: report of four cases.

Hereditary angioedema (HAE) is characterized by a deficiency in C1 inhibitor protein (C1 INH) and by clinical symptoms of episodic swelling of subcutaneous or mucosal tissue. It has rarely been reported in association with non-systemic lupus erythematosus (SLE) glomerulonephritis (GN). A recent report of two cases indicates the prognosis to be poor, with both patients progressing to chronic renal failure 8 and 20 years after diagnosis. This report describes the 5-year follow-up of a previously unreported case of an 8-year-old boy with HAE and non-SLE membranoproliferative glomerulonephritis (MPGN). The patient developed macroscopic hematuria, azotemia, and a vasculitic rash. Treatment included prednisone and cyclophosphamide, resulting in clinical improvement. The present report also summarizes the long-term follow-up of three previously reported cases of HAE and non-SLE GN, 25, 16, and 10 years after their initial presentation. Patients monitored for 25 and 16 years had MPGN and normal renal function and received no therapy. The third patient, monitored for 10 years, had segmental MPGN. This patient presented with urinary abnormalities and, after treatment with prednisone, had improvement in her hematuria. None of these four patients developed chronic renal failure. These observations indicate a variable outcome in patients with HAE and non-SLE GN.

Study of the idiotypic response to autoantibody to the alternative pathway C3/C5 convertase in normal individuals, patients with membranoproliferative glomerulonephritis, and experimental animals.

We studied the fluctuation of autoantibody to the alternative pathway C3/C5 convertase (C3NeF) and its autoantidiotypic antibodies, Ab2 alpha and Ab2 beta, in normal individuals, patients with membranoproliferative glomerulonephritis (MPGN), and New Zealand white rabbits. In normal individuals, serum levels of Ab2 alpha (anti-id Ab without internal imagery to the native antigen) are several times higher than those for Ab2 beta (anti-id Ab bearing the internal image of the native antigen). When normal rabbits are immunized with Ab2 beta, Ab3 is produced which has strong C3NeF activity. Ab3 acts to stimulate the prompt production of Ab4 alpha. Ab3 (C3NeF) titers then fall, followed by the appearance of Ab4 beta. Patients with MPGN and C3NeF activity were also studied. Untreated patients at the time of diagnosis have a two- to fourfold predominance of Ab2 beta which is a direct reversal of the normal situation. When the patients were treated with prednisone, C3NeF levels fell and Ab2 alpha again predominated. These data suggest that the idiotypic network acts to control the production of autoantibody in a balanced fashion. Moreover, these data suggest that the patients' response to Ab1 is quite different than that found in normal individuals.

Autoantibody to the alternative pathway C3/C5 convertase and its anti-idiotypic response. A study in affinity.

In an effort to understand the development and control of autoantibody production, we studied the affinity of autoantibody to the alternative pathway C3/C5 convertase (C3 nephritic factor (C3NeF)) and its autoanti-idiotypic antibodies, Ab2 alpha and Ab2 beta. These were isolated and purified from newborns, normal adults, and patients with membranoproliferative glomerulonephritis. In all cases, both IgG and IgM C3NeF were available for study. The affinity of IgG and IgM C3NeF for their natural Ag (10(8) liters/mol) as well as for the internal image of that Ag displayed on Ab2 beta was high (10(10) liters/mol). Furthermore, the affinity of IgG C3NeF was nearly 100-fold higher in patients than in newborns, whereas there were no significant changes with IgM C3NeF. By contrast, there were not differences in the affinity of IgG Ab2 alpha (which does not display any likeness to the native Ag) from normal adults and patients to any C3NeF isolate. There was, however, a progressive increase in affinity between both Ab2 alpha preparations and IgG C3NeF from newborns, adult normal subjects, and patients, implying an alteration in C3NeF to account for the changes in affinity. These data suggest that Ag-driven affinity maturation occurs with autoantibody but may not occur within the idiotypic network. These data also indicate that as autoantibody affinity matures, it appears to modify its idiotype, perhaps in an effort towards autoregulation.

Human anti-idiotypic antibody responses to autoantibody against the alternative pathway C3 convertase.

Anti-idiotypic antibodies to autoantibody against the alternative pathway C3 convertase (C3NeF) were isolated and purified from normal human serum as well as from serum from six patients with membrano-proliferative glomerulonephritis (MPGN). All preparations of anti-id antibody blocked C3NeF deposition on EC3bBb as well as C3NeF stabilization of EC3bBb functional activity. The Ka of these ant-id antibodies for C3NeF was 10(9) liters/mol which is comparable to the Ka of C3NeF for its antigen. In addition, 90% of anti-id antibody isolated from patients with MPGN and 20% isolated from normal individuals resembled Bb and bound to C3b as well as to antibody specific for the Bb portion of Factor B. These anti-id antibodies also resembled C3b and bound to antibody specific for the C3c portion of C3b. Immunization of rabbits with this latter form of anti-id antibody led to the production of functionally active C3NeF. These data indicate that C3NeF anti-idiotypic antibodies exist in two distinct forms, with and without internal imagery of C3bBb, and can occur in both normal individuals and patients with MPGN.

Production of IgG and IgM autoantibody to the alternative pathway C3 convertase in normal individuals and patients with membranoproliferative glomerulonephritis.

To understand the origin of autoantibody production, we studied the ontogeny of antibody to the alternative pathway C3 convertase (C3 nephritic factor or C3NeF). Peripheral blood mononuclear cells from newborns, normal adults, and patients with membranoproliferative glomerulonephritis produced IgM and IgG C3NeF after culture for 14 days with pokeweed mitogen. Both IgM and IgG moieties appear to have the same paratope and are able to inhibit each other's binding and function. The affinity constant for each of the C3NeF molecules was moderately high (10(8) liters/mol) and there appeared to be little difference between the Ka values for the IgG and the IgM autoantibodies or between Ka values for autoantibodies isolated from newborns, adults, and patients. These data, then, indicate that the ability to produce C3NeF autoantibody is present from the time of birth in normal individuals. The high affinity of these autoantibodies under normal conditions suggests that C3NeF may play a more important physiological role than previously anticipated.

Evidence that production of autoantibody to the alternative pathway C3 convertase is a normal physiologic event.

The origin of autoantibody production was studied with the use of antibody to the alternative pathway C3 convertase (C3 nephritic factor (C3NeF), as a model. Pokeweed mitogen stimulation of peripheral mononuclear cells from newborn infants, normal adults, and patients with membranoproliferative glomerulonephritis indicated that the ability to make C3NeF is apparently present in everyone from the time of birth. In addition, C3NeF appeared to express a single or very limited idiotope (21/21 isolates). The data also suggest that the elaboration of C3NeF may approximate an antibody response after immunization. Thus the C3NeF fraction of the total IgG or IgM produced in culture by pokeweed mitogen-stimulated mononuclear cells from normal neonates and adults, as well as from patients, was in the range of the production of specific antibody. Further, both IgG and IgM C3NeF produced by cells from these normal individuals, including newborn infants, had an affinity for antigen (10(8) to 10(9) L/mol) that was also in the range of specific antibody. Most of the autoantibody molecules (5/7) from serum were IgG3; two B cell clones producing C3NeF were CD5-negative. These experiments indicate that unmutated germline genes are used in the production of C3NeF and that a limited spectrum of antiidiotypic antibodies regulate its production.

Human polyclonal and monoclonal IgG and IgM complement 3 nephritic factors: evidence for idiotypic commonality.

Complement 3 nephritic factors (C3NeF) were isolated from the sera of patients with membranoproliferative glomerulonephritis (MPGN) and the supernatants of pokeweed mitogen-stimulated mononuclear cells from patients with MPGN. Three human monoclonal C3NeF antibodies (two IgGs, CK and PH, and one IgM, K3C4) were established. Using an exhaustive series of affinity columns, we isolated anti-C3NeF idiotypic antibodies (anti-IdNeF) (three from normal and two from patient sera). Anti-IdNeF preparations bound to F(ab')2-NeF and prevented its ability to stabilize C3bBb convertase. We have used the above reagents to address questions on the genesis and the diversity of C3NeF antibodies. The following results were obtained: All anti-IdNeF preparations bound to C3NeF isolated from patient sera, cell culture supernatants, and IgG and IgM monoclonal C3NeF. None of the monoclonal C3NeF bound to an extensive battery of common antigens, including Fc portion of IgG, TNP, beta-galactosidase, DNA, and bacterial products. These data indicate that C3NeF express one common idiotype and that these antibodies are not raised in response to an obvious antigen.

On the origin and control of C3NeF.

Nephritic factors are a relatively recent addition to the growing list of autoantibodies. C3NeF was the first of these to be described and acts by stabilizing the alternative pathway C3/C5 convertase. IgG and IgM C3NeF as well as anti-idiotypic antibody to C3NeF, may be found in normal individuals at all ages. In addition, internal image anti-idiotypic antibody can also be demonstrated in patients with glomerulonephritis. Thus, C3NeF appears to fulfill all of the criteria for being a product of normal B-cell function. The potential dysregulation of this process and the role of C3NeF in glomerulonephritis remains theoretical at best.

Sonography of the kidneys in hemolytic uremic syndrome.

The sonographic appearance of kidneys in patients presenting with acute renal failure due to hemolytic-uremic syndrome (HUS) was evaluated and compared with that in an age-matched control series. In nine patients with HUS, the renal cortex was typically hyperechogenic and the renal pyramids appeared sonolucent. The renal size was normal or enlarged compared with 35 normal controls. While not specific, the sonographic patterns described in this study support the diagnosis of HUS in the appropriate clinical setting.

Components of the alternative pathway of complement in otitis media with effusion.

An evaluation of the alternative pathway of complement was undertaken in patients with otitis media with effusion (OME). Middle ear fluid (MEF) and serum specimens were obtained from 34 patients at the time of elective myringotomy for OME. Bacterial, viral, and mycoplasma cultures were made for all specimens of the fluids. Immunochemical determinations by radial immunodiffusion were performed for C3, C5, factor B, properdin, factor H, factor I, and albumin. Each patient's recent clinical course and past history were reviewed. The results of all viral and mycoplasma cultures were negative. Three of 55 bacterial cultures were positive for type B Haemophilus influenzae. All components of the alternative pathway measured were found to be present in varying amounts in MEF. When the levels of the complement components were compared to the clinical factors studied, there were no observable differences. These data suggest that components of the alternative pathway of complement are present in OME and are not useful in predicting the clinical course or outcome of this disorder.

An abnormality of the fourth component of complement associated with benign recurrent hematuria.

A group of 9 children with persistent isolated hematuria in the absence of other evidence of renal disease is presented. The children represent about one third of our patients with benign recurrent hematuria and are unique in that their C4s, in a functional assay, are lower than one would expect from their immunochemical level, resulting in an abnormally low C4 hemolytic efficiency. This feature occurs in the absence of any other evidence of complement dysfunction. Similar abnormalities were not seen in patients with other hypocomplementemic renal disease or congenital C4 deficiency. The patients' C4 molecules were identical to normal in charge, mobility, antigenic configuration, and ability to participate in an immune reaction. All patients had at least one C4 null gene on genetic analysis and 3 had double null genes. A similar pattern of C4 abnormality was not found in other patients with hypocomplementemic renal disease.

Evaluation of Streptococcus pneumoniae type XIV opsonins by phagocytosis-associated chemiluminescence and a bactericidal assay.

The relative roles of serum factors required for opsonization of type XIV Streptococcus pneumoniae were investigated by means of luminol-enhanced chemiluminescence (CL), bactericidal, and immunofluorescence assays employing adult sera containing high (>1,000 ng of antibody nitrogen per ml) or low (<200 ng of antibody nitrogen per ml) antibody concentrations as determined by radioimmunoassay. Specific antibody concentration correlated directly with both total and heat-labile CL activity (P < 0.005) and with the bactericidal index (P < 0.05) at a serum concentration of 10%. The importance of specific antibody as an opsonin was confirmed by the abolition of CL activity and immunoglobulin immunofluorescence observed after absorption of heated sera with type XIV pneumococcal cells and by the dose response in CL and bactericidal activity observed with the addition of immunoglobulin G to hypogammaglobulinemic serum. A role for the classical complement pathway in opsonization was indicated by significantly greater CL integrals for high-antibody sera than for low-antibody sera depleted of factor D and by the bactericidal activity noted for untreated, but not magnesium ethylene glycol-bis(beta-aminoethyl ether)-N,N-tetraacetic acid-chelated low-antibody sera. The alternative pathway contributed more than half of the CL activity of both high- and low-antibody sera. However, after magnesium ethylene glycol-bis(beta-aminoethyl ether)-N,N-tetraacetic acid chelation, only sera with high antibody concentrations or agammaglobulinemic serum reconstituted with immunoglobulin G with high specific antibody levels supported significant bactericidal activity. Therefore, type-specific antibody and complement promote opsonization of type XIV S. pneumoniae, and this may occur via either complement pathway. These results suggest that CL is a suitable tool to delineate serum factors and their contribution to opsonization, but results must be related to other functional assays.