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Antibodies to surface determinants of human lymphocytes, recognized both by cytotoxicity of fluorescent antibody analysis, were shown to be specifically enriched over the serum levels in cryoprecipitates from patients with systemic lupus erythematosus (SLE). The antilymphocyte antibody was shown to be cold reactive and was exclusively IgM. It was distinct from IgM anti-IgG, which was also variably concentrated in the cryoprecipitates. The question whether the antilymphocyte antibodies appear in the cryoprecipitates as complexes because of interaction with surface membrane antigens, or simply because of cold reactive properties, remains to be determined. The possible clinical relevance of the cryoprecipitation of these antibodies in systemic lupus erythematosus is discussed.

Members of the American Rheumatism Association (ARA) were surveyed to determine their views on the content and quality of their annual meeting. Responses were received from 1,208 members. ARA members were generally positive about the major components of the annual meeting program, but there were significant differences in opinion between practitioners and academicians. Practitioners favored a reduction in basic science material and an increase in didactic clinical sessions. Academicians were more likely to feel that recent annual meetings have adequately addressed their educational needs. These findings have implications for the planning of future annual meetings of the ARA.

Using indirect immunofluorescence and flow cytometry, we determined the proportion and number of T3+, T4+, and T8+ cells in the peripheral blood of patients with systemic lupus erythematosus whose sera were positive for cold-reactive antilymphocyte antibodies versus values in patients whose sera were negative for these antibodies. There was a disproportionate reduction in T4+ peripheral lymphocytes when cold-reactive antilymphocyte antibodies preferentially cytotoxic for this subpopulation were present in autologous serum. The decrease in this subset was responsible for a reduction in the T4:T8 ratio; variation in the number and proportion of T8+ cells was insignificant. A similar, but autoantibody-independent, alteration in the T4+ subpopulation was found in patients who were receiving prednisone therapy. A relationship between T cell population abnormalities and systemic lupus erythematosus disease activity, per se, was not observed.

Systemic lupus erythematosus (SLE) is characterized by increased numbers of circulating B cells activated polyclonally to secrete immunoglobulin. Because T cells secrete, or shed, various factors that are functionally important in regulating immunoglobulin production by B cells, a reverse hemolytic plaque assay was developed to quantitate such activated T cells. In this technique, we used a rabbit antiserum raised to supernatants of concanavalin-A--stimulated human lymphocytes. The relevant antigenic specificity of this antiserum is directed toward the shed surface membrane determinant(s) preferentially expressed on activated T cells. Freshly isolated peripheral blood mononuclear cells from 14 SLE patients contained more than 10 times the number of endogenously activated T cells than cells from normal subjects. Within the SLE group, plaque-forming T cells were particularly increased in patients with active disease. By linear regression analysis, a significant positive correlation was revealed between such activated T cells and immunoglobulin-secreting B cells, also measured by a reverse plaque assay (r = 0.83). It appears that both activated B cells and T cells circulate in increased numbers in SLE. Additional investigation will be required to define the molecular nature of the T cell product(s) being measured and to clarify the relationship of these findings to the immunoregulatory abnormalities in this disorder.

Brief digestion of human peripheral blood lymphocytes by vibrio cholera neuraminidase (VCN) revealed hidden components of the membrane. Autologous human serums contained antibodies directed to these components that were readily demonstrated by immunofluorescence. Antibodies of similar specificity were found in all normal serums. The antibodies were principally of the IgM variety with lesser amounts of the IgG class present. They were equally active at 4 degrees C and 37 degrees C. The VCN revealed membrane determinants were present in normal B and T lymphocytes, monocytes, lymphocytes of patients with chronic lymphatic leukemia and cells of lymphoid lines. The newly revealed determinants slowly disappeared upon culture of the lymphocytes. These hidden components were similarly demonstrated in erythrocyte membranes and represent the T antigen long known for the red blood cells. Absorption by either VCN treated autologous lymphocytes or erythroyctes removed all of the antibodies capable of reacting with both cell types. Absorption by VCN digested isologous lymphocytes removed all reactivity with autologous lymphocytes.

Cold-reactive lymphocytotoxic autoantibodies are present in the serum of most patients with active systemic lupus erythematosus (SLE) and may be important for the development of the lymphopenia and T cell dysfunction characteristic of this disorder. Neither the mechanisms of autoantibody action in this regard, nor the nature of the relevant T cell membrane target molecules have been defined, however. In the present investigation, preincubation of T cells with SLE serum at 37 degrees C reduced their reactivity with SLE IgM anti-lymphocyte autoantibodies, as demonstrated by indirect immunofluorescence and complement-dependent cytotoxicity. Modulation was restricted to SLE IgM autoantibody-reactive antigen; monoclonal antibody staining of various T cell differentiation and activation antigens remained unchanged. Loss of antigen from the surface membrane was rapid, but transient. A nadir was reached after approximately 120 min of 37 degrees C incubation, followed by essentially complete reexpression of antigen several hours later. Although modulation occurred spontaneously at 37 degrees C in the absence of SLE serum, loss of antigen was enhanced by IgM anti-lymphocyte autoantibodies, despite their low thermal amplitude. Modulation was inhibited by sodium azide, by fixation of cells with paraformaldehyde, and by low incubation temperatures. Colchicine and cytochalasin D had no effect on this process, suggesting that the integrity of the cytoskeleton was not essential. Cycloheximide did not prevent loss of antigen, but inhibited its reexpression. In experiments to determine the fate of modulated antigen, both intracytoplasmic accumulation and shedding from the cell surface were demonstrated. Only shedding was increased by the presence of anti-lymphocyte antibodies, however. These studies delineate modulation of T cell membrane antigen as a new mechanism for anti-lymphocyte autoantibody action in SLE. The occurrence of modulation at physiologic temperatures in vitro suggests that a similar phenomenon of potential relevance to T cell dysfunction may obtain in patients with this disorder.

Serologic studies were performed on 25 patients with systemic lupus erythematosus (SLE) during 29 acute episodes of central nervous system (CNS) disease. Increased anti-DNA antibody and decreased total serum hemolytic complement activity were observed only in those patients with associated extra-CNS disease manifestations. Patients with isolated CNS disease were otherwise in apparent clinical and serological remission regarding these two indices. No special association of cold-reactive IgM antilymphocyte antibodies was demonstrable in patients with ongoing CNS injury. Of special interest was an increased incidence of anti-Sm antibodies in the patients with CNS dysfunction relative to that in a large group of patients without neuropsychiatric disease. The incidence of anti-RNP was not increased. The data do not support direct involvement in SLE brain injury of either DNA/anti-DNA complexes or of lymphocytotoxic antibodies cross-reactive with brain cells, but do suggest an association of anti-Sm with CNS disease in this disorder.

Large percentages of the lymphocytes from some patients with rheumatoid arthritis and systemic lupus erythematosus were densely covered with Ig demonstrable by immunofluorescence, which was occasionally present in the form of caps. The amount and character of the Ig staining depended largely on the procedures used in the isolation and washing of the lymphocytes. Cold-reactive antilymphocyte antibodies present in many sera wre primarily responsible for these variations. Overnight culture of the lymphocytes proved to be an efficient procedure for the removal of adsorbed antibody. Some evidence was also obtained for the presence of circulating immune complexes and exogenous rheumatoid factor molecules on the lymphocyte surface. Thus on freshly isolated cells the demonstration of surface Ig proved to an unreliable marker of bone marrow-derived (B) cells in these disease: the actual percent of B cells with intrinsic surface Ig was often markedly decreased. In patients with systemic lupus erythematosus, this reduction was in agreement with the low numbers of cells that had a receptor for aggregated IgG. The mean percentage of thymus-derived (T) cells in both diseases was slightly greater than the normal level.The concentrations of lymphocytes in joint fluids from patients with rheumatoid arthritis were often greater than levels found in blood. T cells primarily accounted for this increase. The T cells typically formed unusually dense rosettes with sheep erythrocytes. B lymphocytes were proportionally much diminished. Evidence was obtained for the existence of a major joint fluid lymphocyte population that lacked all assayed surface markers.

Antilymphocyte antibodies in serum from patients with systemic lupus erythematosus (SLE), as detected by microcytotoxicity and indirect immunofluorescence, were predominantly cold reactive and of the IgM class. These IgM antibodies were most active at 4 degrees C. IgG antibodies were infrequent, and were only minimally lymphocytotoxic. Most sera were cytotoxic for autologous lymphocytes and were equally reactive with normal and SLE lymphocytes, as well as with B- and T-cell preparations. Separate T- and B-cell specificities, which appeared not to be related to HL-A determinants, were identified by differential absorption experiments. The functional significance of these antilymphocyte antibodies is discussed.

IgG anti-T cell autoantibodies are common in SLE serum, react preferentially with activated lymphocytes, and exert early-phase inhibitory effects on antigen-induced T cell proliferation. Little is known about the target molecules in this system, however, because the low titer and low avidity of the most interesting antibodies limit their utility in conventional immunoprecipitation analyses. Therefore, Western blotting was used to demonstrate binding of IgG in anti-T cell antibody-positive SLE sera to four surface membrane molecules shared by peripheral T cells and HSB-2 cells. Molecules of Mr 90,000 and 55,000 were particularly reactive: each target was stained by IgG anti-lymphocyte antibodies in 11 patient sera (approximately 85%) in the panel. Targets of Mr 37,000 and 105,000 were encountered less frequently (six of 13 and one of 13 patients, respectively). It is unlikely that alloantibodies contributed to the staining patterns observed because reactivity with the four targets was consistently present when cell preparations from multiple unrelated donors were examined. The target molecules were localized to the plasma membrane by whole cell absorption/elution experiments, by the failure of chromatin (DNA/histone) to absorb antibodies to these antigens, and through the use of purified membranes as substrate for Western blotting. With the possible exception of the 105,000 Mr molecule, which is a major target in the IgM anti-T cell antibody system, evidence for the existence of neoantigens as a basis for increased reactivity of SLE IgG with activated T cells was not obtained. The identity of the IgG antibody-reactive molecules with respect to known T cell antigens was not determined, although evidence against the existence of antibodies to Tac (IL 2 receptor) and the transferrin receptor was obtained in monoclonal antibody pre-clearing experiments. Nonetheless, the observation that a limited number of major IgG autoantibody target antigens on activated peripheral T cells are shared by HSB-2 cells, a primitive T cell line expressing few of the differentiation antigens characteristic of mature T cells, should provide a basis for more definitive characterization of antigens in this system in the future.

A young woman ingested approximately 13 gm of an over-the-counter acetaminophen preparation and died seven days later in profound coma. Postmortem findings included massive centrilobular hepatic necrosis and renal tubular necrosis. Although hepatotoxicity secondary to acetaminophen overdose is well recognized in Britain, this entity has received little emphasis in the United States. Recognition of the potentially severe consequences is important because early treatment with sulfhydryl compounds may prevent serious liver injury.

To assess the role of genetic factors in systemic lupus erythematosus (SLE), 12 twon pairs (seven definitely monozygotic, three definitely dizygotic) of which one or both twins had SLE, were studied and compared to 17 twin pairs (12 definitely monozygotic) previously described. In the present series, four of seven (57 per cent) definitely monozygotic pairs were clinically concordant for SLE, satisfying the preliminary criteria of the American Rheumatism Association (ARA). Concordance for the presence of antinuclear factor (ANF) and hypergammaglobulinemia was 71 and tinuclear factor (ANF) and hypergammaglobulinemia was 71 and 87 per cent, respiectively. These data closely agree with those on the 12 definitely monozygotic sets previously described. All three of the dizygotic sets in the present series were discordant for clinical SLE, although one clinically well twin had marked serologic abnormalities. Comparison of these data with thos from other first degree relatives of out twins clearly suggests a strong genetic component in the pathogenesis of SLE. The relative contribution of nongenetic and environmental factors to the expression of the disease is discussed.

Neonatal rats born of and nursed by mothers immunized with Torpedo acetylcholine receptor protein developed a defect of neuromuscular transmission as indicated by reduced miniature endplate potential amplitudes. It is likely that antibodies to the Torpedo receptor protein were passively transferred to the neonates in the milk. With the exception of the route of transfer, this neonatal form of experimental autoimmune myasthenia gravis appears to be similar to its human counterpart, and thus can serve as an experimental model.

Patients with systemic lupus erythematosus frequently develop antilymphocyte antibodies as measured by complement-dependent cytotoxicity and immunofluorescence assays. Highest titres of both of the major IgM and IgG classes occur during phases of active disease, and their presence is associated with essentially the entire spectrum of immune system functional abnormalities in this disorder. While the full range of antibody specificities requires further clarification, antibodies to many discrete lymphocyte populations have been described, including B cells, T cells, and T cell subsets. Antibodies to T cell subsets are of special interest because of their relationship with subset depletion in vivo, and their capacity to reproduce, through effects on normal cells in vitro, the same types of immunoregulatory abnormalities characteristic of lymphocytes isolated from patients with SLE. Suppressor/inducer and suppressor/effector T cells appear to be the main targets in this regard. Antibodies specific for activated T lymphocytes exist as well, and this type has the unusual property of interfering with events operant in production of/response to interleukin-2, a critical step controlling the expansion of specifically-reactive T cells and the induction of other lymphokines. In addition to complement-mediated lysis and antibody-dependent cell-mediated cytotoxicity, anti-lymphocyte antibodies have the potential to influence immune system function by several non-cytotoxic mechanisms, including surface antigen modulation and ligand/receptor triggering. Despite the large amount of data which has been accumulated concerning the cell type specificity and functional effects of anti-lymphocyte antibodies in SLE, little is known about the nature of the surface membrane molecules with which they react. Application of cell cloning and molecular biology technology should rectify this deficiency in the near future. Although it is likely that antilymphocyte antibodies are of relevance to immune system pathophysiology in SLE, it remains to be determined whether these interesting antibodies reflect secondary events, or have some more fundamental significance.

The influence of genetic and nongenetic factors on abnormalities of the immune response in systemic lupus erythematosus (SLE) was studied in 9 sets of twins in which one or both twins had SLE. Particular emphasis was placed on contrasting the results between the sibs of 3 monozygotic pairs discordant for clinical SLE. Depression of cell-mediated immunity, determined by lymphocyte blastogenic response to mitogens, was associated with the clinical expression of illness. In contrast, autoreactive antilymphocyte antibodies and lymphocyte tubuloreticular structures were found in both clinically affected and unaffected subjects and were more prominently associated with the presence of other serologic abnormalities. No evidence of antigens or cell surfaces determinants unique to affected sibs was encountered.

Rabbit antiserum to culture supernatant of concanavalin A-stimulated peripheral blood mononuclear cells was used in a reverse hemolytic plaque assay to detect activated T lymphocytes in human peripheral blood. The number of plaque-forming cells in fresh, unstimulated lymphocyte preparations was approximately 400/10(6) cells, and increased 7- to 14-fold after stimulation with a variety of mitogens and antigens. The kinetics of the increase paralleled 3H-thymidine incorporation, with a maximum on day 3 of culture. Plaques were eliminated by treatment of cells with anti-Leu-I + complement or by depletion of E-rosettes. The activated T cells were not restricted to a given inducer, suppressor, or Ia+ T cell subset, however. Both mitogen-stimulated culture supernatants and mitogen-activated lymphocytes, but not resting lymphocytes, were effective in absorbing the capacity of the rabbit antiserum to develop plaques. This suggested that the predominant specificity of this antiserum was directed toward surface membrane determinant(s) of T cells actively shed into the medium. This process was shown to require ongoing protein synthesis, and, in mitogen-stimulated lymphocyte preparations, DNA synthesis as well. This approach has enabled detection of a surprisingly large number of endogenously activated T cells in normal human blood, and should be useful for the analysis of immunoregulatory events at the single-cell level.

Monoclonal antibodies to known surface antigens on B cells and on resting and activated T cells of various types were used in several approaches to examine the specificity of IgM antilymphocyte antibodies in systemic lupus erythematosus (SLE). Surface determinants that were sought included: T3, T11, Leu-1, Leu-8 (pan-T); T4, T8 (T subset); beta 2-microglobulin (beta 2m); L243, Leu-10 (DR and DS/DC framework, respectively); anti-Tac (interleukin-2 receptor); 5E9 (transferrin receptor); and 4F2, AA1 (other activation antigens). The first strategy was based on inhibition of rosette formation between mouse monoclonal antibody-coated targets and anti-mouse IgG-coated erythrocytes by SLE sera, either directly at 4 degrees C or after modulation of IgM antilymphocyte antibody-reactive target cell antigen at 37 degrees C. Significant rosette inhibition, defined as greater than 2 standard deviations from the mean value for 10 control sera, was seen only for beta 2m (13 of 20 SLE sera were positive; inhibition = 15-58%). Next, relative fluorescence intensity of lymphocyte staining by monoclonal antibodies was assessed by flow microfluorometry after preincubation of cells with SLE serum at 4 degrees C or after modulation of SLE antibody-reactive antigen. Modulation markedly reduced or eliminated SLE antilymphocyte antibody IgM staining. Except for beta 2m, neither cold nor warm temperature preincubations altered the relative fluorescence intensity for the known surface antigens. These data confirm anti-beta 2m as a common antibody specificity in SLE and suggest that antilymphocyte antibodies in this disorder are not directed to Ia or to certain other defined lymphocyte antigens of functional interest.

Anti-lymphocyte antibodies have been demonstrated in patients with SLE and RA by immunofluorescence. They differ from those arising through iso-immunization by being cold reactive and chiefly of the IgM class. Separate anti specificities were shown for determinants on either T or B cells.

One of the fundamental immunologic characteristics of systemic lupus erythematosus (SLE) is a depressed T cell proliferative response to various specific and nonspecific stimuli. Both intrinsic cellular defect(s) and inhibitory influences of humoral factors, e.g., antilymphocyte autoantibodies or immune complexes, have been postulated to underly this functional abnormality. Because patient serum can induce SLE-like T cell dysfunction in normal cells, an extrinsic mechanism is probably responsible, but the nature and site of action of this humoral activity has not been defined. This laboratory recently described a novel antibody specific for activated T cells in SLE, which raised the possibility that suppression of T cell proliferation by SLE serum involved antibodies directed to surface determinants expressed during the process of activation. In experiments to examine this concept further, relatively warm-reactive antibodies to T cell blasts were found to inhibit strongly the well-characterized T cell response to tetanus toxoid. These antibodies were distinct from conventional cold-reactive IgM antibodies to resting T cells, which exhibited little inhibitory activity. Inhibition involved noncytotoxic effects on early activation events at the level of the responding T cell, which markedly reduced the expression of receptors for interleukin 2. Inhibitory effects on antigen-pulsed macrophages or on T cells already committed to proliferate were not demonstrable. Anti-T blast antibodies were characteristic of active SLE and were detected only occasionally in patients with inactive disease or non-SLE rheumatic disorders. Although the exact antigenic specificity was not identified, considerable evidence was obtained against the presence of antibodies to Ia and certain other surface determinants of functional relevance. Our observations concerning the suppressive effects of anti-T blast antibodies in SLE serum on the T cell response to tetanus toxoid should provide new insight into mechanisms of in vivo T cell dysfunction in this and other immunologic disorders.

Artificial cryoprecipitates have been prepared from SLE sera by incubation of anti-double stranded DNA with 3H-PM2 DNA. IgM rheumatoid factor, and complement to study further the relationships between natural cryoprecipitation and the DNA/anti-DNA system in this disorder. Addition of rheumatoid factor and/or complement to a mixture of SLE serum and 3H-DNA results in increased precipitation of the 3H-DNA following incubation at 4 degrees C. Precipitation formed with added complement or complement plus rheumatoid factor could be rapidly resolubilized or reprecipitated by warming to 37 degrees C or recooling to 4 degrees C, i.e., exhibited the behavior of natural cryoprecipitates. The resolubilized cryoprecipitates contained excess antibody, because they bound significant quantities of additional 3H-DNA in the Farr assay. Of special interest was the observation that DNA binding activity frequently could be recovered from solubilized cryoprecipitates even though 3H-DNA was not added during the original cryoprecipitate preparation. With certain cryoprecipitations prepared in this way, the DNA binding activity was further increased by deoxyribonuclease treatment, raising the possibility that endogenous DNA may be constituent of such cryoprecipitates. The implications of these findings with respect to the nature of antibody-DNA immune complexes in SLE are discussed.

Fifteen serum cryoprecipitates from 28 patients with systemic lupus erythematosus (SLE) contained DNA demonstrable by counterimmunoelectrophoresis after exhaustive digestion of cryoimmunoglobulin with pronase. The majority of these cryoprecipitates also exhibited increased DNA binding activity in a modified Farr assay. Both DNA and anti-DNA antibody were enriched in the cryoprecipitates relative to the supernatant serum level. These data provide direct evidence for the presence of DNA/anti-DNA complexes in the circulation of patients with SLE.

The Medical Research Education Subcommittee of the American Rheumatism Association surveyed a random selection of large and small programs in internal medicine and family practice residency programs in order to evaluate their rheumatology training. Formal rheumatology training is offered in 90% of these residency programs, but many available positions are not being filled. A full-time staff rheumatologist was present at 69% of large internal medicine programs, 32% of small internal medicine programs, and 11% of family practice programs. The methods of rheumatology training are similar in most programs, although small internal medicine programs and family practice programs more often utilize physicians' offices or outside medical centers for the rheumatology elective training. A majority of the directors of these residency programs thought that many basic skills and techniques were not taught adequately and that the training of their rheumatology residents was not equal to that of residents in cardiology or gastroenterology.

Shedding of cell-surface antigens that react with anti-lymphocyte autoantibodies in systemic lupus erythematosus (SLE) is well-recognized, but the nature of such molecules is unknown. The present investigation demonstrates the rapid shedding of three IgG antibody target molecules of Mr 55,000, 37,000, and approximately 32,000 from the surface of mitogen-activated peripheral T cells during brief incubation at 37 degrees C. Sera lacking IgG anti-lymphocyte antibodies stained none of the three antigens. Absorption of antibody-positive sera with viable HSB-2 cells, a primitive T-cell line lacking HLA antigens and many CD antigens characteristic of mature peripheral T cells, eliminated staining of the shed molecules. These data delineate the number and estimated molecular mass of anti-lymphocyte autoantibody target molecules that are shed from the surface of T cells, and provide further insight into potential mechanisms by which anti-lymphocyte antibodies contribute to the pathogenesis of SLE and related disorders.

The clinical and serologic characteristics of 17 patients with onset of systemic lupus erythematosus (SLE) after age 50 were compared with those of 49 younger patients. All patients were followed prospectively for a mean duration of 47 months. A clinical and serological data base was established for each patient, and information collected at each visit to a lupus clinic was analyzed by computer. The clinical featured distinguishing old-age SLE were a low incidence of significant renal disease and prominent pleuropericarditis and arthritis throughout the period of followup. Serologic abnormalities were milder in older patients. Thus, hypocomplementemia, anti-DNA antibodies, and C1q precipitins occurred less frequently, and rheumatoid factor was more often present than in younger patients with SLE. Regression analysis suggested linear change in disease expression with age rather than distinct age-related subgroups.

Serum from patients with systemic lupus erythematosus (SLE) frequently contain IgM and IgG autoantibodies to the constitutively expressed 73-kD/pI 5.5 member of the hsp70 family of heat shock proteins, as determined by one-dimensional (SDS-PAGE) and two-dimensional (IEF/SDS-PAGE) immunoblotting, and by solid-phase SLE Ig immunoprecipitation experiments using hsp70 protein-specific mAbs as probes. Autoantibodies to hsp70 also were detected in a minority of sera from patients with other rheumatic or viral diseases, but not in normal sera. These data may provide additional insight into etiologic and pathophysiologic mechanisms in this and related autoimmune disorders.

Sera from majority of patients with seropositive rheumatoid arthritis, which generally lacked detectable anti-double stranded DNA in Farr, Crithidia luciliae, and microcomplement fixation assays, exhibited high levels of dsDNA binding in the presence of 3.5% polyethylene glycol when using intrinsically labeled 3H-PM2 DNA as antigen. Except for SLE, such increased dsDNA binding was absent in normal and a variety of other disease sera, including those from patients with seronegative rheumatoid arthritis. In contrast to the situation in SLE, in which dsDNA binding is mediated by specific anti-DNA antibody, the increased dsDNA binding activity in seropositive rheumatoid arthritis was shown to be dependent upon complex low avidity interactions involving DNA, IgG, IgM rheumatoid factor, and low density lipoproteins. Analysis of the composition of the polyethylene glycol serum precipitates by 2-dimensional gel diffusion, immunoelectrophoresis, and sodium dodecyl sulfate polyacrylamide gel electrophoresis failed to reveal the presence of additional DNA-binding proteins unique to seropositive rheumatoid arthritis. The only feature distinguishing high DNA binding sera from those with low DNA binding activity was an increased amount of polyethylene glycol-insoluble IgG in the former, presumably reflecting IgG/IgG and/or IgG/IgM complexes. The significance of these unusual DNA/low density lipoprotein/IgG/rheumatoid factor complexes with respect to the diagnostic specificity and pathophysiology of the DNA/anti-DNA system is discussed.

A large number of disease states are characterized by cryoglobulinemia. Quantification and immunochemical classification of cryoglobulins in serum provide information of diagnostic and pathophysiologic utility. Thus, type I cryoglobulins consist of a monoclonal immunoglobulin of a single class and are associated with lymphoproliferative disorders, such as multiple myeloma. Type II (mixed) cryoglobulins contain monoclonal IgM or rheumatoid factor and polyclonal IgG, and occur in patients with Waldenstrom's macroglobulinemia or chronic active hepatitis, for example. In type III cryoglobulins, both the IgM and rheumatoid factor and the IgG components are polyclonal. A large number of autoimmune or infectious diseases exhibit type III cryoglobulinemia. In certain well-studied situations, type II and type III cryoglobulins have been shown to contain antigen-antibody complexes directly involved in tissue injury in vivo, e.g., DNA and anti-DNA in systemic lupus erythematosus.

In addition to previously described cold-reactive IgM anti-lymphocyte antibodies maximally cytotoxic for resting cells at 15 degrees C, sera from patients with systemic lupus erythematosus (SLE) were found to contain a new type of antibody preferentially reactive at physiologic temperatures with mitogen-activated lymphocytes. This antibody lacked specificity for unstimulated lymphocytes, and was shown to be of the IgG class both by indirect immunofluorescence and in immunochemical experiments. Certain SLE sera also contained IgG antibodies with the capacity to develop plaques with mitogen-activated T lymphocyte preparations used in a reverse hemolytic plaque assay, indicating reactivity with products released by activated cells. The elimination of the ability of SLE sera to develop plaques after absorption with viable mitogen-stimulated lymphocytes, but not with resting cells, suggested that these antibodies were directed toward activation "neoantigen(s)" shed from the cell surface membrane. Surface membrane phenotype analyses performed by using a variety of monoclonal antibody reagents indicated that the plaque-forming cells (PFC) detected with SLE sera were activated T lymphocytes not restricted to single OKT4+, OKT8+, or Ia antigen+ subpopulations. Essentially all PFC expressed transferrin receptors. The present data raise the possibility that certain of the interesting effects of anti-lymphocyte antibodies on immunologic function in SLE may be mediated by interactions of these new type(s) of antibodies with activated lymphocytes or their products, rather than through blocking or depletion effects on resting precursor cells.

The subclass distribution of anti-Sm antibodies in the serum of MRL/Mp-Ipr/Ipr and MRL/Mp- +/+ mice was investigated with a sensitive ELISA technique. In both strains, but particularly in the Ipr mice, anti-Sm antibodies were predominantly of the IgG2a isotype. This preponderance was not an artifact of the sensitivity or specificity of the subclass-specific anti-Sm ELISA, nor did it reflect the subclass distribution of total serum IgG. A plaque-forming cell assay for anti-Sm antibodies also showed a predominance of the IgG2a isotype, indicating that the serum findings could not be explained by differential catabolism of IgG subclasses. Finally, antibodies to double-stranded DNA, as detected by the Crithidia luciliae assay, did not show a restricted subclass distribution. The isotype expression of anti-Sm antibodies in MRL mice must reflect in vivo mechanisms regulating the production of these autoantibodies. The IgG2a restriction in particular suggests an important in vivo role for T cells.

Human peripheral blood lymphocytes were digested briefly with protease prior to application of indirect immunofluorescence techniques for detecting alloantibodies in sera of patients with chronic renal failure on maintenance hemodialysis. Background staining of intrinsic surface IgM and cytophilic IgG bound to Fc receptors was eliminated or greatly reduced, enabling detection of B cell specific antibodies, including cold-reactive types not demonstrable by conventional immunofluorescence or complement-dependent lymphocytotoxicity. The antigenicity of HLA and other surface membrane determinants was not decreased by protease, although reactivity with certain sera was enhanced. In experiments comparing indirect immunofluorescence using protease-treated cells with complement-dependent lymphocytotoxicity and antibody-dependent, lymphocyte-mediated cytotoxicity assays, indirect immunofluorescence was more sensitive and comprehensive, but not less specific, in defining alloantibodies of a variety of types.

Monoclonal anti-beta 2-microglobulin (beta 2m) inhibited in a specific, dose-dependent fashion both in vitro tetanus toxoid-induced human-T-cell proliferation and sheep erythrocyte (E)-rosette formation, a function of the 50 kDa T11 molecule. In these respects, anti-beta 2m exhibited effects similar to those of sera from patients with SLE. Although 10 of 16 SLE sera contained antibody to beta 2m in monoclonal rosette inhibition assays, the presence of antibody of this specificity contributed only partially to the capacity of SLE serum to inhibit E-rosette formation or the T-cell response to tetanus toxoid. Removal of anti-beta 2m from SLE serum by solid phase absorption with beta 2m-Sepharose 4B reduced inhibition of the tetanus toxoid response and E-rosette formation in certain cases, but to a lesser extent than that observed following absorption with T-cell blasts, which completely eliminated inhibitory activity. Neither SLE antilymphocyte antibodies (including anti-beta 2m) nor heterologous anti-beta 2m were directed to the E receptor binding site (T11(1) epitope), as indicated by failure to inhibit OKT11 monoclonal antibody rosette formation or to reduce the relative intensity of OKT11 immunofluorescent staining. These data suggest an interesting functional relationship between beta 2m, the E receptor, and T-cell activation. While anti-beta 2m antibodies in SLE exert some inhibitory effect on antigen-induced T-cell proliferation, other distinct autoantibody systems to T-cell activation antigens appear to play the predominant role in this regard.

Three cell lymphocyte antigens of m.w. 55,000, 70,000, and 105,000 to 110,000 were identified by Western blotting as targets of IgM autoantibodies in serum from a group of 49 patients with systemic lupus erythematosus. The 55- and 70-kDa antigens were well expressed on unstimulated peripheral T cells, whereas the 105- to 110-kDa target was demonstrable only on mitogen-activated T cells and lymphoblastoid T cell lines. Localization of these molecules to the plasma membrane was established by cytoabsorption experiments in which IgM antibody staining of blotted antigens was specifically absorbed from systemic lupus erythematosus serum during 4 degrees C incubations with intact lymphocytes, and by their detection in purified lymphocyte plasma membranes. While the identity of these target antigens vis a vis known surface determinants was not defined, their expression on peripheral T cells from multiple donors and on cell lines of both undifferentiated (HSB-2) and phenotypically mature (Jurkat; HUT 78) types excluded alloantigens, major histocompatibility complex-encoded determinants, and most T cell differentiation antigens as candidates in this regard. Expression of the IgM autoantibody targets on HSB-2 cells argues against discrete T subset specificities as well. IgM reactivity with the 55-, 70-, and 105- to 110-kDa antigens by blotting was highly correlated with antilymphocyte antibody activity in complement-dependent cytotoxicity assays (Fisher's p less than 0.001), and paralleled flow microfluorimetric and microcytotoxicity quantitation of IgM antibody activity in serial observations of individual patients studied during different phases of disease activity. Taken together, these data suggest that IgM lymphocytotoxic antibodies in systemic lupus erythematosus are directed predominantly against a limited number of non-T cell subset-specific antigens.

IgG Fc receptors on human peripheral blood lymphocytes (PBL) were characterized by immunofluorescence studies with defined rabbit IgG b4 allotype/anti-allotype complexes. Three discrete types of Fc receptor-bearing cells, totaling approximately 33% of PBL, were identified. Fc receptors of the three types differed in their sensitivity to trypsin and in either absolute or localized density (topography) as determined by variable requirements for anti-IgC cross-linking in order to visualize bound complexes microscopically. The question of additional heterogeneity related to differences in individual Fc receptor affinity for complexed IgG was not approached in this study. Ten to 15% of PBL had pronase-sensitive, trypsin-resistant Fc receptors readily detected by direct immunofluorescence by using large fluorescein-conjugated complexes prepared near equivalence. Double label and lymphocyte fractionation experiments established this population to be largely distinct from suface IgM+ B cells and T cells, and identical to EA Ripley rosette-forming cells. Approximately 50% of surface IgM+ B cells and approximately 10% of T cells had lower density Fc receptors identified by indirect immunofluorescence with small complexes prepared in antigen excess or by cross-linking fluorescein-conjugated complexes with anti-rabbit IgG anti-serum. An additional approximately 15% peripheral T and B cells had very low density Fc receptors detectable by complexing the IgG on the cell surface by sequential incubations of cells with b4 IgG and anti-b4. Fc receptors on B and T cells were sensitive to both pronase and trypsin digestion. The heterogeneity of IgG Fc receptors on different lymphocyte subpopulations as defined by these these experiments may be of relevance for further analysis of normal and abnormal immune function.

Paired serum and cerebrospinal fluid specimens from 19 patients with SLE and central nervous system dysfunction were studied with respect to cerebrospinal fluid IgG index (a measure of intrathecal IgG synthesis), isoelectric focusing using immunoperoxidase staining techniques to detect oligoclonal IgG, and determination of the cerebrospinal fluid/serum albumin quotient (Q albumin) as a measure of blood-brain barrier integrity. Twenty-five patients without neurologic disease and 70 patients with a variety of non-SLE neurologic disorders were also studied for comparison. Of most interest was the observation that 42 percent of the patients with SLE had cerebrospinal fluid oligoclonal IgG, usually in association with elevation of the cerebrospinal fluid IgG index. In addition, two of the cerebrospinal fluid specimens that exhibited oligoclonal IgG also had increased titers of alpha-interferon. Q albumin was normal (under 9.0) in 12 of 13 patients with SLE, who had seizure, psychosis, or cranial neuropathy as principal central nervous system manifestations (mean +/- SD = 5.3 +/- 2.4), but was significantly elevated (mean +/- SD = 27.4 +/- 18.8, p less than 0.001) in five of six patients with diffuse, major central nervous system injury, for example, encephalopathy with coma, transverse myelopathy, paraparesis. Blood-brain barrier impairment was not correlated either with presence of circulating immune complexes or with other clinical or serologic evidence for extra-central nervous system disease activity. Taken together, the data suggest that, within the limitations of the techniques used, impairment of the blood-brain barrier in SLE may be secondary to the central nervous system lesion, rather than a result of systemic immune complex injury. In addition, substantial evidence is provided for an ongoing humoral immune response within the central nervous system in this disorder, which, in certain patients, may be associated with the production of intrathecal alpha-interferon.

Albumin and immunoglobulin G (IgG) were determined in cerebrospinal fluid (CSF) and serum, and the CSF/serum albumin Index (CSF X 10(3)/serum albumin concentration ratio) and IgG Index [(CSF/serum IgG)/(CSF/serum albumin)] were calculated. Data for these indices and oligoclonal banding are described in 23 cases of multiple sclerosis (MS), 19 of systemic lupus erythematosus (SLE), eight of sarcoidosis, 48 cases of miscellaneous disease, and 25 control patients with nonspecific complaints. Of the MS, SLE, and sarcoidosis patient groups, 8.5%, 26%, and 12.5% showed an abnormally high CSF/serum albumin Index; 87%, 16%, and 0% an increased IgG Index; and 87.5%, 42% and 0% showed positive oligoclonal banding. IgG Index and oligoclonal banding results for MS patients differed significantly from the sarcoidosis (p less than .001) and SLE (p less than .05) groups. When the CSF/serum albumin Index is considered also, the control and sarcoidosis patient results differ significantly from the MS group (p less than .001 and p less than .01). A strong correlation between the IgG Index and oligoclonal banding is implicated.

Circulating T cells bearing receptors for the Fc portion of IgG (T gamma) were identified by sensitive immunofluorescent techniques with rabbit IgG b4 allotype/anti-b4 complexes. A twofold decrease in both proportion and absolute number of T gamma cells was found in patients with active systemic lupus erythematosus (SLE) relative to values obtained during disease remission. The reduction in T gamma cells was most evident in patients with severe hypocomplementemia. A deficit of T gamma cells in active patients was not demonstrated. The percentage of total T cells rosetting with sheep erythrocytes was reduced in peripheral blood of most patients regardless of disease activity status, but particularly during SLE exacerbation. Cells lacking intrinsic surface immunoglobulin, IgG Fc receptors, and receptors for sheep erythrocytes were increased. These cells, operationally termed null, exhibited an inverse linear relationship with T cells that was not apparent in regression analyses performed against other lymphocyte subpopulations. Such differences were not found for B cells and IgG receptor-bearing non-B/non-T cells which were present in normal proportions in virtually all patients. The origin and functional significance of these unusual lymphocyte subpopulation abnormalities are discussed.

Sera from patients with active systemic lupus erythematosus caused the retention of the radioactively labeled synthetic polynucleotide 14C-dAT on cellulose nitrate filters. The amount of 14C-dAT bound correlated with the presence of hypocomplementemia and active renal disease. Sera containing only anti-single-stranded DNA antibodies did not bind 14C-dAT but did bind 125I DNA from E coli that had been prefiltered to remove ssDNA contamination. Inhibition experiments also indicated the presence of antigenic differences between 14C-dAT and prefiltered 125I-E coli DNA. The data suggest that dAT carries one of the major antigenic specificities for antinative DNA antibodies in SLE, and that cellulose nitrate filtration fails to remove a significant proportion of contaminating single-stranded DNA determinants in non-synthetic DNA.

The relative specificity of systemic lupus erythematosus antilymphocyte antibodies for T cell subsets bearing the OKT4, OKT8, or autologous erythrocyte rosette (A-RFC) markers was examined in complement-dependent microcytotoxicity assays and by indirect immunofluorescence. Target cells included normal OKT4+ or OKT8+ T cell clones established from mitogen-activated blasts, resting or phytohemagglutinin-activated peripheral T cells highly enriched for OKT4+ cells or OKT8+ cells, and A-RFC+ and A-RFC- populations. In the majority of sera, cytotoxicity for T4+ cells was greater than that for T8+ cells regardless of cellular activation status. Overall cytotoxicity was considerably higher for activated cells, however, especially when warm assay temperatures were used. A-RFC+ targets were more reactive than nonrosetting T cells, and this was associated with strikingly higher relative fluorescence intensity of IgM staining. Despite these consistent differences in relative cytotoxicity or staining, antibody titers against all cell types were strongly correlated in individual sera. Absorption experiments failed to demonstrate distinct antibody specificities for T4+ and A-RFC+ cells. These data suggest that the major determinant of cytotoxic reactivity may be a single or limited number of surface antigens common to all T cells. Superimposed on this dominant system(s) is a special reactivity with certain distinct subsets and with activated T cells generally.

In a prospective study 26 of 29 patients with systemic lupus erythematosus had cold-reactive antilymphocyte antibodies cytotoxic for autologous lymphocytes and lymphocytes from normal subjects. The level of antilymphocyte antibodies was highly correlated, by linear regression analysis, with lymphopenia in these patients. The data suggested that both the avidity and the concentration of these antibodies were important determinants in this relationship. A clear association between increased antilymphocyte antibody activity and exacerbation of SLE was demonstrated. Apart from lymphopenia, however, neither type of clinical manifestation nor any particular serologic abnormality appeared to be related to the presence of antilymphocyte antibodies.

Unmeasurable total haemolytic complement (C) was observed in serum of a patient with untreated chronic lymphocytic leukaemia and recurrent non-hereditary angioedema. Analysis of C components immunochemically demonstrated a marked reduction of C1q and C1s inhibitor, undetectable C1r, C1s and an elevated B. Haemolytic C1, C4 and C2 were less than 5 percent of normal, functional C1s inhibitor was absent. Cryoglobulin and C1q precipitins were present in the serum. Of special interest was the presence of high levels of cold-reactive antilymphocyte antibody, determined by both C-dependent cytotoxicity and indirect immunofluorescence. The antibody exhibited specificities for both autologous lymphocytes and lymphocytes from normal donors; cytotoxic activity for autologous leukaemia cells was removed by absorption with normal isologous tonsil lymphocytes. Specific enrichment of this antibody relative to the serum level was demonstrated in the cryoglobulin and its isolated 19S fractions. Free lymphocyte surface antigen was also demonstrated by gel diffusion using specific rabbit antilymphocyte antiserum. These data strongly suggest the presence of pathogenetically significant circulating complexes of lymphocyte surface antigen and specific antibody in certain patients with CLL.

Using the direct migration inhibition test, response to measles antigen in patients with systemic lupus erythematosus (SLE) was found to be decreased when compared with that of normal subjects. No alteration was observed in similar experiments using parainfluenza type 1 and rubella antigens. The specific decrease in measles antigen effect showed no obvious correlation with activity of SLE or with the presence of lymphocytotoxic antibodies. Whether the specificity of the decrease in reactivity is due to some particular relationship between the measles virus or antigen and SLE, or to the possibility that measles reactivity is a more sensitive indicator of a generalized defect of cell-mediated immunity, remains unclear.