Induction of antibodies to nuclear antigens in rabbits by immunization with hydralazine-human serum albumin conjugates.

The antihypertensive drug hydralazine can induce in man a syndrome similar to spontaneous systemic lupus erythematosus (SLE). The pathogenesis of this drug-induced syndrome is not understood. In this investigation, five groups of rabbits were studied: group I, 10 rabbits hyperimmunized with hydralazine conjugated to human serum albumin (HSA) in complete Freund's adjuvant (CFA); group II, four rabbits with HSA in CFA; group III, four rabbits with CFA alone; group IV, five rabbits with hydralazine conjugated to rabbit serum albumin (RSA); and group V, four rabbits with a major metabolite of hydralazine conjugated to HSA. The rabbits immunized with hydralazine-HSA developed rising titers of antibodies to hydralazine and progressively increasing amounts of antibodies to both single-stranded and native DNA. The antibodies to DNA were cross-reactive with hydralazine as determined by inhibition of DNA binding and DNA hemagglutination tests. Similar results were obtained in rabbits immunized with the metabolite-HSA compound except the major hapten antibody response was to the metabolite. The DNA antibodies in this group were also capable of being absorbed by metabolite-HSA as well as hydralazine-HSA, indicative of the cross-reactivity between hydralazine and its metabolite. Immunization with hydralazine-RSA caused rabbits to produce antibodies to hydralazine but not to DNA, indicating the requirement for an immune response to the carrier protein in order for antibodies reactive with DNA to be produced. Thus, hyperimmunization of rabbits with hydralazine-protein conjugates may provide a useful animal model of SLE. The data suggests that an immune response to hydralazine may be important in human hydralazine-induced SLE.

Cellular immunity to nuclear antigens in systemic lupus erythematosus.

Cellular immune repsonses were determined by skin testing and mitogen- and antigen-induced blastic transformation of peripheral blood lymphocyte cultures in 24 patients with systemic lupus erythematosus (SLE) and 24 normal subjects. The incidence of positive skin tests with Candida albicans, PPD (tuberculin-purified protein derivative) intermediate strength, Trichophyton and histoplasmin was not significantly different in the two groups nor was lymphocyte stimulation by the mitogen phytohemagglutinin-M (PHA-M), implying that cellular immunity is normal in SLE. However, the SLE patients had a significantly increased incidence of positive skin tests and stimulated lymphocyte cultures to a number of nuclear antigens compared with normal subjects. No correlation could be made between the test results and the activity of the SLE at the time of study except for a significant association between lymphocyte culture stimulation by rabbit thymus native DNA and active SLE nephritis. Patients with a membranous antinuclear factor (ANF) pattern had positive skin tests with rabbit thymus native DNA and usually had active disease.

Quantitative studies of human monokine-induced endothelial cell elongation.

Leucocyte-endothelial cell interactions are important in the inflammatory response. In this study, the effect of peripheral blood mononuclear cell (PBMC) products on endothelial cell (EC) shape was examined and quantified. PBMC were obtained from normal donors by Ficoll-Hypaque separation of heparinized whole blood and cultured for 72 hr in media containing 10% fetal calf serum with and without concanavalin A (Con A). Media conditioned by PBMC or control, nonconditioned media were then added to preconfluent, first passage EC cultures derived from human umbilical veins. Conditioned media from Con A-stimulated PBMC resulted in a dose-dependent, marked elongation and whorling of cultured EC. The minimum effective concentration found to elicit a response was 1.25%, with a maximum response occurring at 10%. Quantitative morphometric analyses of treated EC indicated that the elongation was highly significant (p less than 0.001) when compared to EC incubated with control, nonconditioned media. In addition, EC elongation was accompanied by a highly significant (p less than 0.001) increase in cell area. Although less dramatic, conditioned media from unstimulated PBMC also elicited a similar, significant dose-dependent change in EC shape. Significant changes in EC shape were evident within 6 hr and continued over the time course of the experiment (40 h). Cell shape changes were partially reversible at 18 h after removal of the PBMC-conditioned media and replacement with control, nonconditioned media. The change in EC morphology induced by a PBMC-derived factor(s) suggests a mechanism by which activated leucocytes may modulate cellular traffic at the blood-vessel wall interface.

Release of estradiol from fetal bovine serum by rat thymus, spleen, kidney, lung and lung macrophage cultures.

High levels of estradiol (E2) were detected by radioimmunoassay (RIA) in supernatants of rat thymus and spleen, kidney, lung and lung macrophages cultured in the presence of fetal bovine serum(FBS). A competitive inhibitor of E2 synthesis and no effect on the production of E2 by thymic cultures. Acid hydrolysis of estrogen conjugates in FBS released enough free E2 to account for the high level found in culture supernatants. Removal of free estrogens and estrogen conjugates by charcoal stripping the serum before addition to media of rat tissues by hydrolysis of estrogen conjugates in FBS.

Prospective study of immunologic effects of hydralazine in hypertensive patients.

Twenty-seven hypertensive patients (23 of whom were black) were treated with hydralazine as their major antihypertensive drug and were followed for evidence of autoimmunity and clinical systemic lupus erythematosus (SLE). Only one patient developed SLE but many, although asymptomatic, had serologic evidence of autoimmunity: antibodies to single- and double-stranded ribonucleic acid (RNA), single-stranded deoxyribonucleic acid (DNA), histones, and lymphocytes. Acetylation phenotype profoundly influenced this response; slow acetylators had a higher incidence and larger amounts of autoantibodies. Antibodies to both types of RNA were a more sensitive index of autoimmunity than antinuclear antibodies. Hydralazine treatment did not alter cell-mediated immune responses. The hydralazine SLE patient had large amounts of autoantibodies that were predominantly IgG, while in the others IgM autoantibodies were predominant. No antibodies, but positive lymphoproliferative responses to hydralazine, were found in half the patients tested.

Nonsteroidal anti-inflammatory drugs and modulation of cartilaginous changes in osteoarthritis and rheumatoid arthritis. Clinical implications.

Nonsteroidal anti-inflammatory drugs have a potential for modifying the complex pathophysiologic events leading to cartilage destruction in various forms of arthritis. Following an evaluation of basic mechanisms in the pathogenesis of cartilaginous destructive lesions, the effects of nonsteroidal anti-inflammatory drugs on normal chondrocyte metabolism are discussed. Their capacity to modulate cartilage and bone lesions in experimental forms of arthritis is addressed, as is the manner in which they may modify the pathophysiology of cartilage destruction in human forms of arthritis. Different classes of nonsteroidal anti-inflammatory drugs produce different effects in certain in vivo or in vitro settings.

Cartilage metabolism and anti-inflammatory drugs in osteoarthritis.

The issue of selecting nonsteroidal anti-inflammatory drugs (NSAIDs) for the management of osteoarthritis based upon their direct effect on normal cartilage metabolism and their ability to modulate mechanisms that disturb such metabolism is addressed. This article provides an overview of the anatomic and biochemical changes that occur in diseased synovial tissue and cartilage, and the pathophysiologic mechanisms presumed to function in the induction and perpetuation of cartilage failure. Studies using experimental animal models of osteoarthritis are reviewed to evaluate the natural history of the disease, as well as the direct effect of NSAIDs on anabolic and catabolic cartilage function. Experimental evidence strongly indicates that NSAIDs of different classes, as well as members within a given class, function differently and manifest activity in addition to cyclo-oxygenase inhibition.

Prospective study of the evolution of Raynaud's phenomenon.

Seventy-four patients with Raynaud's phenomenon and no associated illness were followed prospectively to determine whether a secondary disease would develop, and clinical and laboratory assessments were performed at study entry to determine their association with the subsequent development of disease. After an average of 2.7 years of follow-up (range 0.5 to 5.7 years), outcome information was available on 58 persons (78.4 percent). A connective tissue disease developed in 11 (19.0 percent): three systemic sclerosis and eight CREST syndrome. The two variables at entry most strongly associated with the subsequent development of a connective tissue disease were an abnormal nailfold capillary pattern (adjusted odds ratio = 26.82, 95 percent confidence interval = 4.69, 153.2) and an abnormal pulmonary function test result (odds ratio = 4.78, 95 percent confidence interval = 1.02, 22.41). The positive association of an abnormal barium esophagram, presence of antinuclear antibodies, and cutaneous abnormalities did not reach statistical significance. The development of connective tissue diseases in this group of patients is not rare. An abnormal nailfold capillary pattern is strongly associated with the subsequent development of systemic sclerosis or CREST syndrome in patients with Raynaud's phenomenon.

3-Hydroxymethyl-s-triazolo[3,4-a]phthalazine, a novel urinary hydralazine metabolite in man.

The elucidation of the structure of a new major metabolic product of hydralazine, 3-hydroxymethyl-s-triazolo[3,4-a]-phthalazine, is described. The structures of several other previously described metabolites of the drug, phthalazone, s-triazolo[3,4-a]phthalazine, and 3-methyl-s-triazolo[3,4-a]phthalazine, are confirmed. A metabolic pathway of hydralazine is also proposed.

Genetic and immunologic studies of patients on procainamide.

Forty (40) patients with cardiac arrhythmias receiving procainamide (PA) therapy and 24 patients who were receiving other drugs for their cardiac disorders were investigated for class II HLA phenotypes and their DRB1*04 and DQB1*03 subtypes. Other genetic marker evaluations in the PA patients included: 1) class III MHC C4A and C4B null alleles of complement; and, 2) acetylation phenotype. Twenty (20) of the PA patients were also tested for the ability of their stimulated cells to secrete Interleukin-1 (IL-1 beta) and tumor necrosis factor (TNF alpha). We also examined the spontaneous production of these cytokines by peripheral blood leukocytes (PBL) from patients who were receiving chronic PA treatment. The results revealed no association of acetylation phenotypes with the class II HLA phenotypes nor class III MHC C4 allotypes in these patients. The results did show a significant increase in class III C4 complement allotypes in the PA patients when compared to the controls. The results also showed a significant increase in autoantibodies and DQw3 phenotypes in the PA patient group when compared to control populations. Results of spontaneous IL-1 and TNF production suggested there may be an association of select class II HLA phenotypes in some patients and this may be relevant to host responsiveness to PA treatment.

Are there environmental forms of systemic autoimmune diseases?

A large number of drugs and an increasing number of environmental agents reportedly result in the appearance of a number of autoantibodies and in many instances in the appearance of a range of autoimmune clinical syndromes. The major disorders so recognized have marked resemblances to the autoimmune disease systemic lupus erythematosus. The commonly used term is drug-induced lupus; a better term is drug-related lupus. There is considerable interest at the present time in an increasing number of environmental agents. There have been two epidemics in recent years--one in Spain to a contaminant of rapeseed oil and one in the United States to a contaminant of l-tryptophan that caused an eosinophilic myositis. It is important for physicians and others involved in health care to recognize the potential associations of these diseases of unknown cause or causes.

Erosive osteoarthritis.

Erosive osteoarthritis is a disorder that most often involves the hands of postmenopausal women. It can begin abruptly with pain, swelling, and tenderness. Distal interphalangeal joints are involved most frequently, followed by proximal interphalangeal joints. Occasionally there is metacarpophalangeal, carpal, or large joint involvement. The female-to-male ratio is approximately 12:1. There are no known HLA associations. Laboratory studies generally are negative. A mild elevation of the sedimentation rate may occur. Radiologically, the disorder is characterized by central erosions and the "gull wing" deformity. Synovial pathology has shown changes consistent with both rheumatoid arthritis and osteoarthritis and manifests the stage of disease at the time of biopsy. The etiology remains obscure, but hormonal influences, metabolic disorders, and autoimmunity have been implicated. Treatment is largely supportive with physical therapy, nonsteroidal antiinflammatory drugs, and occasionally prednisone. Overall prognosis is good, although deformity and impairment of hand function may occur. For this reason, a reassessment of treatment strategies may be in order.

In vitro studies in reproductive immunology. - 2. Demonstration of the inhibitory effect of male genital tract constituents on PHA-stimulated mitogenesis and E-rosette formation of human lymphocytes.

The effect of male genital tract components (human spermatozoa, intact and chromatographed seminal plasma fractions) on in vitro cell-mediated immune reactions was examined. Their addition to PHA-containing lymphocyte cultures resulted in a marked degree of inhibition of DNA synthesis, the response varying with the fractions employed. The interference by such components with a terminal cell event during blast transformation was suggested by the failure of stimulated cultures to incorporate thymidine irrespective of the time during the incubation period at which the constituents were added. Seminal plasma and sperm extracts were also shown to inhibit T-cell associated E-rosette formation. The inhibitory properties of genital fractions remained unaltered after repetitive freeze-thaw cycling when tested in the transformation and E-rosette assays. Heating to 100 degrees C modified the response with some of the fractions.

An evaluation of commercial kits for the detection of antibodies to double-stranded DNA.

Antibodies to double-stranded (ds) DNA may provide useful information in the management of patients with systemic lupus erythematosus. commercial radioimmunoassay (RIA) kits for the detection of (ds) DNA antibodies (Lupo-Tec, Wampole Laboratories; anti-DNA kit, Amersham Corporation) were compared with a modified Farr assay and checked for intra-lot, inter-lot and inter-assay variation. Purity of DNA preparations was assessed using rabbit antibody to single-stranded (ss) DNA. Selected sera with low (less than or equal to 15%), moderately elevated (less than or equal to 40%), or markedly elevated (greater than or equal to 44%) (ds) DNA binding values (Farr assay) were tested. Positive and negative control sera when supplied with the kits also were evaluated. Normal sera were used as internal controls. A variable degree of intra-lot, inter-lot, and inter-assay correlation was observed. At low antibody levels, discordance was observed but values greater than or equal to 25% (Farr assay) were abnormally elevated in all kits tested. Clinical and laboratory personnel should be aware of potential pitfalls in RIA methods and carefully interpret results when commercial DNA kits are used.

Induction of autoantibodies by human immunodeficiency virus infection and their significance.

Although many of the changes in HIV and AIDS appear to be global, on closer investigation the defects are specific and can be explained by recognized mechanisms. Despite a demonstrated concurrent polyclonal B-cell activation, studies fail to show any evidence to date of significant dysregulation of the immune system leading to serologic or clinical autoimmunity.

Drug-related lupus. Incidence, mechanisms and clinical implications.

Adverse side effects to drugs and chemicals in which immune mechanisms may be responsible have been described in drug-related lupus (DRL). The spectrum of drugs that may elicit DRL includes such classes as the hydrazines, arylamines, and chemicals that can be metabolised to amines. The 2 major pathways of metabolism--acetylation and N-hydroxylation--are described in detail. The events leading to autoantibody production are not well understood; however, specific consideration of the genetic makeup of patients who are candidates for treatment with these drugs may help identify those at risk of developing DRL.

Advances in rheumatology.

This article reviews the major contributions to our understanding of the rheumatic diseases. These include the first definite evidence of linkage between genes controlling the immune response and the major histocompatibility complex genotype; the recognition of the types and the role of the many crystals in arthritis; the application of sophisticated immunologic techniques to the understanding of the antinuclear autoantibodies; the recognition of a new disorder--Lyme arthritis; and the contributions of joint replacement and other surgeries, and the numerous drugs and procedures now available for treatment of patients.

Felty's syndrome in black Americans: a case report.

This is a case report of a black woman with Felty's syndrome, who had recurrent, severe leg ulcers refractive to the usual treatment. The leg ulcers healed promptly after splenectomy and there has not been a recurrence of these lesions after three years' follow-up. The rarity of Felty's syndrome in blacks is highlighted and the literature on this aspect is reviewed. Information on seven patients with Felty's syndrome seen at the University of Cincinnati Medical Center is provided and the relationship of skin ulcers to splenectomy reviewed.

Cytokine modulation of chondrocyte metabolism--in vivo and in vitro effects of piroxicam.

Changes in the macromolecular proteolycan (PG) and collagen of the cartilage matrix may culminate in irreparable tissue destruction. Molecular modifications appear to result from: (A) exogenous proteinases, (B) endogenous chondrocyte proteinases whose synthesis and release is modulated by exogenous non-enzymatic cytokines (CKs) and (C) quantitative and/or qualitative alterations in chondrocyte PG and collagen synthesis which are potentially induced by exogenous CKs. Studies have recently been initiated to determine the effect of piroxicam on the synthesis and activity of such metabolic regulatory CKs in patients with rheumatoid arthritis and osteoarthritis, and in age-, sex-, and race-matched controls. Therapeutic doses of piroxicam alone had no effect on the anabolic or catabolic function of chondrocytes. Current studies concern the effect of piroxicam on: (a) spontaneous and lectin-driven production by peripheral blood monocytes and T-cells of trypsin-sensitive, heat-labile CKs (interleukin 1, lymphokine) which, in a protein- and RNA-synthesis-dependent manner, induce a concentration and duration of substrate exposure dependent release of chondrocyte PG- and collagen- degrading neutral proteinases in cartilage organ and chondrocyte suspension culture systems; (b) spontaneous and lectin-driven synthesis by peripheral blood T-cells of lymphokines capable of suppressing chondrocyte PG, glycosaminoglycan, protein, collagen and nucleic acid synthesis in a quantitatively reversible manner; (c) pathological synovial membrane synthesis of such catabolic-inducing and anabolic-modulatory CKs. These experimental model system are reviewed together with preliminary data on the effect of piroxicam.

Alterations in human endothelial cell morphology, proliferation and function by a macrophage-derived factor.

Changes in endothelial cell (EC) morphology occur at sites of physiological lymphocyte traffic and in areas of chronic inflammation. Previous studies have shown that EC shape changes also occur in vitro following exposure of EC monolayers to peripheral blood mononuclear cell (PBMC)-derived conditioned media (CM). In the present study, quantitative image analysis is used to define the cell of origin of the elongating factor(s), to examine changes in EC proliferation and function accompanying PBMC-induced human EC elongation and to identify the active PBMC-derived products responsible for this elongation. By separating mononuclear cells into subpopulations (macrophages, B cells and T cells) and adding conditioned media derived from these subpopulations to cultured ECs, the macrophage (M phi) is shown to be the primary cell of origin of the elongating factor(s). Furthermore, EC elongation is accompanied by both a dose-dependent decrease in cellular proliferation and an increase in prostacyclin production. These findings suggest that PBMC-induced changes in EC morphology may be associated with a shift from a proliferative state to a more secretory phase of the EC cycle. Finally, using recombinant factors it is shown that TNF alpha acting in combination with IL-1 may be the active PBMC-derived products which contribute to EC elongation.