Anti-interleukin-2 receptor monoclonal antibody therapy supports a role for Th1-like cells in HgCl2-induced autoimmunity in rats.

Brown-Norway (BN) rats injected with HgCl2 develop an autoimmune disease characterized by a T-dependent polyclonal B-cell activation. Increase in major histocompatibility complex class II molecule expression on B cells concomitant with enhancement of serum IgE concentration supports the involvement of the T helper 2 (Th2)-like subset in the induction of the disease. The mercury disease is autoregulated and does not develop in Lewis (LEW) rats. Considering the reciprocal regulation, well defined in mice, between the Th1 and Th2 subsets, we addressed the role of the Th1-like subset in this disease. Brown-Norway and LEW rats injected with HgCl2 were treated with NDS61, a mouse anti-rat-IL-2R MoAb that blocks mainly Th1 cells. Data reported herein show that: (1) HgCl2 treatment does not modify either the percentage of IL-2R+ cells or IL-2R expression in both BN and LEW rats; (2) treatment of BN rats with NDS61 MoAb does not modify the induction phase of the mercury disease but delays in part the regulation phase; (3) such a treatment leads to some immune abnormalities in LEW rats; (4) HgCl2 markedly potentiates the anti-mouse Ig antibody response in BN rats which probably limits the effect of this treatment. This study supports a role for the Th1-like subset in HgCl2-induced autoimmunity in the rat.

Evidence for a role of antilaminin-producing B cell clones that escape tolerance in the pathogenesis of HgCl2-induced membranous glomerulopathy.

In Brown-Norway rats HgCl2 induces an autoimmune disease due to a T-dependent B cell polyclonal activation. This disease is marked by the production of numerous antibodies including antiglomerular basement membrane (GBM) antibodies. Rats exhibit a biphasic glomerulopathy with heavy proteinuria. Initially anti-GBM antibodies are found linearly deposited; they precede the appearance of membranous glomerulopathy. Rats recover spontaneously even if HgCl2 injections are pursued, but mechanisms at play are unclear. We have assessed the effects of transplanting the spleen from a BN rat, either at the acme of the disease or at the time of convalescence, into naive BN rats, some of which were then injected with HgCl2. Transplantation of a spleen from HgCl2-injected rats at the acme of the disease dramatically protects BN rats from all the manifestations of the mercury disease. BN rats transplanted with a spleen from HgCl2-injected rats at the time of convalescence only exhibited a typical membranous glomerulopathy with heavy proteinuria but without circulating anti-GBM antibodies. Antilaminin antibodies were eluted from the glomeruli. This study shows that spleen cells from HgCl2-injected rats are able to confer tolerance to HgCl2-induced autoimmunity. It also shows that some B cell clones escape this tolerance. Finally, this study strongly suggests that membranous glomerulopathy, responsible for proteinuria in this model, is related to the presence of antilaminin antibodies.

Effect of the thiol group on experimental gold-induced autoimmunity.

Brown Norway rats injected with aurothiopropanolsulfonate sodium salt develop systemic autoimmunity. The aim of this study was to assess the influence of the sulfur-containing group in this experimental model of gold-induced autoimmunity. It was shown that the sulfur-containing group does not induce autoimmunity of itself, but potentiates the immunotoxic effects of gold.

Increased expression of class II major histocompatibility complex molecules on B cells in rats susceptible or resistant to HgCl2-induced autoimmunity.

Administration of HgCl2 to the susceptible Brown-Norway (BN) rats induces an autoimmune disease characterized by a T-dependent polyclonal activation of B cells responsible for a dramatic increase in serum IgE concentration. The resistant Lewis (LEW) rats injected with HgCl2 do not exhibit such autoimmune manifestations. We show here that, upon HgCl2 injections, major histocompatibility complex (MHC) class II molecule expression is increased very early in lymph nodes and spleen B cells from both strains. So far, it is the earliest marker (day 3) of the effect of HgCl2 on the immune system. In both strains this enhancement is transient, but regulatory mechanisms are much more efficient in the resistant LEW strain than in the susceptible BN strain. In addition, we observed that MHC class II molecule expression on B cells differs according to the organ and the rat strain tested. All these findings are discussed in an attempt to underline the role of MHC class II molecule expression in the occurrence of mercury-induced autoimmunity.

Prevention of experimental autoimmune uveoretinitis and experimental autoimmune pinealitis in (Lewis x Brown-Norway) F1 rats by HgCl2 injections.

Mercuric chloride (HgCl2) induces in Brown-Norway (BN) and (Lewis x Brown-Norway) F1 hybrid rats a transient autoimmune disease characterized by the production of various antibodies to self and non-self antigens and by a dramatic increase of serum IgE. Experimental autoimmune uveoretinitis (EAU) can be induced in Lewis (LEW) and (LEW x BN) F1 hybrid rats by a single immunization with retinal S-antigen (S-Ag). Besides uveoretinitis, animals immunized with S-Ag develop an autoimmune pinealitis (EAP). We demonstrate in this study that (LEW x BN) F1 hybrid rats, injected with HgCl2 7 days before S-Ag immunization, are quite efficiently protected against EAU and EAP. We also show that HgCl2-induced protection is neither due to a cytotoxic effect of HgCl2 nor to CD8+ T-cell dependent mechanisms nor to the HgCl2-induced increase of serum IgE concentration. The role of other hypothetical mechanisms, such as anti-S-Ag anti-idiotypic antibodies and/or HgCl2-induced unbalance between T-helper cell subsets, is discussed.

Beneficial effect of human therapeutic intravenous immunoglobulins (IVIg) in mercuric-chloride-induced autoimmune disease of Brown-Norway rats.

Administration of HgCl2 to the susceptible Brown-Norway (BN) strain of rats induces an autoimmune disease characterized by polyclonal B cell activation, increased serum levels of IgE and the occurrence of anti-glomerular basement membrane antibody-mediated glomerulonephritis. We have observed that the simultaneous administration to BN rats of normal human polyspecific immunoglobulins for therapeutic use (IVIg) with HgCl2 significantly decreased the occurrence and severity of proteinuria, and reduced serum IgE levels in diseased animals. Hypergammaglobulinaemia was potentiated in animals receiving HgCl2 and IVIg, compared with animals receiving HgCl2 alone. In vitro experiments indicated that F(ab')2 fragments from IVIg inhibited the binding to laminin of pathogenic anti-laminin antibodies from diseased rats, as did antibodies from the resistant Lewis strain of rats but not antibodies from susceptible BN rats. These observations suggest that IVIg may interfere with the immune regulatory mechanisms involved in mercury-induced autoimmune disease in an analogous fashion to the ability of IVIg to suppress the expression of certain pathological autoimmune responses in humans.

Graft-versus-host reactions in the rat mimic toxin-induced autoimmunity.

Gold salts, D-penicillamine or mercurials induce autoimmunity in Brown Norway (BN) rats and provoke an immunosuppression in Lewis (LEW) rats. It has been suggested that immunologically mediated manifestations induced by drugs could result from graft-versus-host (GVH) like reactions. We show that BN spleen cells transferred into (LEW x BN)F1 hybrids induce a chronic GVH reaction (GVHR). This reaction led to an autoimmune disease quite similar to that induced by drugs in BN rats. In both situations, a common part of the B cell repertoire is triggered. In contrast, LEW spleen cells transferred into (LEW x BN)F1 hybrids provoke a lethal GVHR. This is to be compared with the CD8-mediated immunosuppression observed in LEW rats injected with HgCl2. These findings are in agreement with the prediction that immune dysregulation induced by drugs leads to GVH-like reactions either stimulatory or suppressive depending upon the strain tested.

D-penicillamine-induced autoimmunity in Brown-Norway rats. Similarities with HgCl2-induced autoimmunity.

D-penicillamine (DP) has been previously shown to induce an autoimmune disease in Brown-Norway (BN) rats, characterized by a dermatitis, by the production of antinuclear antibodies, by the formation fo circulating immune complexes, and by linear IgG deposits along the glomerular basement membrane. These manifestations are quite similar to those observed in mercuric chloride (HgCl2)-induced autoimmunity. The mechanism of the latter disease has been recently partly elucidated. The aim of this study was to compare DP and HgCl2-induced autoimmunity in BN rats and to compare the mechanisms involved in both situations. A transient increase in the number of spleen cells, affecting B cells and CD4+ T cells, and an increase in serum IgE concentration, previously reported in HgCl2-induced autoimmunity, were observed during DP treatment. Autoreactive anti-class II T cells able to proliferate not only in the presence of autologous B cells but also in the presence of syngeneic normal B cells were found in DP-treated BN rats. Spontaneous regulation occurred, associated with the disappearance of autoreactive T cells. Suppressor CD8+ T cells were not involved in this phenomenon. Mechanisms involved in both the induction and the regulation of DP-induced autoimmunity seem to be quite similar to those reported in HgCl2-induced autoimmunity.

Inhibition of experimental autoimmune uveoretinitis by mercuric chloride injections in (Lewis x Brown Norway) F1 hybrid rats.

Experimental autoimmune uveoretinitis (EAU), induced in (LEW X BN) F1 rats by immunization with S antigen (S-Ag) is T cell and antibody (ab) mediated and anti-S-Ag IgE ab have been involved in the occurrence of ocular lesions. (LEW X BN) F1 rats repeatedly injected with HgCl2 develop an autoimmune disease characterized by numerous auto-ab and a high increase of serum IgE level. We hypothesize that large amounts of non anti-S-Ag IgE induced by HgCl2 would compete with anti-S-Ag induced by S-Ag immunization so as to prevent EAU to occur. Indeed (LEW X BN) F1 rats immunized with S-Ag 7 days after the first HgCl2 injection are strongly protected against EAU. The putative role of the different mercury-induced autoimmune phenomena in the protection against EAU are discussed.

Role of CD8+ T cells in mercury-induced autoimmunity or immunosuppression in the rat.

In Brown-Norway (BN) rats mercuric chloride induces an autoimmune disease characterized by an increase in serum IgE concentration, and by the production of anti-glomerular basement membrane antibodies responsible for a glomerulonephritis with a heavy proteinuria. (i) This disease results from a B-cell polyclonal activation probably due to frequent anti-class II T cells. (ii) The self limitation observed in this model is associated with both a decrease in the frequency of anti-class II T cells and the emergence of CD8+ T cells able to suppress these autoreactive T cells. (iii) In Lewis (LEW) rats which do not develop autoimmunity, HgCl2 provokes the appearance of non-antigen-specific CD8+ T cells responsible for a depression of T-cell functions. The aim of this work was to test the effect of treatment with an anti-CD8 monoclonal antibody (MoAb) in both BN and LEW rats. Anti-CD8 MoAb-treated rats were effectively depleted in CD8+ T cells. However, neither the induction nor regulation phases of mercury-induced autoimmunity were modified in BN rats. Mercury-induced immunosuppression in LEW rats was abrogated; however, depletion in CD8+ T cells did not allow the disease to occur in that strain. Finally, CD8 depletion induced in normal BN rats the appearance of rare anti-class II T cells showing that these cells are normally present in that strain but negatively controlled by suppressor T cells.

HgC12 induces T and B cells to proliferate and differentiate in BN rats.

Mercuric chloride induces in Brown-Norway (BN) rats an autoimmune disease characterized by the production of various autoantibodies and by a marked increase in the IgE serum concentration. This agent is responsible for a T dependent polyclonal activation of B cells, which is probably due to the emergence of autoreactive T cells. The aim of this study was to evaluate the effect of HgCl2 injections on lymphoid organs and on the serum concentration of the various Ig isotypes. HgCl2 induced (1) a lymphoproliferation in spleen and lymph nodes involving B and T helper cells while the number of T suppressor/cytotoxic cells was not modified, (2) an increase in the number of Ig containing cells resulting in a rise in all serum Ig isotypes, and (3) an early thymic atrophy probably immunologically mediated, which was not involved in the induction phase of the disease since adult thymectomy had no effect. These findings demonstrate that the polyclonal effect of HgCl2 is not isotype-restricted although the IgE response is predominantly affected and they support evidence for a major role for an excess of T help in the HgCl2-induced polyclonal activation of B cells. It was also observed that B cell areas are present in normal BN rat thymuses, the potential role of which in the induction of autoimmunity remains to be investigated.

Effect of HgCl2 on experimental allergic encephalomyelitis in Lewis rats. HgCl2-induced down-modulation of the disease.

HgCl2 induces autoimmunity in Brown-Norway rats and immunosuppression in Lewis rats. In the latter rats, HgCl2 triggers the proliferation of T suppressor/cytotoxic (OX8+) cells which actively suppress T cell functions. This led us to study the effect of HgCl2 on experimental allergic encephalomyelitis (EAE), a T cell-mediated autoimmune disease obtained following immunization with basic protein (BP). It will be shown that HgCl2 attenuates or even prevents clinical manifestations of EAE and inhibits both the proliferative response of T cells to BP and the anti-BP antibody response. This immunosuppression was not due to a defect at the T helper cell or antigen-processing cell level but to the emergence of T suppressor cells.

Autoreactive T cells in mercury-induced autoimmunity. Ability to induce the autoimmune disease.

It has been previously shown that autoreactive T cells appear during mercury-induced autoimmunity in Brown-Norway (BN) rats. In the present work, it is shown that: 1) T cells and T helper cells from HgCl2-injected BN rats are able to actively transfer autoimmunity in normal BN rats; the disease transferred is exacerbated when recipients are treated with the antisuppressor/cytotoxic T cell monoclonal antibody (OX8); 2) normal T cells preincubated with HgCl2 are also able to transfer the disease in OX8-treated but not in T cell-depleted rats; and 3) T cells from HgCl2-injected BN rats also transferred the disease in both normal and T cell depleted rats. It is concluded that: 1) autoreactive T cells, and presumably anti-Ia T cells are involved in the pathogenesis of mercury-induced autoimmunity; 2) these autoreactive T cells induce suppressor/cytotoxic T cells to proliferate in normal syngeneic recipients; the fact that this T cell subset did not proliferate in HgCl2-injected BN rats suggests that HgCl2 also affects T suppressor cells; and 3) mercury-induced autoimmunity could result from the additive effect of the emergence of autoreactive T cells and of a defect at the T suppressor level.

Relationship between the liver and lymphocytotoxic alloantibodies in inbred rats. Specific absorption by nonparenchymal liver cells.

Liver allografts have a privileged status with regard to hyperacute rejection. In this experimental study, we have used extracorporeal liver hemoperfusion in sensitized rats in order to analyze reactions between lymphocytotoxic antibodies and the liver. In sensitized BN rats, a donor-specific (Lewis) extracorporeal liver hemoperfusion can delay hyperacute rejection of heart allografts and reduce the level of lymphocytotoxic antibodies. The decrease in the level of antibodies could be due to massive absorption of antibodies by the liver or to release of major histocompatibility complex antigens in a soluble form. Immunofluorescent examination of the hemoperfused liver revealed important deposits of C3 on Kupffer cells and of IgG on sinusoidal cells. On the contrary, in control rats in which a third-party (DA) liver hemoperfusion was performed, heart allograft survival was less prolonged, the decrease in the level of lymphocytotoxic antibodies was not significant, and the deposits of IgG and C3 were much less evident. The level of circulating immune complexes was unchanged after a donor-specific or a third-party liver hemoperfusion. These results support the hypothesis that resistance of the liver to hyperacute rejection might be due to a massive and nontoxic absorption of lymphocytotoxic antibodies on nonparenchymal liver cells.

Plasma exchange in a rat model of autoimmune glomerulonephritis.

Using an original technique permitting repeated plasma exchange in the rat, we have tested this therapeutic approach in animals actively immunised with horseradish peroxidase, and in rats with HgCl2-induced autoimmune glomerulonephritis. Plasma exchange effectively removes circulating IgG anti-horseradish peroxidase antibodies from the sera of immunised rats. When applied to the model of HgCl2-induced antiglomerular basement membrane glomerulonephritis in Brown-Norway rats, this technique is also remarkably effective. In these rats, proteinuria is abolished during the plasma exchange treatment period and no circulating antiglomerular basement membrane antibodies can be detected. These antibodies are, however, found in the ultrafiltrates of exchanged rats. Serum IgE, characteristically elevated in HgCl2-treated rats, is also markedly diminished in exchanged rats. Control rats treated with infusions of fresh frozen plasma or with heparin alone did not show any improvement in disease severity. These results suggest that plasma exchange alone can attenuate antiglomerular basement membrane nephritis in HgCl2-treated rats. This observation may be of relevance for the treatment of human antiglomerular-basement membrane-mediated glomerulonephritis.

Mercury-induced autoimmune glomerulonephritis: requirement for T-cells.

Mercury-induced autoimmunity in Brown-Norway rats has been shown previously to be due to polyclonal activation of B lymphocytes, requiring the presence of T lymphocytes. Autoimmunity in that strain is characterised by the appearance of an autoimmune glomerulonephritis, by the production of a host of autoantibodies, and by an increase in total serum IgE. In the present study, T-cell deprived rats were tested to assess the role of T cells in the appearance of autoimmune abnormalities in vivo. It will be shown that both BN rnu/rnu and BN 'B' rats, who have virtually no T cells, do not develop autoimmunity following HgCl2 injections. In contrast BN 'B' rats reconstituted with normal T cells, and BN rnu/+ rats, exhibit autoimmune manifestations, including autoimmune glomerulonephritis, quite similar to those observed in Brown-Norway rats. These data demonstrate that T cells are essential for mercury-induced autoimmunity to occur in Brown-Norway rats.

Effect of methylprednisolone and cyclophosphamide in mercury-induced autoimmune glomerulonephritis.

The effects of methylprednisolone and of cyclophosphamide were tested in mercury-induced autoimmune disease in Brown-Norway rats. Survival, proteinuria, presence of antiglomerular basement membrane bound antibodies and of immune complex type deposits, amounts of circulating immune complexes, and total serum IgE were studied. Serum IgE represents the most sensitive marker in this drug-induced autoimmune disease. Methylprednisolone alone (1.5 mg/kg per day) affected the course of the disease only slightly. Cyclophosphamide (20 mg/kg every other day) given from day 0 completely prevented all the autoimmune manifestations, but the rats were profoundly immunosuppressed. The same protective effect was obtained with lower cyclophosphamide dosage (15 mg/kg on day 0 and then 2 mg/kg per day). More interestingly, cyclophosphamide given from day 10 or 15 (20 mg/kg twice a week or every other day), at a time when the disease was already expressed, resulted in partial or complete recovery, provided that the rats had not exhibited heavy proteinuria before initiation of treatment. Cyclophosphamide is therefore a powerful agent, able to prevent and even to reduce the consequences of polyclonal activation in this model.

Effect of prostaglandin E1 in brown Norway rats with mercury-induced autoimmune disease.

The effect of prostaglandin E1 on mercury-induced autoimmune disease in brown Norway rats has been investigated. Daily doses of 6 to 24 micrograms prolonged survival and significantly decreased proteinuria, deposition of immune reactants in the glomeruli, circulating anti-glomerular membrane antibody production, total serum IgE, and circulating immune complex level. A dose of 3 micrograms was also effective but to a lesser degree. These results show the efficiency of prostaglandin E1 in yet another autoimmune disease, show that the beneficial effect of prostaglandin E1 in this model is related to its immunosuppressive effects, and suggest that modification of prostaglandin-mediated suppression induced by HgCl2 might play a role in the pathogenesis of this autoimmune disease.

Autoimmunity induced by HgCl2 in Brown-Norway rats. I. Production of monoclonal antibodies.

Mercuric chloride (HgCl2) induces in Brown-Norway rats (BN) a B cell polyclonal activation resulting in autoimmune disease. Spleen cells from BN rats injected with HgCl2 were fused with IR983F, a nonsecreting rat myeloma cell line, in order to obtain monoclonal antibodies reacting with autoantigens or IgE-producing hybridomas. After screening for immunoglobulin-producing clones, we found 5% clones with anti-tissue activity, 8% with anti-TNP activity, and 41% secreting IgE. Among the anti-tissue monoclonal antibodies, one recognizes both TNP and mesangial structures of rat normal glomeruli, which could be an as yet unrecognized mechanism of nephrotoxicity. These experiments 1) confirm that HgCl2 induces polyclonal activation, 2) show that the mercury model is of interest to obtain monoclonal IgE and various autoantibodies, and 3) suggest a new possible mechanism of antibody-mediated renal injury.

Effect of cyclosporine A on mercury-induced autoimmune glomerulonephritis in the Brown Norway rat.

To test the effect of cyclosporine A (CsA) in mercuric chloride (HgCl2)-induced nephritis in the Brown-Norway (BN) rat, we treated groups of intoxicated rats with varying doses of CsA for a period of 2 months. All manifestations of HgCl2-induced disease were prevented in rats treated concurrently with CsA at either 7 or 10 mg/kg/day. Partial suppression was evident at lower daily doses, but not with bi-weekly CsA administration. The initial phase of HgCl2-induced nephritis could be completely suppressed with a short, 15 day course of CsA. The later phase of the disease could be tempered by CsA administration starting on day 10 after the first HgCl2 injection. The optimal regimen of 7 mg/kg/day for 60 days was not associated with any evidence of CsA toxicity. CsA appears to interfere with the polyclonal activation of B cells observed in HgCl2-induced autoimmune disease, accounting for its striking preventive and curative effect in this model.