Effects of methotrexate on radiologic progression in juvenile rheumatoid arthritis.

OBJECTIVE: To assess the effects of methotrexate (MTX) therapy on radiologic progression in juvenile rheumatoid arthritis (JRA). METHODS: We evaluated serial wrist radiographs for carpal length in 23 JRA patients with bilateral wrist involvement, before and during MTX treatment. These carpal length measurements were compared with established norms for carpal length in a healthy pediatric population. RESULTS: Both clinical responders to MTX (17 of 23 patients) and nonresponders (6 of 23) had decreasing carpal length prior to initiation of the treatment. Eleven of the 17 clinical responders had improved carpal length after a mean of 2.5 years of MTX treatment. All 6 clinical nonresponders had progressive loss of carpal length. CONCLUSION: MTX treatment resulted in radiologic improvement, as measured by carpal length, in the majority of children with JRA who had a clinical response to MTX.

Modulation of murine hepatic lipase activity by exogenous and endogenous Kupffer-cell activation.

Deficiency of hepatic lipase (HL) may play a role in the lipoprotein abnormalities in chronic inflammatory states which are characterized by reticuloendothelial-system activation and cytokine release. HL triacylglycerol hydrolase activity was measured in heparin perfusates of livers from autoimmune MRL/lpr mice, which spontaneously develop a condition closely resembling human lupus erythematosis and exhibit spontaneous Kupffer-cell activation after 8 weeks of age, as well as from normal mice treated with Corynebacterium parvum or polyinosinic-polycytidylic acid complex [poly(I.C)] to induce Kupffer-cell activation. HL activity in MRL/lpr mice older than 8 weeks was 29.5% (P = 0.002) of that in age-matched control MRL/++ mice. Treatment of normal mice with C. parvum or poly(I.C) resulted in HL activities 18.6% (P = 0.004) and 13.1% (P = 0.007) respectively of untreated controls. Northern-blot hybridization of liver poly(A)+ RNA showed no differences in HL mRNA abundance in MRL/++ mice compared with the MRL/lpr autoimmune strain after 8 weeks of age, or in normal control mice compared with those treated with C. parvum, indicating attenuation of HL activity at the translational or post-translational level. Deficiency of this enzyme may represent one of the mechanisms contributing to the dyslipoproteinaemia of autoimmune disease and chronic infection.

Liver of MRL/lpr mice contain interleukin-4-producing lymphocytes and accessory cells that support the proliferation of Th2 helper T lymphocyte clones.

Hepatic nonparenchymal cells (NPC) from mice of nonautoimmune strains support the proliferation of only Th1 and not Th2 helper T lymphocyte (HTL) clones. Because of the multiple systemic and liver-specific immune defects in the autoimmune MRL/lpr mouse strain, we have explored the possibility that hepatic accessory cells from MRL/lpr mice are capable of stimulating the proliferation of Th2 HTL. We report here that hepatic NPC from MRL/lpr and C3H/lpr female mice older than 8 weeks, in contrast to hepatic NPC from MRL/++ and C3H/HeN strains, are able to support in vitro the proliferation of both Th1 and Th2 CD4 clones. Additionally, hepatic lymphocytes (HL) from MRL/lpr mice can be stimulated to produce interleukin (IL)-4 to a much higher degree than HL from the nonautoimmune strains. These results suggest that the activation of Th2 cells by hepatic NPC and production of IL-4 by HL may contribute to the immunologic aberrations in the MRL/lpr mouse strain.

Immune abnormalities in the pathogenesis of juvenile rheumatoid arthritis.

Juvenile rheumatoid arthritis (JRA) is the most common rheumatic disease of childhood. Although the etiology remains unknown, immunoregulatory imbalances are thought to be important in the pathogenesis of JRA. Numerous immunologic abnormalities have been described in these patients, but it remains unclear which are fundamental to the pathogenesis of the disease and which are secondary. In this article, the authors review lymphocyte and lymphokine abnormalities in children with JRA with emphasis on the possible role of these immune abnormalities in the pathogenesis of JRA.

Animal models of chronic inflammatory arthritis.

The use of animals has been indispensable to the investigation of the etiology, pathophysiology, and treatment of juvenile arthritis. Because of ethical concerns about studying children, the difficulty of obtaining tissue, and the heterogeneous manifestations and protean course of chronic arthritis in childhood, the scope of potential research has been severely limited. This brief review summarizes a few of the animal models most commonly used in chronic inflammatory arthritis research: subcutaneous air pouch, antigen-induced (including arthritogenic infectious agent), and spontaneous models. In the spontaneous and antigen-induced animal models of arthritis, local and systemic immunoregulatory abnormalities clearly play a major role in the pathogenesis of arthritis. By elucidating the immune response to those antigens, as well as the role of genetics and environment, the pathogenesis of juvenile rheumatoid arthritis may be better understood. At the present time, however, the complexity of these models precludes more definitive interpretation and extrapolation to human diseases.

The liver of MRL/lpr mice contains defective accessory cells and a population of immunosuppressive lymphocytes.

Immunoregulatory abnormalities in the MRL/lpr mouse strain include activation of macrophages and hepatic natural killer cells, spontaneous production of tumor necrosis factor, defective oral tolerance, and impaired production of interleukin-2. Because the liver is the major organ responsible for the clearance, degradation, and presentation of foreign antigens from the gastrointestinal tract, we have investigated antigen presentation activity of hepatic nonparenchymal cells (NPC) from MRL/lpr, MRL/++, and C3H/HeN female mice in the primary immune response as measured by stimulation of allogeneic one-way mixed lymphocyte response (MLR), and allogeneic cell-mediated lympholysis (CML). Whereas adherent NPC from C3H/HeN, MRL/++, and young MRL/lpr mice were effective stimulators, NPC from MRL/lpr mice older than 9 weeks were defective stimulators of both MLR and CML responses. This abnormality was not observed in splenic accessory cells from these mice. Moreover, a population of hepatic NPC from older MRL/lpr mice are immunosuppressive: mixing of MRL/lpr NPC with splenic stimulators from MRL/++ mice profoundly inhibited primary allogeneic CML responses. The inhibitory hepatic nonparenchymal cell population was nonadherent, radioresistant and was removed by pretreatment with antibodies to either asialoAGM-1 or Lyt-2 plus complement. This inhibition was not observed with the addition of MRL/++ NPC or supernates from cultured MRL/lpr NPC. These findings suggest a selective organ-specific and age-dependent impairment of antigen presentation and the presence of an immunosuppressive lymphocyte population in the liver of MRL/lpr mice which may contribute to the autoimmune process.

Murine hepatic accessory cells support the proliferation of Th1 but not Th2 helper T lymphocyte clones.

The liver is the major site of clearance and degradation of foreign antigens from the portal circulation. Despite the presence of hepatic accessory cells, antibody responses to orally administered antigens are uncommon. To ascertain if hepatic accessory cells are incapable of stimulating specific subsets of T lymphocytes, freshly isolated hepatic nonparenchymal and splenic cells were cultured with a panel of antigen-specific, H-2-restricted Th1 and Th2 HTL clones. Whereas spleen cells stimulated the proliferation of both Th1 and Th2 clones, hepatic nonparenchymal cells (NPC) stimulated the proliferation of only Th1 and not Th2 clones. Adding rIL-1, rIL-6, and rIL-7, alone or in combination, to the cultures did not result in proliferation of the Th2 clones. Despite the absence of Th2 proliferation, NPC were able to stimulate the secretion of IL-3 and IL-4 by Th2 clones in the presence of antigen. Moreover, adding hepatic NPC did not inhibit spleen cells from stimulating Th2 clones in the presence of antigen. Thus, the inability of liver cells to stimulate the proliferation of Th2 helper T lymphocytes appears to be secondary to an absence of either an unknown accessory cell cofactor or an accessory cell that preferentially presents antigen to Th2 cells. The selective activation of Th1 and not Th2 cells by liver accessory cells may result in suppression of antibody responses to orally administered antigens.

Intermittent intravenous cyclophosphamide therapy for lupus nephritis.

We carried out a preliminary study to determine whether intermittent intravenous cyclophosphamide therapy could be safely and effectively used in the treatment of childhood lupus nephritis. Sixteen children (4 to 18 years of age) with lupus nephritis were treated with cyclophosphamide monthly for 6 months and then every 3 months. Eight children were treated because of corticosteroid-unresponsive active lupus nephritis, with a fall in their creatinine clearance to less than 100 ml/min/1.75 m2, and eight children were treated because of corticosteroid-dependent nephrotic syndrome or active lupus nephritis with unacceptable corticosteroid-induced side effects. Cyclophosphamide treatment was associated with significant improvement at 1 year in mean levels of hemoglobin (11.3 +/- 0.5 to 13.1 +/- 0.3 gm/dl), C3 (52 +/- 5.9 to 108 +/- 13.7 mg/dl), and C4 (7.6 +/- 0.9 to 15.9 +/- 2.2 mg/dl) (all p less than 0.005), despite a significant reduction in mean prednisone dosage (31 +/- 5 to 14 +/- 2 mg/day; p less than 0.005). There was a decrease in 24-hour urine protein excretion from 3121 +/- 913 to 1016 +/- 364 mg/24 hours (p less than 0.05). For children whose initial creatinine clearance was less than 100 ml/min/1.75 m2, creatinine clearance also improved significantly (57.5 +/- 11 to 121 +/- 24.5 ml/min/1.75 m2; p less than 0.05). The long-term safety of intravenous cyclophosphamide therapy and its long-term efficacy in comparison with prednisone alone remain to be established. In the interim, intravenous cyclophosphamide therapy should be reserved for children with severe, unacceptable corticosteroid side effects or with corticosteroid-resistant and potentially life-threatening disease.

A comparison of murine hepatic accessory cells and splenic dendritic cells.

Accessory cells are required for proliferation and antibody synthesis of B lymphocytes and proliferation of T lymphocytes in primary immune responses in vitro. The obligatory cells derived from the spleen are referred to as dendritic cells. Accessory cells were isolated from normal adult livers which were functionally interchangeable with splenic DC. Both hepatic accessory cells (AC) and splenic DC adhere firmly to plastic culture dishes or wells within 2 hr; but hepatic AC, unlike splenic DC, do not detach during 22 hr additional incubation. Hepatic AC, unlike splenic DC, are not lysed or inactivated by monoclonal antibody 33D1 and C'. Hepatic AC and splenic DC are similarly sensitive to irradiation in vivo and insensitive to irradiation in vitro. Hepatic AC are separated with cells which are predominantly phagocytic and FcR+ and contain nonspecific esterase. Both hepatic AC and splenic DC are suppressed or eliminated by activation of NK cells in vivo, a phenomenon prevented by prior elimination of NK cells.

Spontaneous production of tumor necrosis factor alpha by Kupffer cells of MRL/lpr mice.

We report that freshly isolated, unstimulated Kupffer cells (KC) from MRL/lpr female mice in short-term culture spontaneously produce high levels of TNF-alpha. TNF production was first detected in KC cultures at age 6 wk and increased with the age of the mice. Moreover, the levels of spontaneous TNF production by KC directly correlated with the age of the MRL/lpr mice. Although TNF production by KC could be induced with C. parvum in vivo or LPS in vitro in all nonautoimmune C3H/HeN, BALB/c, DBA/2, C57B16 mice, the only other strain in which spontaneous TNF production by KC was observed was MRL/++ mice greater than 10 mo old.

Immunoregulatory aberrations in patients with polyarticular juvenile rheumatoid arthritis.

The presence of hypergammaglobulinemia and various circulating autoantibodies in children with polyarticular juvenile rheumatoid arthritis (JRA) implies an immunoregulatory disorder. We report here experiments planned to elucidate the underlying cellular aberrations in this disease. Twelve children with polyarticular JRA were studied. Percentages of Leu-1, Leu-2, and Leu 3 T cells were comparable to those of normal individuals. Immunofluorescent double staining studies demonstrated elevated numbers of activated (DR+) T cells of both Leu-2 and Leu-3 phenotype. B cells characterized both phenotypically (Leu-12) and functionally (as spontaneous plaque-forming cells, PFC) were elevated. In vitro PFC responses to pokeweed mitogen (PWM) and Epstein-Barr virus (EBV) were diminished. The levels of concanavalin A-induced suppressor cells of the PWM-stimulated PFC responses were comparable to control values. In contrast, the EBV-associated suppressor T cells were significantly impaired in both EBV-seropositive and EBV-seronegative patients. These studies indicate that peripheral blood B-cell activity is abnormal in polyarticular JRA. Defective T-cell responses in vitro suggest that this may be due to disruption of normal regulatory circuits between B and T cells and may contribute to the pathogenesis of this disease.

High hepatic natural killer cell activity in murine lupus.

This study demonstrates a profound elevation of NK activity, as measured by cytotoxicity to YAC-1 targets in a 4-h incubation 51Cr-release assay, of freshly isolated hepatic NPC from both MRL/lpr and (NZB X NZW)F1 mice. This marked increase was not observed in splenic or peripheral blood NK. The hepatic NK were nonadherent, radioresistant, Ly-1-,2-, and AGM1+. Furthermore, biologic response modifiers can further augment hepatic NK activity in these autoimmune strains.

Hepatic reticuloendothelial system activation in autoimmune mice: differences between (NZB X NZW)F1 and MRL-lpr/lpr strains.

5'-Nucleotidase (5'N) activity and Ia expression of hepatic nonparenchymal cells (NPCs) of the autoimmune (NZB X NZW)F1 and MRL-lpr/lpr and nonautoimmune C3H/FeJ, DBA/2J, and A/J strains were assayed to determine endogenous activation of the hepatic reticuloendothelial system (RES). Pretreatment of the nonautoimmune strains with Corynebacterium parvum resulted in decreases in Fc receptor expressor and 5'N and an increase in Ia expression of NPCs. Endogenous hepatic RES activation, as measured by low 5'N and high Ia expression, was observed in both the autoimmune MRL-lpr/lpr and (NZB X NZW)F1 strains even without C. parvum pretreatment. However, in the MRL-lpr/lpr strain, age is a dependent variable in activation and correlates with delayed clearance from the circulation of soluble IgG immune complexes. In the (NZB X NZW)F1 strain, the observation of decreased 5'-nucleotidase activity and increased percentage of Ia-positive cells at all ages suggests a primary abnormality. Therefore, different genetic or exogenous factors may be responsible for the activation of the hepatic RES in these autoimmune strains.

Cellular immunity in patients with systemic juvenile rheumatoid arthritis.

We studied the cellular immune responses in 10 patients with systemic form of juvenile rheumatoid arthritis. The numbers of peripheral T lymphocytes and their helper/inducer and cytotoxic/suppressor subpopulation were within normal levels. Activated T lymphocytes (DR+) were slightly increased but not at statistically significant levels. In contrast to the T cells, B lymphocytes were increased; both the percentage of B cells (B1+) and the number of cells spontaneously secreting IgG, IgA, and IgM were increased. Stimulation of peripheral mononuclear cells in vitro with pokeweed mitogen induced poor plaque-forming cell responses, which were partially improved upon removal of monocytes. The presence of concanavalin A in the cultures led to complete suppression. We conclude that patients with systemic JRA are characterized primarily by B-cell rather than T-cell abnormalities.

The immunologic basis of lupus.

This article reviews the immunologic basis of systemic lupus erythematosus. It covers basic immunologic abnormalities, possible role of diet in autoimmune diseases, and the basis for specific abnormalities, such as renal and central nervous system disease.

Abnormal binding of soluble IgG immune complexes to hepatic nonparenchymal cells of autoimmune mice.

Because the liver is the major organ responsible for removal of soluble immune complexes (IC), the surface binding characteristics of preformed model IC to unstimulated mouse liver nonparenchymal cells (NPC) in suspension were studied. NPC of non-autoimmune C3H/FeJ, C3H/HeJ, A/J, DBA/2 and the autoimmune NZB/W F1 and MRL/lpr female mice of various ages were isolated by perfusion of the portal vein with collagenase followed by separation of NPC from hepatocytes with a metrizamide gradient. Thirty-five percent of NPC of all mouse strains were nonspecific esterase-positive and phagocytosed latex beads. Radiolabeled mouse IgG anti-DNP covalently cross-linked stable IC were separated by gel filtration and bound to NPC under various conditions. Marked differences were noted in maximal number of IC bound per cell between the autoimmune and non-autoimmune mouse strains: 3.3 to 4.0 X 10(5) in the non-autoimmune strains vs 0.3 to 1.4 X 10(5) molecules of IC bound per cell in the autoimmune strains at 1 to 6 mo. Insignificant differences were noted in Ka by Scatchard plot analysis (3.5 to 5.0 X 10(8) M-1) and rate of reversibility of binding as determined by dissociation of surface-bound IC with an excess of heat-aggregated gamma-globulin (T 1/2:1.5 to 2 min). These data demonstrate a decreased number of available binding sites for IC in unstimulated NPC from NZB/W F1 and MRL/lpr female mice throughout their life spans. Although the findings are consistent with saturation of binding sites of the NPC with native IC, the abnormality found in the 1-mo-old autoimmune mice (who do not have detectable autoantibodies) suggests a primary defect in FC receptor expression or an altered state of activation of NPC that may contribute to the disease process.