Monocytes Of Systemic Lupus Erythematosus Patients Research Articles

MicroRNA-98-5p Inhibits IFI44L-Mediated Differentiation of Dendritic Cells and Activation of Interferon Pathway in Systemic Lupus Erythematosus

ABSTRACT MicroRNA-98-5p (miR-98-5p) plays a protective role in the pathogenesis of autoimmune diseases through anti-inflammatory effects, but little is known about its role in Systemic lupus erythematosus (SLE). Our previous study suggested Interferon-inducible 44 like (IFI44L) overexpressed in monocytes which contributes to the pathogenesis of SLE by enhancing the maturation and functions of monocyte-derived dendritic cells (Mo-DCs), and miR-98-5p can regulate the expression of IFI44L. In this study, we identified miR-98-5p lowly expressed in both peripheral blood mononuclear cells (PBMCs) and monocytes of SLE patients along with high expression of IFI44L. IFI44L serves as target gene of miR-98-5p which inhibits differentiation of Mo-DCs and IFI44L-mediated activation of interferon pathway. We further showed that miR-98-5p promotes methylation of the IFI44L promoter to down-regulate its expression in SLE. Our results reveal an important role for miR-98-5p in the IFI44L-mediated immune imbalance of SLE and suggest a potential therapeutic target for SLE in the future.

α-Ketoglutarate-Dependent KDM6 Histone Demethylases and Interferon-Stimulated Gene Expression in Lupus.

We aimed to investigate the hypothesis that interferon (IFN)-stimulated gene (ISG) expression in systemic lupus erythematosus (SLE) monocytes is linked to changes in metabolic reprogramming and epigenetic regulation of ISG expression. Monocytes from healthy volunteers and patients with SLE at baseline or following IFNα treatment were analyzed by extracellular flux analysis, proteomics, metabolomics, chromatin immunoprecipitation, and gene expression. The histone demethylases KDM6A/B were inhibited using glycogen synthase kinase J4 (GSK-J4). GSK-J4 was tested in pristane and resiquimod (R848) models of IFN-driven SLE. SLE monocytes had enhanced rates of glycolysis and oxidative phosphorylation compared to healthy control monocytes, as well as increased levels of isocitrate dehydrogenase and its product,α-ketoglutarate (α-KG). Because α-KG is a required cofactor for histone demethylases KDM6A and KDM6B, we hypothesized that IFNα may be driving "trained immune" responses through altering histone methylation. IFNα priming (day 1) resulted in a sustained increase in the expression of ISGs in primed cells (day 5) and enhanced expression on restimulation with IFNα. Importantly, decreased H3K27 trimethylation was observed at the promoters of ISGs following IFNα priming. Finally, GSK-J4 (KDM6A/B inhibitor) resulted in decreased ISG expression in SLE patient monocytes, as well as reduced autoantibody production, ISG expression, and kidney pathology in R848-treated BALB/c mice. Our study suggests long-term IFNα exposure alters the epigenetic regulation of ISG expression in SLE monocytes via changes in immunometabolism, a mechanism reflecting trained immunity to type I IFN. Importantly, it opens the possibility that targeting histone-modifying enzymes, such as KDM6A/B, may reduce IFN responses in SLE.

LncRNA GAS5 as an Inflammatory Regulator Acting through Pathway in Human Lupus.

To investigate the contribution of GAS5 in the pathogenesis of SLE. Systemic Lupus Erythematosus (SLE) is characterized by aberrant activity of the immune system, leading to variable clinical symptoms. The etiology of SLE is multifactor, and growing evidence has shown that long noncoding RNAs (lncRNAs) are related to human SLE. Recently, lncRNA growth arrest-specific transcript 5 (GAS5) has been reported to be associated with SLE. However, the mechanism between GAS5 and SLE is still unknown. Find the specific mechanism of action of lncRNA GAS5 in SLE. Collecting samples of the SLE patients, Cell culture and treatment, Plasmid construction, and transfection, Quantitative real-time PCR analysis, Enzyme-linked immunosorbent assay (ELISA), Cell viability analysis, Cell apoptosis analysis, Western blot. In this research, we investigated the contribution of GAS5 in the pathogenesis of SLE. We confirmed that, compared to healthy people, the expression of GAS5 was significantly decreased in peripheral monocytes of SLE patients. Subsequently, we found that GAS5 can inhibit the proliferation and promote the apoptosis of monocytes by over-expressing or knocking down the expression of GAS5. Additionally, the expression of GAS5 was suppressed by LPS. Silencing GAS5 significantly increased the expression of a group of chemokines and cytokines, including IL-1β, IL-6, and THFα, which were induced by LPS. Furthermore, it was identified the involvement of GAS5 in the TLR4-mediated inflammatory process was through affecting the activation of the MAPK signaling pathway. In general, the decreased GAS5 expression may be a potential contributor to the elevated production of a great number of cytokines and chemokines in SLE patients. And our research suggests that GAS5 contributes a regulatory role in the pathogenesis of SLE, and may provide a potential target for therapeutic intervention.

Long noncoding RNA MALAT-1 is a novel inflammatory regulator in human systemic lupus erythematosus

Despite growing evidence that Long noncoding RNAs (lncRNAs) can regulate gene expression and widely take part in autoimmune and inflammatory diseases, our knowledge of systemic lupus erythematosus (SLE)-related lincRNAs remains limited. In this study, we aimed to explore the contribution of the lncRNA metastasis associated lung adenocarcinoma transcript 1 (MALAT1) to the pathogenesis of SLE. PBMCs were obtained from SLE patients and healthy donors. The expression levels of MALAT-1 were measured by quantitative PCR. Small interfering RNA (siRNA) was then used to knock down the expression of MALAT1 in order to determine the role of MALAT1 in the expression levels of IL-21 and SIRT1 signaling pathway in primary monocytes of SLE patients. Here, we found MALAT-1 expression was abnormally increased in SLE patients and predominantly expressed in human monocytes. Additionally, silencing MALAT-1 significantly reduced the expression of IL-21 in primary monocytes of SLE patients. Furthermore, MALAT-1 exerts its detrimental effects by regulating SIRT1 signaling. Our results demonstrate that MALAT-1 is the key regulatory factor in the pathogenesis of SLE and provides potentially novel target for therapeutic intervention.

Enhanced Inflammasome Activity in Systemic Lupus Erythematosus Is Mediated via Type I Interferon-Induced Up-Regulation of Interferon Regulatory Factor 1.

The inflammasome complex is a driver of organ damage in patients with systemic lupus erythematosus (SLE). Although type I interferons (IFNs) are well established as mediators of SLE pathogenesis, their role in inflammasome activation in SLE has not been assessed. The aim of this study was to examine type I IFNs as regulators of the inflammasome. SLE patients fulfilled ≥4 American College of Rheumatology criteria and were recruited from the University of Michigan Lupus Cohort. Primary monocytes were isolated from SLE patients or healthy controls by negative selection, treated with inflammasome activators in the presence or absence of IFNα, and IL-1β secretion was measured by enzyme-linked immunosorbent assay. Expression levels of IFN and inflammasome-related molecules were assessed by real-time polymerase chain reaction and Western blotting. IFN regulatory factor 1 (IRF-1) expression was specifically down-regulated by small interfering RNA (siRNA) transfection and a chemical inhibitor. Monocytes from patients with SLE exhibited increased expression and enhanced activation of the inflammasome by ATP when compared with control monocytes. Expression of inflammasome and IFN-regulated genes was significantly correlated in monocytes from SLE patients but not in control monocytes. Inflammasome activity was increased after prolonged exposure to IFNα. Reduction of IRF-1 expression via siRNA blocked caspase 1 up-regulation after treatment with IFNα. Importantly, hyperactivity of the inflammasome in the monocytes of SLE patients was significantly reduced after knockdown or inhibition of IRF-1. Prolonged type I IFN exposure, as seen in SLE patients, primes monocytes for robust inflammasome activation in an IRF-1-dependent manner. IRF-1 inhibition may serve as a novel target for treatment of SLE-associated inflammation and organ damage.

MicroRNA-302d targets IRF9 to regulate the IFN-induced gene expression in SLE

Systemic lupus erythematosus (SLE) is a complex disease targeting multiple organs as a result of overactivation of the type I interferon (IFN) system, a feature currently being targeted by multiple biologic therapies against IFN-α. We have identified an estrogen-regulated microRNA, miR-302d, whose expression is decreased in SLE patient monocytes and identify its target as interferon regulatory factor (IRF)-9, a critical component of the transcriptional complex that regulates expression of interferon-stimulated genes (ISGs). In keeping with the reduced expression of miR-302d in SLE patient monocytes, IRF9 levels were increased, as was expression of a number of ISGs including MX1 and OAS1. In vivo evaluation revealed that miR-302d protects against pristane-induced inflammation in mice by targeting IRF9 and hence ISG expression. Importantly, patients with enhanced disease activity have markedly reduced expression of miR-302d and enhanced IRF9 and ISG expression, with miR-302d negatively correlating with IFN score. Together these findings identify miR-302d as a key regulator of type I IFN driven gene expression via its ability to target IRF9 and regulate ISG expression, underscoring the importance of non-coding RNA in regulating the IFN pathway in SLE.

Validating the expression of miRNAs in healthy controls and SLE patients and to predict possible gene targets

Background Systemic Lupus Erythematosus (SLE) is a multisystem autoimmune connective tissue disease that has a strong female predominance (9:1) seen especially in women of child-bearing years [1]. Although the exact mechanisms of disease are not completely understood, the role of dysregulated microRNA (miRNA) has been implicated in the pathogenesis of SLE [2]. Clinically, the disease is unpredictable, with the current therapy for symptomatic patients associated with undesirable side effects and toxicity affecting multiple organs [3]. Therefore, in order to bridge the gap between disease and cure, an in depth understanding of disease pathogenesis is required to manufacture medications that tackle the underlying cause of the disease. In this study, we set out to validate differentially expressed miRNAs in SLE patient monocytes versus healthy controls, previously identified in a microRNA screen carried out in the group. Furthermore we aim to perform bioinformatics analysis to predict potential gene targets for these miRNAs that may play a role in the disease pathogenesis. Methods RNA was extracted from monocytes from SLE patients and healthy controls previously collected in the laboratory. Bioinformatics analysis was undertaken to identify potential genes targeted by miRNAs of interest. The primers specific to the miRNAs were designed and optimised following which gene induction of miRNAs was determined by PCR. In order to investigate the expression of miRNAs, densitometric analysis of the gel electrophoresis was determined. Results In the study, the expression of miRNA-107 and miRNA132 were investigated. The expression of miR-107 was significantly greater in SLE patients than in healthy controls (p<0.00362), whilst there were no appreciable differences in miRNA-132 expression in SLE patients and healthy controls. SMURF1 and SOCS5 were identified as potential genes targeted by miRNA-107, however no significant change in expression in either genes were observed in SLE patients versus controls. Conclusions There was a significant increase in the expression levels of miRNA-107 in SLE patients, while no significant changes in miRNA-132 was demonstrated. Further investigation into potential gene targets for miR-107 is required in order to understand the pathological importance of dysregulation of this miRNA in SLE and its therapeutic implications

ID: 209: Single cell interferon signatures in lupus patient monocytes reveal a differential impact of interferon signaling between monocyte subtypes

Type I interferon (IFN) is a primary pathogenic factor in human systemic lupus erythematosus (SLE). IFN signatures have been observed in immune cell populations. We examined gene expression in individual SLE patient monocytes in this study. CD14+ + CD16− classical monocytes (CLs) and CD14dimCD16+ non classical monocytes (NCLs) from SLE patients were purified by magnetic separation. The Fluidigm C1 System was used for single cell capture and target gene pre-amplification. Rt-PCR was used to quantify expression of 87 monocyte-related genes. An individual cell IFN score was generated based upon the expression of 17 IFN-induced genes. Monocytes from the same SLE patient blood sample demonstrated varying levels of IFN-induced gene expression. In CLs, high IFN score correlated with CD32a, IL1B, and IL8 expression. In NCLs, high IFN score was correlated with inflammatory mediators, including cytokines such as IL12, IL23, and IL15; the immune receptors CD36, CD32a, CD80, and TLR7; and inflammatory signaling genes such as RELA, STAT2, IRAK1, and MyD88. CD16 transcripts were detected in a small group of classical monocytes, despite a lack of surface CD16 expression. In these cells, CD16 expression was positively correlated with IFN score (p = 0.019). This study reveals diverse IFN responses of individual monocytes, supports the idea that IFN signaling has distinct effects upon classical and non-classical monocytes. IFN may contribute to the transition from classical to non-classical subtype in SLE. Single cell studies can reveal effects of IFN on single immune cells which may be masked in whole blood or mixed cell populations.

Single cell gene expression in lupus patient monocytes reveals a differential impact of interferon signaling between monocyte subtypes (TECH2P.920)

Abstract Background/Purpose Interferon (IFN) gene expression signatures have been observed in whole blood and mixed immune cell populations in systemic lupus erythematosus (SLE), but the significance of this IFN signaling in immune cell subsets is not well understood. We examined gene expression in individual SLE patient monocytes to explore the impact of increased IFN-induced transcription in single cells. Methods CD14++CD16- classical monocytes and CD14dimCD16+ non classical monocytes from SLE patients were purified by magnetic separation. Single cells were isolated, and gene expression in each individual cell was examined by rtPCR. Results Monocytes from the same SLE patient blood sample demonstrated varying levels of IFN-induced gene expression. In classical monocytes, high IFN score correlated with CD32a, IL1B, and IL8 expression. In non-classical monocytes, high IFN score was correlated with a larger number of inflammatory mediators, including cytokines such as IL12, IL23, and IL15; the immune receptors CD36, CD32a, CD80, and TLR7; and inflammatory signaling genes such as RELA, STAT2, IRAK1, IRAK4, and MyD88. Conclusion We observed striking diversity in the IFN responses of individual monocytes, demonstrating that IFN signaling has distinct effects upon classical and non-classical monocytes. Single cell studies can reveal effects of IFN on single immune cells and uncommon populations such as non-classical monocytes, which would be masked in whole blood or mixed cell populations.

Interferon regulatory factor 1 marks activated genes and can induce target gene expression in systemic lupus erythematosus.

Interferon regulatory factor 1 (IRF-1) mediates both induction of interferons (IFNs) and responses to type I IFNs. It has been implicated as a critical mediator of inflammation in murine lupus models. In a previous study of chromatin modifications in monocytes from patients with systemic lupus erythematosus (SLE), IRF-1 was implicated as being associated with increased histone acetylation in this disease. The present study was undertaken to directly investigate IRF-1 binding sites on chromatin. Cells from 9 female SLE patients and 7 female controls were examined. Monocytes were purified from peripheral blood and subjected to library preparation using a validated antibody to IRF-1. IRF-1 binding sites on chromatin were identified by chromatin immunoprecipitation followed by sequencing. The effect of IRF-1 on target gene expression was confirmed using an overexpression system in cell lines, and coimmunoprecipitation was used to identify protein interactions. IRF-1 binding around transcribed regions was increased in SLE patient monocytes, but histone modifications at potential IRF-1 binding sites without detectable IRF-1 binding were increased as well. Overexpression of IRF-1 was sufficient to drive transcription of target genes. IRF-1 overexpression was also able to alter histone modifications at a focus set of target genes, and treatment with an IRF-1 inhibitor reduced both expression and histone modifications at target genes. IRF-1 was found to interact with a select set of histone-modifying enzymes and other transcription factors. IRF-1 is an important signaling protein in the interferon pathway. It not only activates gene expression as a transcription factor, but may perpetuate disease by leading to a dysregulated epigenome.

Generation of Tolerogenic Dendritic Cells from Systemic Lupus Erythematosus Patients

Generation of Tolerogenic Dendritic Cells from Systemic Lupus Erythematosus Patients

AB0480 Towards A Novel Tolerogenic Dendritic Cell Based Immunotherapy for Systemic Lupus Erythematosus

BackgroundTolerogenic dendritic cells (tolDCs) can be considered as an attractive strategy to treat autoimmune diseases. The tolerogenic potential of autologous tolDCs has not yet been evaluated to treat systemic lupus...

Interferon regulatory factor 5 activation in monocytes of systemic lupus erythematosus patients is triggered by circulating autoantigens independent of type I interferons

Genetic variants of interferon regulatory factor 5 (IRF-5) are associated with susceptibility to systemic lupus erythematosus (SLE). IRF-5 regulates the expression of proinflammatory cytokines and type I interferons (IFNs) believed to be involved in the pathogenesis of SLE. The aim of this study was to determine the activation status of IRF-5 by assessing its nuclear localization in the immune cells of SLE patients and healthy donors, and to identify SLE-associated triggers of IRF-5 activation. IRF-5 nuclear localization in subpopulations of peripheral blood mononuclear cells from 14 genotyped SLE patients and 11 healthy controls was assessed using imaging flow cytometry. The activation and function of IRF-5 were examined after ex vivo stimulation of healthy donor monocytes with SLE serum or components of SLE serum. Cellular localization was determined by ImageStream flow cytometry, and cytokine expression was analyzed by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. IRF-5 was activated in a cell type-specific manner; monocytes from SLE patients had constitutively elevated levels of nuclear IRF-5, as compared to natural killer cells and T cells. SLE serum was identified as a trigger for IRF-5 nuclear accumulation; however, neither IFNα nor SLE immune complexes could induce nuclear localization. Instead, autoantigens composed of apoptotic/necrotic material triggered IRF-5 nuclear accumulation in monocytes. Production of the cytokines IFNα, tumor necrosis factor α, and interleukin-6 in monocytes stimulated with SLE serum or autoantigens was distinct, yet showed a correlation with the kinetics of IRF-5 nuclear localization. This study provides the first formal proof that IRF-5 activation is altered in the monocytes of SLE patients, which can be attributed, in part, to the SLE blood environment.

Suppression of Mycobacterium Tuberculosis Induced Reactive Oxygen Species andTumor Necrosis Factor-Alpha Activity in Human Monocytes of Systemic LupusErythematosus Patients by Reduced Glutathione

The etiology and pathogenesis of systemic lupus erythematosus remains unknown, evidence exists for the involvement of mycobacterial antigen. This study is aimed to determine the effect of Mycobacterium tuberculosis on clinical course of SLE patients and the role of ROS and TNF-α in the pathogenesis of tuberculosis associated SLE patients. This study was done on 100 patients divided into SLE group (n=30), TB group (n=30), SLE-TB group (n=30) and control group (n=10). All patients underwent clinical, biochemical and immunological evaluation by employing techniques such as SDS-PAGE, direct binding and competition ELISA, PBMC and cell culture. Fever, arthritis, skin rash, photosensitivity were more common in both SLE and SLE-TB group. Reduced glutathione showed amelioration of ROS and TNF-α induced action, which in turn, subsequently suppressed the immune-bindings observed in monocytes of TB and SLE patients cultured without glutathione. Data shows that SLE patients are more susceptible to developing Mycobacterium tuberculosis, as ROS and TNF-α in SLE patients could activate the replication of mycobacterial Ag85B (30 kDa) after bacilli infection.

Disease activity in systemic lupus erythematosus is associated with an altered expression of low‐affinity Fcγ receptors and costimulatory molecules on dendritic cells

Dendritic cells (DCs) play a pivotal role in the interface between immunity and maintenance of peripheral tolerance. The capture of immunoglobulin G (IgG)-containing immune complexes (ICs) by low-affinity Fc gamma receptors (Fc gammaRs) expressed on DCs may influence the immunogenicity/tolerogenicity of these cells, depending on the activating/inhibitory potential of Fc gammaRs. Because of the key role that low-affinity Fc gammaRs play in determining the magnitude of the response in IC-driven inflammation, these receptors are likely to play a role in autoimmune diseases, such as systemic lupus erythematosus (SLE). To evaluate if an altered expression of costimulatory molecules and/or Fc gammaRs could account for disease severity, we evaluated the expression of these molecules on immature and mature DCs derived from peripheral blood monocytes of SLE patients and healthy donors. Our results show an increased expression of the costimulatory molecules CD40 and CD86. Furthermore, the ratio of CD86/CD80 is higher in SLE patients compared with healthy donors. Conversely, while the expression of activating Fc gammaRs was higher on DCs from SLE patients, expression of inhibitory Fc gammaRs was lower, compared with DCs obtained from healthy donors. As a result, the activating to inhibitory Fc gammaR ratio was significantly higher in DCs from SLE patients. The altered ratio of activating/inhibitory Fc gammaRs on mature DCs showed a significant correlation with the activity of SLE, as determined by the SLE Disease Activity Index (SLEDAI) score. We postulate that the increased ratio of activating/inhibitory Fc gammaRs expressed on DCs from SLE patients can contribute to the failure of peripheral tolerance in the IC-mediated phase of autoimmune pathogenesis.

Flow cytometry analysis of glucocorticoid receptor expression and binding in steroid-sensitive and steroid-resistant patients with systemic lupus erythematosus

IntroductionGlucocorticoid (GC) therapy is the main treatment for systemic lupus erythematosus (SLE). However, some patients are resistant to these agents. Abnormalities of glucocorticoid receptor (GR) seem to be related to steroid resistance. This study evaluated GRs in T lymphocytes and monocytes of SLE patients by flow cytometry (FCM) using a monoclonal antibody (mAb) and FITC-Dex probes.MethodsThirty-five patients with SLE before treatment and 27 age- and sex-matched normal controls were studied. Disease activity scores were determined before and after treatment and used to divide the patients into steroid-resistant (SR) and steroid-sensitive (SS) groups. GRs in T lymphocytes (CD3+) and monocytes (CD14+) were examined by FCM with GR-mAb and FITC-Dex probes before treatment. Peripheral blood mononuclear cells (PBMCs) were isolated for in vitro GCs sensitivity assays. The validity of FCM analysis of intracellular staining for GR with GR-mAb and FITC-Dex probes was evaluated through comparison with western blot and radioligand binding assay (RLBA) in U937 and K562 cells in vitro. One-way ANOVA, student's t test, linear regression and spearman correlation were performed.ResultsA significant decrease in GR binding and the expression in K562 and U937 cells with 10-6 M dexamethasone (Dex) was found compared with those without Dex. In addition, a positive correlation was found between FCM and RLBA as well as FCM and Western blot. The expression and binding of both CD3/GR and CD14/GR in SR patients with SLE, detected by FCM, were all lower than those in SS patients with SLE, whereas there was no significant difference in SS patients and controls. In vitro corticosteroid sensitivity assay indicated that PHA-stimulated tumour necrosis factor-α (TNF-α), IL-12 and interferon-γ (IFN-γ) secretion was significantly inhibited by 10-6 M Dexamethasone in all controls and SS patients, compared with that in SR group, which confirms patient classification as SR and SS by disease activity index (SLEDAI) score.ConclusionsAbnormalities of expression and binding of the GR may be involved in tissue resistance to steroids in SLE patients. Determination of GR expression and binding by FCM may be useful in predicting the response to steroid treatment of SLE patients.Trial registrationClinical trial registration number NCT00600652.

Abnormal cell-specific expressions of certain protein kinase C isoenzymes in peripheral mononuclear cells of patients with systemic lupus erythematosus: effect of corticosteroid application.

We have studied the expressions of various protein kinase C (PKC) isoenzymes in T cells and monocytes from patients with systemic lupus erythematosus (SLE), in comparison to those of healthy controls and patients with other immunological disorders. As measured by Western blotting, the levels of PKCbeta, delta, eta, epsilon, theta and zeta (but not of PKCalpha) significantly decreased in T cells of SLE patients. In monocytes, however, we observed marked suppressions only in the expressions of PKCdelta, epsilon and zeta but not in the expressions of other PKC isoforms. In vivo corticosteroid application, as well as in vitro steroid treatment of monocytes, elevated the expressions of most isoforms close to normal values; however, the decreased levels of PKCtheta and zeta were not affected by steroid application. These alterations were characteristic to SLE because we could not detect any changes in the PKC levels in mononuclear cells of primary Sjögren's syndrome and mixed connective tissue disease patients. These results suggest that impaired PKC isoenzyme pattern may exist in the T cells and monocytes of SLE patients. Furthermore, the clinically efficient glucocorticoid application in SLE can increase the expression of some members of PKC system.

IL-10 stimulates production of platelet-activating factor by monocytes of patients with active systemic lupus erythematosus (SLE).

IL-10 displays modulatory properties on the synthesis of platelet-activating factor (PAF), a potent inflammatory mediator of vascular injury. Despite the fact that IL-10 is considered to be an anti-inflammatory cytokine, IL-10 levels correlate with disease activity in SLE. Moreover, in SLE IL-10 is unable to exert its immunosuppressive and anti-inflammatory effects. We have investigated the ability of IL-10 to stimulate PAF production from monocytes of SLE patients. Spontaneous and IL-10-stimulated PAF production by peripheral blood monocytes was measured in active (n = 13) and inactive (n = 14) SLE patients and in 15 normal control subjects. We observed that monocytes derived from patients with active SLE, but not from controls or inactive SLE, spontaneously produced significant amounts of PAF. Moreover, IL-10 enhanced the synthesis of PAF from monocytes of active SLE patients only. IL-10-induced PAF production correlated with the severity of the disease and with the extent of proteinuria. These results indicate that IL-10 only stimulates the synthesis of PAF from monocytes of SLE patients when immunologically active, suggesting that IL-10 may possess a paradoxical proinflammatory effect in SLE by promoting the production of PAF, a secondary mediator of inflammation.

IMPAIRED PRODUCTION OF IL-12 SYSTEMIC LUPUS ERYTHEMATOSUS. I. EXCESSIVE PRODUCTION OF IL-10 SUPPRESSES PRODUCTION OF IL-12 BY MONOCYTES

Interleukin 12 (IL-12) plays a crucial role in defensive immune responses, modulation of cytokine production and is involved in the pathogenesis of some autoimmune diseases. The authors investigated whether decreased in vitro production of IL-12 occurs in systemic lupus erythematosus (SLE), in which the cytokine secreting pattern is predominantly type 2. IL-12 production by SLE peripheral blood mononuclear cells (PBMC) was significantly impaired compared with normal PBMC, and this was not due to decreased numbers of monocytes. After depletion of non-adherent cells from PBMC, monocytes of SLE patients produced significantly less IL-12 than those of controls, but IL-12 levels in SLE and control non-adherent cell supernatants were not significantly different. Exogenous recombinant (r)IL-10 strongly inhibited IL-12 production by both SLE and normal PBMC and anti-IL-10 neutralizing antibody significantly reversed the IL-12 deficiency of SLE PBMC and SLE monocytes, while not affecting normal PBMC. Recombination interferon γ (rIFN-γ) considerably enhanced IL-12 production in both SLE and normal PBMC, but it did not significantly reverse the inhibitory effect of rIL-10 on IL-12 production. IL-12 production was significantly lower in patients with active SLE than those in remission. These results suggest that SLE monocytes may be deficient in IL-12 production and that this is secondary to abnormal production of various cytokines, especially excessive production of IL-10.

Binding affinity and quantity of estrogen receptor in peripheral blood monocytes of patients with systemic lupus erythematosus

Binding affinity and quantity of the estrogen receptor in monocytes of patients with systemic lupus erythematosus (SLE) were studied. The tritiated-estradiol binding assay was performed using peripheral blood adherent cells (> 95% monocytes) derived from six lupus patients (SLEDAI score: 2-30) and five age-comparable normal women during the mid-follicular phase of the menstrual cycle. Dissociation constant (Kd) and number of binding sites (Ro) were estimated by Scatchard analysis. The specificity and sensitivity of the assay were verified by using estrogen receptor-positive ZR75-1 human breast cancer cells. Kd and Ro of the type I receptor for the SLE patients were 12.2 +/- 6.5 (nM) and 69.0 +/- 42.4 (x 1000/cell), respectively, while those of the normals were 14.5 +/- 3.7 and 86.8 +/- 23.4, respectively. Three patients displayed relatively low Kd or Ro values. While those low values fell within the mean -3s.d. of the normal controls, precise statistical comparison was not possible. No clear correlation between the receptor parameters and the SLEDAI scores was noted. Although further studies of a larger number of samples are needed to conclude, these results suggest that peripheral blood monocytes of SLE patients express the estrogen receptor whose Kd and Ro are similar to those of normals.