Elias Toubi

Intravenous immunoglobulin therapy affects T regulatory cells by increasing their suppressive function.

Intravenous Ig therapy (IVIg) is reported to be a useful regimen in treating autoimmune diseases. In this study, we asked whether IVIg (in vitro) could increase the expression of TGF-β, IL-10, and the transcription factor FoxP3 in T regulatory (Treg) cells, and the idea that IVIg could enhance suppressive properties of these cells. CD4+ T cells from 12 healthy individuals were cultured in the presence or absence of IVIg vs human control IgG during 16, 24, and 36 h. Using FACS analysis and gating on CD4+CD25high Treg cells, we assessed the expression of intracellular TGF-β, IL-10, and FoxP3. In addition, the production of TNF-α by stimulated CD4+ T cells alone or in culture with CD25+ by itself or together with IVIg was also assessed. The presence of IVIg with Treg cells in culture significantly increased the intracellular expression of TGF-β (17.7 ± 8.5% vs 29.8 ± 13%; p = 0.02), IL-10 (20.7 ± 4.7% vs 34.2 ± 5.2%; p = 0.008) and FoxP3 (20.8 ± 5.2% vs 33.7 ± 5.9%; p = 0.0006) when compared with cells cultured alone or with control human IgG. The suppressive effect of CD4+CD25+ T cells presented as the decrease of TNF-α production by stimulated CD4+CD25− (effector T cells) was further increased by adding IVIg to cell culture. We hereby demonstrate an additional mechanism by which IVIg could maintain self-tolerance and decrease immune-mediated inflammation.

Accelerated Fas-Mediated Apoptosis of Monocytes and Maturing Macrophages from Patients with Systemic Lupus Erythematosus: Relevance to In Vitro Impairment of Interaction with iC3b-Opsonized Apoptotic Cells

Impaired handling of apoptotic cells has been suggested as an important factor in the development of systemic lupus erythematosus (SLE), and a role for complement in the removal of apoptotic cells was shown recently. We studied the in vitro function of macrophages from 40 patients with SLE and their matched controls in the removal of heterologous apoptotic cells opsonized by iC3b. Interaction index of apoptotic cells opsonized by iC3b was significantly lower in patients with SLE and averaged 71% ± 37 of that of healthy individuals (p < 0.002) and 69% ± 35 of patients with rheumatoid arthritis (p < 0.007). SLE patients had increased apoptosis of both freshly isolated monocytes (p < 0.001) and maturing macrophages (p < 0.04) that led to decreased density of monocyte-derived macrophages. Apoptosis was inhibited by adding soluble Fas receptor indicating Fas-mediated apoptosis. As demonstrated in both healthy controls and patients with SLE, decreased macrophage density by itself caused significant decreased uptake of apoptotic cells by the remaining macrophages. Maintaining normal density in SLE patients either by an increased initial density or by using soluble Fas restored the interaction capacity of the individual macrophages in the majority of patients. We concluded that impaired in vitro interaction of iC3b-opsonized apoptotic cells with macrophages from patients with SLE was mainly associated with Fas-dependent accelerated apoptosis of the monocytes/macrophages. Accelerated apoptosis of phagocytes may represent a novel in vitro mechanism of impairment of interaction with apoptotic cells that, apart from reducing the number of professional phagocytes, alters the function of the remaining macrophages.

Regulatory T cells (CD4+CD25brightFoxP3+) expansion in systemic sclerosis correlates with disease activity and severity

BACKGROUND The role and function of T regulatory (Treg) cells have not been fully investigated in patients with systemic sclerosis (SSc). METHODS Ten patients with SSc donated 20ml of peripheral blood. Activity (Valentini) and severity (Medsger) scores for SSc were calculated for all patients. Healthy volunteers (controls) were matched to each patient by gender and age. CD4(+) cells were separated using the MACS system. The numbers of Treg cells were estimated by flow cytometry after staining for CD4, CD25, and FoxP3 and calculated as patient-to-control ratio separately for each experiment. Correlations with activity and severity indices of the disease were performed. Twenty-four-hour production of TGF-beta and IL-10 by activated CD4(+) cells was measured by ELISA in culture supernatants. RESULTS The numbers of Treg cells, expressed as patient-to-control ratio, correlated significantly with both activity and severity indices (r=0.71, p=0.034 and r=0.67, p=0.044, respectively). ELISA-measured production of TGF-beta and IL-10 by CD4(+) cells was similar in patients and controls. CONCLUSIONS Increased numbers of Treg cells are present in patients with SSc, correlating with activity and severity of the disease. This expansion of Treg cells was not accompanied, however, by heightened TGF-beta or IL-10 production. Further studies to elaborate the causes and functional significance of Treg cell expansion in SSc are needed.

Changes in macrophage function after rituximab treatment in patients with rheumatoid arthritis

Objective: To assess changes in macrophage phenotype and function after rituximab-induced B cell depletion in patients with rheumatoid arthritis (RA). Methods: 10 patients with RA were treated with rituximab, achieving significant B cell depletion 4 months later. Clinical improvement, rheumatoid factor (RF), anti-cyclic citrullinated peptide (anti-CCP) antibodies, mRNA of B cell activating factor (BAFF), interleukin (IL) 10 and CD86 in human monocyte-derived macrophages (HMDMs) and tumour necrosis factor α (TNFα) secretion from cultured HMDMs were assessed at baseline and after the depletion. Results: A clinical response of American College of Rheumatology (ACR) 50% improvement was noted in six patients, and another two patients responded with moderate improvement, equivalent to ACR 20–50% improvements. RF and anti-CCP antibodies were positive at baseline in seven of ten patients. RF disappeared or declined in six patients 4 months after treatment, correlating with clinical improvement. By contrast, anti-CCP remained unchanged in six patients. After rituximab treatment, and in association with clinical improvement, BAFF, IL10 and CD86 mRNA expression in HMDM were significantly upregulated compared with values at baseline. A significant decrease in TNFα in the supernatant of cultured HMDM was also noted. Conclusions: In addition to B cell depletion and attenuation in some of the specific autoantibodies, clinical improvement in rituximab-treated patients with RA occurred in association with changes in macrophage function.

Rituximab: Beyond Simple B Cell Depletion

Rituximab, a chimeric anti-CD20 monoclonal antibody, has a proven track record for over a decade in the treatment of lymphomas, where it has been used to eradicate malignant lymphocytes. In appreciation of the putative role of B cells, especially with respect to autoantibody production, in the pathogenesis of autoimmune diseases, successful trials of B-cell depletion therapy in RA, SLE, and other autoimmune diseases have been carried out. In these trials, clinical benefit has generally correlated with the extent and duration of B-cell depletion, but at times imperfectly, and autoantibody reduction only selectively. Additional mechanisms whereby rituximab may assert its clinical benefit in autoimmune diseases have been examined including a look at B-cell functions as T-cell modulator and antigen-presenting cell, T-regulatory cell behavior, NK cell activity, and macrophage activities in immune inflammation. The available data on rituximab’s action in autoimmune diseases is reviewed.

Semaphorin 3A is a marker for disease activity and a potential immunoregulator in systemic lupus erythematosus

IntroductionSemaphorin 3A (sema3A) and neuropilin-1 (NP-1) play a regulatory role in immune responses and have a demonstrated effect on the course of collagen induced arthritis. This study was undertaken to evaluate the role of sema3A and NP-1 in the pathogenesis of systemic lupus erythematosus (SLE) and the specific effect of sema3A on the auto-reactive properties of B cells in SLE patients.MethodsThirty two SLE and 24 rheumatoid arthritis (RA) patients were assessed and compared with 40 normal individuals. Sema3A serum levels were measured and correlated with SLE disease activity. The in vitro effect of sema3A in reducing Toll-like receptor 9 (TLR-9) expression in B cells of SLE patients was evaluated.ResultsSema3A serum levels in SLE patients were found to be significantly lower than in RA patients (55.04 ± 16.30 ng/ml versus 65.54 ± 14.82 ng/ml, P = 0.018) and lower yet than in normal individuals (55.04 ± 16.30 ng/ml versus 74.41 ± 17.60 ng/ml, P < 0.0001). Altered serum sema3A levels were found to be in inverse correlation with SLE disease activity, mainly with renal damage. The expression of both sema3A and NP-1 on B cells from SLE patients was significantly different in comparison with normal healthy individuals. Finally, when sema3A was co-cultured with cytosine-phosphodiester-guanine oligodeoxynucleotides (CpG-ODN)-stimulated B cells of SLE patients, their TLR-9 expression was significantly reduced, by almost 50% (P = 0.001).ConclusionsThis is the first study in which a reduced serum level of sema3A was found in association with SLE disease activity. It also raises the possibility that sema3A may have a regulatory function in SLE.

The role of T regulatory cells in human sepsis.

It is well-known that septic shock undermines immune homeostasis by inducing an initial intense systemic inflammatory response that is rapidly followed by a negative feedback of anti-inflammatory process. This secondary immunoparalysis state is characterized by decreased phagocytic cells, T cells, natural killer cells and B cells function and proinflammatory cytokine release. This persistence of immunoparalysis increased the risk for fatal outcome. In recent studies it was found that following the onset of septic shock, a relative increase in T regulatory cells number and suppressive function appears and makes them an important participant in the inhibition of immune responsiveness during sepsis. Consequently, a question emerging from these findings concerns the degree to which the manipulation of T regulatory cells might improve the outcome of patients with sepsis. Preliminary studies in animal models suggest that more work is needed to understand the conditions under which such a therapy may be effective.

Semaphorins: Their Dual Role in Regulating Immune-Mediated Diseases

The semaphorin family members were originally considered to play a role in neurodevelopment, angiogenesis, tumor development, and metastasis. Over the past few years, a growing body of data indicates that semaphorins are involved in the regulation of the immune system, identified in this case as the “immune semaphorins.” These semaphorins are involved in almost all phases of both normal and pathological immune responses and were demonstrated to participate in allergic diseases as well as in auto-immune pathologies. Some of them, such as semaphorin 3A (sema3A), is important in downregulating autoimmune diseases by suppressing the over-activity of both T and B cell autoimmunity. In addition, sema3A was shown to enhance the ability of T and B cell regulatory properties and by doing so to control autoimmune diseases, such as systemic lupus erythematosus. Other semaphorins, such as semaphorins 4D and 4A are important in stimulating T and B cells, thus keeping these immune responses on-going. However, when overexpressed, they can induce the induction of many immune-mediated diseases. The importance of all this is to develop targeting therapies that could possibly enhance or alternatively suppress these molecules. In this review, we will focus on several immune semaphorins—their role in immune homeostasis and in immune-mediated diseases.

Cisplatin treatment triggers familial mediterranean fever attacks

A 42-year-old familial Mediterranean fever (FMF) patient who was treated with cisplatin-based chemotherapy for adenocarcinoma of the lung developed severe and frequent attacks of FMF during treatment. Abdominal pain, arthralgia and fever occurred for a few days following each cisplatin cycle. His FMF worsened, the abdominal pain and fever lasted longer and treatment with colchicine was ineffective. It has been hypothesized that the link between cisplatin treatment and FMF attacks lies in an increased production of serotonin, IL-6, IL-1, IL-8 and TNF-alpha. These inflammatory cytokines have been reported to be overproduced during cisplatin treatment and are known to play an important role in FMF relapse. The oncologist should be made aware of the possibility of disease aggravation in FMF patients during cisplatin-based chemotherapy.

Antibody clustering helps refine lupus prognosis.

OBJECTIVE Our aim was to investigate a possible association between patterns of anti-dsDNA antibody isotypes (IgG, IgM, and IgA), rheumatoid factor (RF) isotypes (IgG, IgM, IgA), and IgG anti-C reactive protein (CRP) and systemic lupus erythematosus (SLE) disease activity (SLEDAI). METHODS Our study group included 98 patients, 86 women and 12 men, with a mean SLEDAI score of 7.9 +/- 4.1. We divided the patients into 4 groups by the serum anti-dsDNA antibody isotype intensity level. RESULTS We found that patients in group 1 (IgG > IgM, 42 patients) had a statistically significantly higher SLEDAI score than group 2 (IgG < IgM, 13 patients) (10.57 +/- 4.62 versus 5.6 +/- 4, P = 0.0012), group 3 (IgG = IgM, 8 patients) (10.57 +/- 4.62 versus 6.2 +/- 1.98, P = 0.04), and group 4 (none, 35 patients) (10.57 +/- 4.62 versus 6 +/- 1.5, P = 0.0001). SLE patients with IgG RF or IgM RF isotype present had a significantly higher SLEDAI score compared with those without IgG RF or IgM RF (10.57 +/- 4.8 versus 7.6 +/- 4.1, P = 0.03, 10.6 +/- 5 versus 7.6 +/- 3.9, P = 0.046). The presence of IgA RF isotype was not associated with a higher SLEDAI score. IgG anti-CRP did not correlate differentially with SLEDAI scores. CONCLUSIONS A combination of high-titer IgG anti-dsDNA with a positive RF of IgM isotype may serve as a marker for more active SLE.