Alma-Martina Cepika

Effect of steroids on the frequency of regulatory T cells and expression of FOXP3 in a patient with systemic lupus erythematosus: a two-year follow-up.

Regulatory T cells (Tregs) maintain peripheral tolerance by restricting the activity of nearby immunocytes. The loss of tolerance can induce development of autoimmune diseases, such as systemic lupus erythematosus (SLE). It has previously been shown that SLE patients have lower levels of Tregs than controls. Herein we show that glucocorticoid therapy in a newly diagnosed SLE patient was accompanied with the increase in the percentage of CD4+CD25+Foxp3 T cells to the higher levels than in healthy controls. No effect of the therapy was observed on the percentage of CD4+CD25+ T cells. The observed difference in correlation of the CD4+CD25+ and CD4+CD25+Foxp3+ T cells levels with disease onset in a patient receiving steroids emphasized the importance of critical evaluation of Treg phenotype and their role in the control of SLE.

Type I cytokine profiles of human naïve and memory B lymphocytes: a potential for memory cells to impact polarization

B cells bifurcating along ‘type 1’ or ‘type 2’ pathways under the influence of polarizing cytokines can, in turn, influence the direction of an immune response. Here, we compare the capacity of human B cells residing within naïve and memory compartments to participate in type 1 polarizing responses. B‐cell receptor (BCR) engagement provided the main signal for interleukin (IL)‐12Rβ1 expression in the two subsets: this was potentiated by CD154 together with interferon‐γ (IFN‐γ) but inhibited by IL‐12. IL‐12Rβ2 could be induced on a minority of B cells by the same signals, and also by IFN‐γ alone. WSX‐1, a receptor for IL‐27, was expressed in both subsets with no evidence for its regulation by the signals studied. While neither subset was capable of secreting much IL‐12 p70, memory B cells could produce a small amount of IL‐12 p40 on CD40 ligation. Memory B cells also, exclusively, expressed IL‐23 p19 mRNA on BCR triggering. Importantly, products of appropriately stimulated memory – but not naive – B cells were shown to promote the synthesis of IFN‐γ in uncommitted T‐helper cells. The data indicate an equal capacity for naïve and memory B cells to respond within a type 1 polarizing environment. Although poorly equipped for initiating type 1 responses, B cells – by virtue of the memory subset – reveal a capacity for their maintenance and amplification following T‐dependent signalling.

Monocyte response to LPS after exposure to corticosteroids and chloroquine with implications for systemic lupus erythematosus

Essential part of a response to infection is early pathogen recognition and adequate initiation of innate immunity. One of the hallmarks of systemic lupus erythematosus (SLE) is reduced resistance to infection despite overall hyperactivity of the immune system. Immunosuppressive drugs (high‐dose corticosteroids and cytotoxic agents) are independent risk factors for infection in SLE, with bacteria as predominant cause. To investigate whether less aggressive immunomodulatory treatment may still affect recognition and response to Gram‐negative bacteria, we measured TLR4 expression in monocytes of untreated SLE patients and patients on chloroquine and low‐dose steroid therapy and examined the drugs’ influence on monocyte TLR4 expression in peripheral blood mononuclear cell (PBMC) culture. Additionally, we determined whether induction of monocyte NF‐κB signalling, TNF‐α and IL‐6 production with lipopolysaccharide (LPS), a TLR4 ligand, can be altered with dexamethasone, chloroquine or both. There was no statistically significant difference in TLR4 expression between patients with SLE and controls, even though treated SLE patients tended to have lower frequency of TLR4+ monocytes and TLR4 mean fluorescence intensity than healthy controls. However, neither dexamethasone nor chloroquine had major influence on TLR4 expression in vitro or suppressed LPS‐induced NF‐κB activation in monocytes, although dexamethasone decreased TNF‐α and IL‐6 production. Therefore, even if low‐dose steroids or chloroquine do not seem to affect TLR4 expression and signalling, steroids might decrease cytokine production in response to LPS.

Decrease in circulating DNA, IL-10 and BAFF levels in newly-diagnosed SLE patients after corticosteroid and chloroquine treatment

Arsenal of pattern-recognition receptors alongside antibody production machinery make B cells vulnerable to autoimmune response if an autoantigen elicits both pathways in a self-sustained fashion. Systemic lupus erythematosus is an autoimmune disease characterized by autoantibodies to DNA, RNA and related structures. Murine studies demonstrated autoreactive B cell activation upon TLR9 stimulation with DNA-containing immune complexes. This activation could be abolished with chloroquine, a drug used in SLE treatment that also blocks TLR9 signaling. We investigated whether chloroquine modulates TLR9 expression, circulating DNA levels and B cell-related cytokines in newly discovered, untreated SLE patients. TLR9 was measured in peripheral blood B cells by flow cytometry, serum DNA by real-time PCR, and IL-10 and BAFF by ELISA before treatment, after 3weeks on corticosteroids, and 3months after introduction of chloroquine. We found that circulating DNA is higher in SLE patients than in controls in every time-point and decreases significantly after chloroquine treatment. Untreated patients had higher serum IL-10 than controls or patients on corticosteroids. Also, corticosteroids decreased and chloroquine completely abolished CpG-mediated CD86 upregulation on B cells and IL-10 secretion in PBMC culture. Providing the TLR9 pathway activation demonstrates its importance in pathogenesis of human SLE, this data supports continuation of chloroquine in SLE treatment protocol. In addition, observed modulation of cytokine and DNA levels after immunomodulatory treatment prompts for inclusion of untreated patients in studies of human immune disorders.

Expression of chemokine receptor CX3CR1 in infants with respiratory syncytial virus bronchiolitis

Respiratory syncytial virus (RSV) glycoprotein G mimics fractalkine, a CX3C chemokine, which mediates chemotaxis of leukocytes expressing its receptor, CX3CR1. The aim of this study was to examine the relationship between RSV infection and expression of perforin and IFN‐γ in CX3CR1‐expressing peripheral blood CD8+ T cells. Samples were collected from infants with RSV bronchiolitis, both in the acute and convalescence phase (n = 12), and from their age‐ and sex‐matched healthy controls (n = 15). Perforin expression and IFN‐γ secretion in CX3CR1+ CD8+ T cells were assessed by four‐color flow cytometry. The NF‐κB p50 and p65 subunit levels were also determined as markers of RSV‐induced inflammation. Study results showed perforin and CX3CR1 expression to be significantly lower in the convalescent phase of infected infants than in healthy controls. There was no significant difference in IFN‐γ secretion and NF‐κB binding activity between two time‐points in RSV‐infected infants, or when compared with healthy controls. Infants with prolonged wheezing had lower acute‐phase CX3CR1 levels in peripheral blood. These data indicate existence of an event persisting after acute RSV infection that is able to modulate effector functions of cytotoxic T cells, and also link disease severity with CX3CR1 expression.