Spleen Tyrosine Kinase (Syk) Regulates Systemic Lupus Erythematosus (SLE) T Cell Signaling

Engagement of the CD3/T cell receptor complex in systemic lupus erythematosus (SLE) T cells involves Syk rather than the zeta-associated protein. Because Syk is being considered as a therapeutic target we asked whether Syk is central to the multiple aberrantly modulated molecules in SLE T cells. Using a gene expression array, we demonstrate that forced expression of Syk in normal T cells reproduces most of the aberrantly expressed molecules whereas silencing of Syk in SLE T cells normalizes the expression of most abnormally expressed molecules. Protein along with gene expression modulation for select molecules was confirmed. Specifically, levels of cytokine IL-21, cell surface receptor CD44, and intracellular molecules PP2A and OAS2 increased following Syk overexpression in normal T cells and decreased after Syk silencing in SLE T cells. Our results demonstrate that levels of Syk affect the expression of a number of enzymes, cytokines and receptors that play a key role in the development of disease pathogenesis in SLE and provide support for therapeutic targeting in SLE patients

Therapeutic options for Burkholderia cepacia infections beyond co-trimoxazole: a systematic review of the clinical evidence

Burkholderia cepacia complex (BCC) is an important group of pathogens affecting patients with cystic fibrosis and chronic granulomatous disease as well as immunocompromised and hospitalised patients. Therapeutic options are limited owing to high levels of resistance of the organism, either intrinsic or acquired, to many antimicrobial agents. Co-trimoxazole (trimethoprim/sulfamethoxazole) has been a drug of choice. However, in some cases it cannot be administered because of allergic or hypersensitivity reactions, intolerance or resistance. We systematically searched for relevant publications including clinical data in PubMed and Scopus. The search identified 48 relevant case reports (57 cases) and 8 cohort studies or trials. Nineteen (33.3%) of 57 patients included in the case reports received ceftazidime-based regimens, 14 (73.7%) of whom were cured. Meropenem was administered in seven patients (12.3%), one (14.3%) of whom improved and five (71.4%) were cured. Seven (12.3%) of 57 cases were treated with penicillins, four of which were piperacillin (all had a favourable outcome). Based on the data reported in the eight relevant cohort studies or trials identified, favourable outcomes were observed in 68.4% (26/38) to 100% (16/16) of cases treated with ceftazidime and 66.7% (6/9) of cases treated with meropenem. Also, 9/12 (75%) of patients receiving penicillins improved. Thus, Ceftazidime, meropenem and penicillins, mainly piperacillin, either alone or in combination with other antimicrobial agents, may be considered as alternative options for BCC infections, according to the in vitro antimicrobial susceptibility patterns and clinical results. However, the available clinical data are not sufficient and further clinical experience is required to clarify the appropriateness of these antibiotics for BCC infections. (C) 2008 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved

Spleen Tyrosine Kinase (Syk) Regulates Systemic Lupus Erythematosus (SLE) T Cell Signaling

Engagement of the CD3/T cell receptor complex in systemic lupus erythematosus (SLE) T cells involves Syk rather than the zeta-associated protein. Because Syk is being considered as a therapeutic target we asked whether Syk is central to the multiple aberrantly modulated molecules in SLE T cells. Using a gene expression array, we demonstrate that forced expression of Syk in normal T cells reproduces most of the aberrantly expressed molecules whereas silencing of Syk in SLE T cells normalizes the expression of most abnormally expressed molecules. Protein along with gene expression modulation for select molecules was confirmed. Specifically, levels of cytokine IL-21, cell surface receptor CD44, and intracellular molecules PP2A and OAS2 increased following Syk overexpression in normal T cells and decreased after Syk silencing in SLE T cells. Our results demonstrate that levels of Syk affect the expression of a number of enzymes, cytokines and receptors that play a key role in the development of disease pathogenesis in SLE and provide support for therapeutic targeting in SLE patients

Schematic representation of the proposed role of Syk in SLE.

<p>Syk promotes the upregulation of a number of cytokines, receptors and enzymes that play a key role in SLE immunopathogenesis. It affects expression levels of CD44, primarily variant v6, involved in T cell migration; IL-21, involved in antibody production; OAS2, involved in type-I interferon responses; and PP2A, involved in the regulation of IL-2 production.</p

Intracellular levels of Syk affect the expression of genes associated with SLE immunopathogenesis.

<p>a) Healthy blood donor T cells were transfected with a SYK overexpression plasmid and the expression of an array of genes associated with SLE immunopathogenesis was measured using real-time PCR. A number of genes known to be overexpressed in SLE patients were found to be upregulated by SYK overexpression, like IL-21, CD44, OAS2 and PP2A (mean ±SEM fold expression changes between overexpression and empty vector transfected cells in four different experiments are shown). b) SLE patient T cells were transfected with either a SYK-specific siRNA or a control, non-silencing, siRNA. Silencing of SYK resulted in suppression of expression of a number of genes known to be aberrantly upregulated in SLE. Notably, expression levels of IL-21, CD44, OAS2 and PP2A were all found to decrease in SYK-knockdown T cells (mean ±SEM fold expression changes between silencing and control transfected T cells in four different experiments are shown).</p

Induced Syk expression and Syk silencing in T cells.

<p>a) T cells extracted from healthy blood-donors were transfected with either a SYK (SYK) or an empty (EV) expression vector. Following 72h of incubation, cells were lysed, RNA extracted and SYK expression levels measured in real-time PCR. SYK overexpression led to a significant increase in its mRNA levels in all four experiments tested (left panel, normalized expression levels shown). To investigate whether this finding translates into protein expression levels as well, whole T cells were used to be analyzed in flow cytometry. Syk protein was found to significantly increase following SYK overexpression in all experiments tested (right panel, a representative plot is shown, plots gated on CD3+ T cells). b) T cells extracted from SLE patients were transfected with either a SYK-specific (siRNA) or a control (control) siRNA and expression levels of Syk were measured using real-time PCR and flow cytometry. The SYK silencing protocol led to a significant reduction in its expression at both the mRNA (left panel) and protein levels (right panel).</p

Silencing of SYK results in the suppression of aberrantly expressed molecules in SLE.

<p>a) SLE patient T cells were transfected with 15nM of either control or SYK-specific siRNA. Seventy-two hours following transfection cells were harvested and stained with antibodies against IL-21, CD44v3 and CD44v6. Silencing of SYK resulted in the suppression of expression of all the above molecules, particularly IL-21 and CD44v6. Bars show the mean ±SEM of control (control) vs. SYK-siRNA (siRNA) transfected T cells in ≥7 experiments and a representative experiment is shown. b) Protein expression levels of OAS2 and PP2a measured in western blot were found to decrease following silencing of SYK. Mean ±SEM of densitometry analyses of SYK-specific (siRNA) vs. control (control) siRNA transfected cells in ≥6 experiments are plotted. A representative experiment and cumulative data are shown.</p