Background Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by diverse pathogenic autoantibodies and clinical phenotype. Laboratory monitoring of complement activation products (CAP; C3a, C5a, C3d, C4d) is commonly used in diagnosis and follow up. CAP bound to cellular surfaces (CB-CAP) analysed by flow cytometry offer a relatively novel approach to study complement turnover in SLE. Objectives We evaluated the significance and usability of cell bound CAP in monitoring complement turnover in a whole blood staining approach, comparing cell bound CAPs on different leucocyte subpopulations with conventional complement and disease activity parameters. Methods CAP bound to lymphocyte subpopulations were analysed by flow cytometry, compared to serological complement and other SLE activity parameters determined by nephelometry, rocket electrophoresis and ELISA. Results We have analysed CB-CAPs bound to lymphocytes in 50 adult patients with SLE. B-cells and CD4+ T-cells of SLE patients showed significantly higher levels of cell-surface-bound C4d compared to controls. C4d bound to B-lymphocytes was inversely correlated with serum levels of C4 (r = -0.309, p=0.013) and C3 (r= - 0.356, p=0.004) reflecting complement decay. In addition, B-cell C4d showed a significant positive correlation with anti-dsDNA antibody levels (r =0.574, p<0.001) and plasma C3d (r =0.319, p=0.009) thereby corresponding to disease activity. CB-CAPs were stable over 24 hours after blood drawing thus offering more flexibility in sample handling than conventional complement turnover assays. Conclusions Our data show that CB-CAP measurement correlates with established disease activity markers, and given its easier handling, is of possible utility in routine diagnostic and monitoring of patients with SLE. A multicentre study for evaluation of this method in a cohort of paediatric patients is planned. Disclosure of Interest None declared

In children with primary immunodeficiencies, the onset of symptoms precedes the diagnosis and the initiation of appropriate treatment by months or years. This delay in diagnosis is due to the fact that while these disorders are rare, some of the infections seen in immunodeficient patients are common. Defective antibody production represents the largest group among these disorders, with otitis, sinusitis and pneumonia as the most frequent initial manifestation. We performed a prospective study of humoral immunity in children hospitalized due to community-acquired pneumonia in tertiary care hospital. Out of 254 patients (131 boys, 123 girls, median age 4.5 years) recruited over 3 years, we found 2 boys (age 11 and 21 months) lacking serum immunoglobulins and circulating B cells. Subsequent genetic analysis confirmed diagnosis of X-linked agammaglobulinemia. Despite their immunodeficiency, the pneumonia was uncomplicated in both patients and did not call for immunological evaluation. However, the immunoglobulin screening at admission allowed for an early diagnosis of the immunodeficiency and timely initiation of immunoglobulin substitution, the key prerequisite for a favorable course of the disease.Simple and inexpensive immuno-globulin measurement during the manage-ment of hospitalized children with community-acquired pneumonia may help in early identification of patients with compromised humoral immunity and prevent serious complications.

BACKGROUNDSevere combined immunodeficiency (SCID) comprises a heterogeneous group of heritable deficiencies of humoral and cell-mediated immunity. Many patients with SCID have lymphocyte-activation defects that remain uncharacterized.METHODSWe performed genetic studies in four patients, from four families of Northern Cree ancestry, who had clinical characteristics of SCID, including early onset of severe viral, bacterial, and fungal infections despite normal B-cell and T-cell counts. Genomewide homozygosity mapping was used to identify a candidate region, which was found on chromosome 8; all genes within this interval were sequenced. Immune-cell populations, signal transduction on activation, and effector functions were studied.RESULTSThe patients had hypogammaglobulinemia or agammaglobulinemia, and their peripheral-blood B cells and T cells were almost exclusively of naive phenotype. Regulatory T cells and γδ T cells were absent. All patients carried a homozygous duplication--c.1292dupG in exon 13 of IKBKB, which encodes IκB kinase 2 (IKK2, also known as IKKβ)--leading to loss of expression of IKK2, a component of the IKK-nuclear factor κB (NF-κB) pathway. Immune cells from the patients had impaired responses to stimulation through T-cell receptors, B-cell receptors, toll-like receptors, inflammatory cytokine receptors, and mitogens.CONCLUSIONSA form of human SCID is characterized by normal lymphocyte development despite a loss of IKK2 function. IKK2 deficiency results in an impaired response to activation stimuli in a variety of immune cells, leading to clinically relevant impairment of adaptive and innate immunity. Although Ikk2 deficiency is lethal in mouse embryos, our observations suggest a more restricted, unique role of IKK2-NF-κB signaling in humans. (Funded by the German Federal Ministry of Education and Research and others.).

To the Editor: Autoimmune cytopenia and chronic lymphoproliferation are the key manifestations of autoimmune lymphoproliferative syndrome (ALPS), a disorder of lymphocyte apoptosis associated with mutations in the CD95 pathway [1]. Impaired CD95 function and changes in B cell selection allowing the survival of autoreactive B cells may contribute to autoimmunity in ALPS [2, 3]. B cell–activating factor (BAFF) is an important regulator of B cell survival [4, 5]. In mice, overexpression of BAFF results in autoimmunity [6], and elevated BAFF levels have been reported in autoimmune diseases [4, 7, 8] as well as in primary immunodeficiencies associated with autoimmunity [9, 10]. In ALPS, the contribution of BAFF to autoimmune cytopenia has not been investigated. We determined serum BAFF levels in 115 patients with lymphoproliferation (splenomegaly and ⁄ or persistent enlargement of lymph nodes in at least two sites) and autoimmune cytopenia referred for the evaluation of ALPS. Twenty-five healthy donors served to determine normal BAFF serum levels by ELISA (Adipogen ⁄ Axxora, N. AG-46B-0001-KI01), which were in the range reported by others [8]. Twenty-two patients carried germline mutations in CD95 (ALPS-FAS), and 19 of them had BAFF levels above the 95th percentile of controls (Fig. 1). Median BAFF levels were 3.5-fold higher than in healthy donors. BAFF levels were similarly elevated in six patients with increased double-negative T (DNT) cells, abnormal CD95-mediated apoptosis, but no germline or somatic mutation in CD95, CD95L or caspase 10 (ALPS-U), in 55 patients with elevated DNT cells, but normal CD95 mediated apoptosis (ALPS-ph) and in 31 patients with normal DNT cells and normal apoptosis (Others). Among the 22 ALPS-FAS patients, BAFF levels did not correlate with the number of lineages affected by autoimmune cytopenia, the extent of lymphoproliferation or with biomarkers including IgG, IL-10, sCD95L, vitamin B12 or the percentage of DNT cells. However, in the whole cohort, there was a significant correlation between BAFF levels and the percentage of B cells (n = 105, rs = )0.491, P < 0.01) and sCD25 levels (n = 113, rs = 0.287, P < 0.01). It is well known that BAFF is upregulated in response to B cell lymphopenia and median BAFF levels were 20-fold higher in two patients with X-linked agammaglobulinemia (XLA) than in our patients with lymphoproliferation and autoimmunity. BAFF is produced by myeloid cells, and sCD25 is a marker of T cell and monocyte activation, both hallmarks of hemophagocytic lymphohistiocytosis (HLH). Analysis of BAFF levels in five patients with HLH revealed levels even higher than those in XLA patients. In summary, patients with ALPS-FAS and other forms of lymphoproliferation and autoimmunity showed elevated BAFF levels similar to patients with systemic lupus erythematosus or Sjögren Syndrome [8]. It remains unclear whether these levels are relevant for the pathogenesis of autoimmunity. The fact that 50-fold higher levels were observed in HLH, a disease not associated with autoantibodies, argues against a simple correlation. Elevated BAFF levels in patients with lymphoproliferation and autoimmunity including ALPS-FAS may rather reflect monocyte activation or decreased B cell numbers, which can influence BAFF levels, than represent a causal factor for the autoimmune cytopenia.

Autoimmune lymphoproliferative syndrome (ALPS) is mainly caused by defects in the CD95 pathway. Raised CD3+TCRαβ+CD4-CD8- double negative T cells and impaired T cell apoptosis are hallmarks of the disease. In contrast, the B cell compartment has been less well studied. We found an altered distribution of B cell subsets with raised transitional B cells and reduced marginal zone B cells, switched memory B cells and plasma blasts in most of 22 analyzed ALPS patients. Moreover, 5 out of 66 ALPS patients presented with low IgG and susceptibility to infection revealing a significant overlap between ALPS and common variable immunodeficiency (CVID). In patients presenting with lymphoproliferation, cytopenia, hypogammaglobulinemia and impaired B cell differentiation, serum biomarkers were helpful in addition to apoptosis tests for the identification of ALPS patients. Our observations may indicate a role for apoptosis defects in some diseases currently classified as CVID.