Joana Vitte

Component‐resolved diagnosis with commercially available D. pteronyssinus Der p 1, Der p 2 and Der p 10: relevant markers for house dust mite allergy

Commercially available house dust mite components may improve routine testing of allergic patients.

The SNARE Machinery in Mast Cell Secretion.

Mast cells are known as inflammatory cells which exert their functions in allergic and anaphylactic reactions by secretion of numerous inflammatory mediators. During an allergic response, the high-affinity IgE receptor, FcεRI, becomes cross-linked by receptor-bound IgE and antigen resulting in immediate release of pre-synthesized mediators – stored in granules – as well as in de novo synthesis of various mediators like cytokines and chemokines. Soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptors (SNARE) proteins were found to play a central role in regulating membrane fusion events during exocytosis. In addition, several accessory regulators like Munc13, Munc18, Rab GTPases, secretory carrier membrane proteins, complexins, or synaptotagmins were found to be involved in membrane fusion. In this review we summarize our current knowledge about the SNARE machinery and its mechanism of action in mast cell secretion.

Human mast cell tryptase in biology and medicine

The most abundant prestored enzyme of human mast cell secretory granules is the serine-protease tryptase. In humans, there are four tryptase isoforms, but only two of them, namely the alpha and beta tryptases, are known as medically important. Low levels of continuous tryptase production as an immature monomer makes up the major part of the baseline serum tryptase levels, while transient release of mature tetrameric tryptase upon mast cell degranulation accounts for the anaphylactic rise of serum tryptase levels. Serum tryptase determination contributes to the diagnosis or monitoring of mast cell disorders including mast cell activation - induced anaphylaxis, mastocytosis and a number of myeloproliferative conditions with mast cell lineage involvement. Baseline serum tryptase levels are predictive of the severity risk in some allergic conditions.

Oxidative stress level in circulating neutrophils is linked to neurodegenerative diseases.

Alzheimer’s and Parkinson’s diseases are the most common neurodegenerative conditions. Oxidative lesions are a hallmark of both diseases, but the respective roles of systemic and cerebral dysfunction are not elucidated. As circulating neutrophils are the most powerful sources of reactive oxygen species, we measured oxidative stress levels in resting neutrophils from 44 Alzheimer’s and Parkinson’s disease patients and compared them to 40 healthy counterparts. Significantly increased oxidative stress levels were observed in patients’ groups, while control groups had very similar levels irrespective of age. One-third of the neurodegenerative patients presented with oxidative stress levels higher than those of any healthy donor. This increase was not due to an elevated production of reactive oxygen species during the neutrophil oxidative burst. Mitochondrial mass and activity were altered in neutrophils of the Parkinsonian group compared to controls, but not in those from Alzheimer’s disease group. To our knowledge, this is the first report linking oxidative stress and mitochondrial parameters in circulating neutrophils from neurodegenerative and normal donors. Our results indicate that oxidative stress levels in circulating neutrophils are of interest for further mechanistic studies of neurodegenerative diseases and might open the perspective of a diagnostic tool.

Munc18-2 and Syntaxin 3 Control Distinct Essential Steps in Mast Cell Degranulation

Mast cell degranulation requires N-ethylmaleimide–sensitive factor attachment protein receptors (SNARE) and mammalian uncoordinated18 (Munc18) fusion accessory proteins for membrane fusion. However, it is still unknown how their interaction supports fusion. In this study, we found that small interfering RNA–mediated silencing of the isoform Munc18-2 in mast cells inhibits cytoplasmic secretory granule (SG) release but not CCL2 chemokine secretion. Silencing of its SNARE-binding partner syntaxin 3 (STX3) also markedly inhibited degranulation, whereas combined knockdown produced an additive inhibitory effect. Strikingly, while Munc18-2 silencing impaired SG translocation, silencing of STX3 inhibited fusion, demonstrating unique roles of each protein. Immunogold studies showed that both Munc18-2 and STX3 are located on the granule surface, but also within the granule matrix and in small nocodazole-sensitive clusters of the cytoskeletal meshwork surrounding SG. After stimulation, clusters containing both effectors were detected at fusion sites. In resting cells, Munc18-2, but not STX3, interacted with tubulin. This interaction was sensitive to nocodazole treatment and decreased after stimulation. Our results indicate that Munc18-2 dynamically couples the membrane fusion machinery to the microtubule cytoskeleton and demonstrate that Munc18-2 and STX3 perform distinct, but complementary, functions to support, respectively, SG translocation and membrane fusion in mast cells.

Genetic Evidence for the Aggravation of Plasmodium falciparum Malaria by Interleukin 4

BACKGROUND Severe malaria (SM) due to Plasmodium falciparum causes millions of child deaths in sub-Saharan Africa. It comprises a variety of clinical disorders, including cerebral malaria (CM) and severe anemia (SA). In previous work, we have shown that interferon gamma and interleukin 12 protect against CM. Here, we investigated whether interleukin 4 (IL-4) aggravates the risk of severe disease. METHODS We prospectively recruited children with CM (n = 240), SA (n = 101), and uncomplicated malaria (UM) (n = 42) in Bamako, Mali, and measured IL-4 production in plasma by enzyme-linked immunosorbent assay. We then assessed the influence of 11 polymorphisms on predisposition to SM by the family-based association test (FBAT). RESULTS IL-4 concentrations were higher in children with CM than in children with UM during malaria (P = .003). FBAT analyses showed that the most significant association was between the IL4 variable-number tandem repeat (VNTR) 1/2 genotype and SM (P < .001); an association was also observed for IL4 -33 C/T, rs2243267 G/C, rs2243268 C/A, and rs2243282 C/A (P < .05). Interestingly, we found that the plasma concentration of IL-4 was higher in subjects with the IL4 VNTR 1/2 or 1/1 genotype than with the IL4 VNTR 2/2 genotype (P = .003). CONCLUSIONS These results support the view that IL-4 may be a risk factor for SM. IL-4 may aggravate the disease by interfering with type 1 T helper cell differentiation or by promoting local inflammation at sites of parasite sequestration.

Total serum tryptase levels are higher in young infants

To cite this article: Belhocine W, Ibrahim Z, Grandné V, Buffat C, Robert P, Gras D, Cleach I, Bongrand P, Carayon P, Vitte J. Total serum tryptase levels are higher in young infants. Pediatr Allergy Immunol 2011; 22: 600–607.

Ara h 2 and Ara h 6 sensitization predicts peanut allergy in Mediterranean pediatric patients

Peanut allergy (PA) management was improved by the introduction of molecular allergology, but guidelines for Mediterranean patients are lacking. We aimed at evaluating peanut component‐resolved diagnosis as a diagnostic and prognostic tool in children from Southern France.

Direct quantification of the modulation of interaction between cell- or surface-bound LFA-1 and ICAM-1

The functional activity of leukocyte integrins is highly regulated by several mechanisms related to intrinsic molecular properties and receptor interaction with the cell membrane. Here, we present a microkinetic study of the lymphocyte function‐associated antigen‐1‐mediated interaction between flowing Jurkat cells and surface‐ or cell‐bound intercellular adhesion molecule‐1 (ICAM‐1). We conclude that adhesion is initiated by the formation of a single bond with ∼0.3 s–1 dissociation rate, and attachment is subsequently strengthened by the formation of additional bonds during the next 10 s; exposing cells to Mg2+ or Mn2+ resulted in up to a 16‐fold increase of the binding frequency, in line with reported measurements performed on isolated molecules with surface plasmon resonance methodology; cell‐bound ICAM‐1 molecules were more efficient in mediating adhesion than Fc‐ICAM‐1, properly oriented and bound by surface‐adsorbed protein A; and quantitative analysis of binding frequency suggested that adhesion efficiency was ten‐ to 100‐fold lower than the maximum value allowed by previously determined association rates of soluble molecules. It is concluded that the presented methodology provides a simple and unique way of dissecting the initial step of cell adhesion and discriminating between affinity and avidity modulation of adhesion receptors.

Adenosine and Hemodialysis in Humans

Abstract Background Infections and hypotension are serious complications that develop during hemodialysis (HD) treatment. Adenosine (ADO), a strong hypotensive and immunosuppressive agent, may participate in these two HD complications, because high concentrations of ADO metabolites are found in dialyzed human plasma. ADO, which is released by endothelial cells, is quickly transformed into inosine (INO) by plasmatic ADO deaminase (ADA) and mononuclear cell ADO deaminase (MCADA). In plasma, the degradation of ADO into INO and its uptake by red blood cells (RBC) are both very rapid, resulting in the short half-life of ADO in blood. Methods Using liquid chromatography, we evaluated ADO and INO plasma concentrations before and after HD session. Results Before the HD session, ADO and INO plasma concentrations were higher in hemodialyzed patients than in controls and in peritoneally dialyzed patients. At the end of the HD session, ADO plasma concentration was increased. ADO plasma concentration for the undialyzed patients was in the same range as that of the controls. Before HD, ADA activity was higher in hemodialyzed patients (559±349 IU) than in controls (219±48 IU), and the activity rose during the session (665±135 IU). ADA activity in the undialyzed patients (222±80 IU) was in the same range as that of the controls (219±48 IU). Before the HD session, the MCADA activity (247±144 IU) was lower than in controls (624±99 IU). HD did not modify ADO RBC uptake. ADO inhibited mononuclear cell proliferation and interferon-γ production in humans. Finally, as much as 50 μM INO does not inhibit ADO uptake by RBC and does not modify ADA and MCADA activities. Conclusions These data indicate that chronic HD inhibited MCADA activity and increased ADO plasma concentration. Both high ADO plasma concentration and low MCADA activity may be involved in dialysis-induced immune system failure and thereby favor infectious diseases.