Search

To evaluate the outcome of children who received prolonged intravenous immunoglobulin (IVIg) replacement therapy early in life for X-linked agammaglobulinemia (XLA).We performed a retrospective study of the clinical features and outcome of patients with genetic and/or immunologic results consistent with XLA. Patients receiving IVIg replacement therapy within 3 months of the diagnosis and for at least 4 years between 1982 and 1997 were included.Thirty-one patients began receiving IVIg replacement therapy at a median age of 24 months and were followed up for a median time of 123 months. IVIg was given at doses0.25 g/kg every 3 weeks, and mean individual residual IgG levels ranged from 500 to 1140 mg/dL (median, 700 mg/dL). During IVIg replacement, the incidence of bacterial infections requiring hospitalization fell from 0.40 to 0.06 per patient per year (P. 001). However, viral or unidentified infections still developed, including enteroviral meningoencephalitis (n = 3) causing death in one patient, exudative enteropathy (n = 3), and aseptic arthritis (n = 1). At last follow-up, 30 patients were alive at a median age of 144 months (range, 58 to 253 months). Among 23 patients who were evaluated by respiratory function tests and computed tomography, 3 had an obstructive syndrome, 6 had bronchiectasis, and 20 had chronic sinusitis.Early IVIg replacement therapy achieving residual IgG levels500 mg/dL is effective in preventing severe acute bacterial infections and pulmonary insufficiency. More intensive therapy may be required to fully prevent the onset of bronchiectasis, chronic sinusitis, and nonbacterial infections, particularly enteroviral infections, in all cases.

The class II transactivator (CIITA) controls both the constitutive and IFN-gamma inducible expression of HLA-D genes. In addition, through the squelching of another transactivator CREB-binding protein, CIITA was more recently shown to have a wider cellular function, including cell cycle control or cellular response to IFN-gamma and IL-4. However, due to its low expression level, its analysis mainly relies on the study of recombinant overexpressed forms of the protein. We report here the analysis of native CIITA in various cell types. We first show the precise timing of CIITA protein expression in a fibroblast cell line in response to IFN-gamma. This expression is observed 2 h after the cytokine addition with a peak of expression ranging from 16 to 24 h. We next show the existence of two major isoforms of the CIITA protein differentially expressed in fibroblast, B lymphocyte or melanoma cell lines. We present the first demonstration that these isoforms originate from alternative translation initiation codons. We finally show that CIITA isoforms translocate to the nucleus with an apparently similar efficiency. Our data therefore demonstrate the existence of CIITA isoforms whose respective ratio depends on the cell type examined. However, we present evidence for a modulation of this ratio in a melanoma cell line with an abnormal constitutive expression of MHC class II molecules.

International audience; Griscelli syndrome (GS) is a rare autosomal recessive disorder that associates hypopigmentation, characterized by a silver-gray sheen of the hair and the presence of large clusters of pigment in the hair shaft, and the occurrence of either a primary neurological impairment or a severe immune disorder. Two different genetic forms, GS1 and GS2, respectively, account for the mutually exclusive neurological and immunological phenotypes. Mutations in the gene encoding the molecular motor protein Myosin Va (MyoVa) cause GS1 and the dilute mutant in mice, whereas mutations in the gene encoding the small GTPase Rab27a are responsible for GS2 and the ashen mouse model. We herein present genetic and functional evidence that a third form of GS (GS3), whose expression is restricted to the characteristic hypopigmentation of GS, results from mutation in the gene that encodes melanophilin (Mlph), the ortholog of the gene mutated in leaden mice. We also show that an identical phenotype can result from the deletion of the MYO5A F-exon, an exon with a tissue-restricted expression pattern. This spectrum of GS conditions pinpoints the distinct molecular pathways used by melanocytes, neurons, and immune cells in secretory granule exocytosis, which in part remain to be unraveled.

We retrospectively analyzed the B-cell function and leukocyte chimerism of 22 patients with severe combined immunodeficiency with B cells (B+ SCID) who survived more than 2 years after bone marrow transplantation (BMT) to determine the possible consequences of BMT procedures, leukocyte chimerism, and SCID molecular deficit on B-cell function outcome. Circulating T cells were of donor origin in all patients. In recipients of HLA-identical BMT (n = 5), monocytes were of host origin in 5 and B cells were of host origin in 4 and of mixed origin in 1. In recipients of HLA haploidentical T-cell–depleted BMT (n = 17), B cells and monocytes were of host origin in 14 and of donor origin in 3. Engraftment of B cells was found to be associated with normal B-cell function. In contrast, 10 of 18 patients with host B cells still require Ig substitution. Conditioning regimen (ie, 8 mg/kg busulfan and 200 mg/kg cyclophosphamide) was shown neither to promote B-cell and monocyte engraftment nor to affect B-cell function. Eight patients with B cells of host origin had normal B-cell function. Evidence for functional host B cells was further provided in 3 informative cases by Ig allotype determination and by the detection, in 5 studied cases, of host CD27+ memory B cells as in age-matched controls. These results strongly suggest that, in some transplanted patients, host B cells can cooperate with donor T cells to fully mature in Ig-producing cells.

The endothelial dysfunction associated with Hematopoietic Stem Cell Transplantation (HSCT) is responsible for pleomorphic complications that arise during the post-tranplant period: sinusoidal obstruction syndrome (SOS), pulmonary arterial hypertension (PAH), capillary leak syndrome (CLS) and transplantation-associated thrombotic microangiopathy (TA-TMA). Numerous factors may induce these endothelial damages. Circulating endothelial cells (CEC) emerge as a potent marker of endothelial lesion in a variety of disorders but their monitoring in HSCT has been little investigated so far. Methods Thirty-six pediatric patients treated in the department of Immuno-Hematology at the Necker-Enfants-Malades Hospital who received allogeneic HSCT after full intensity conditioning regimen for non-malignant disorders including inherited immunodeficiencies, metabolic diseases or hemoglobin disorders were enrolled in the study between April 2011 and December 2012. CEC were monitored at 5 time points: before conditioning (day -15), at time of transplant (day 0) and during the post-transplantation period (at day +15, day +30 and eventually day +60 if the patient was still hospitalized). Blood samples were not collected in period of sepsis to avoid any confounding factor7. Results Among the thirty-six patient enrolled, four patients were excluded because of incomplete follow-up. Thirty-two patients completed the study, and their data were analyzed. Vascular complications occurred in 10 patients (31% patients) and are detailed in Figure 1C. Of note, all patients who developed SOS were under defibrotide prophylaxis. Two patients developed CLS in association with grade IV GVH. TA-TMA occurred in two patients and resolved in both after cyclosporine withdrawal. Patients with vascular events (VE) had significant higher CEC counts as compared to patients without endothelial complication at day +15 (p = 0.0122) and day +30 (p=0.0046). The increase of CEC counts was significantly higher in patients with VE during the first 15 days (p=0.0175) and during the first month post transplantation (p=0.0005) as compared to patient without VE. Strikingly, detection of high CEC counts in patients with major vascular events (ie. PAH, CLS and TA-TAM) was preceding the onset of clinical manifestations by a median of 18.5 days (range, 8-42). We didn’t find any correlation on univariate analysis between CEC counts and the following variables: the underlying disease, the intensity of conditioning regimen, the existence of CMV reactivation, graft versus host disease or death. Conclusion We observed that the occurrence of vascular complications is associated with an early rise (in the first 15 days) of CEC count. Interestingly, the increase in the number of CEC is preceding the onset of major endothelial complications. Thus, monitoring of CEC in routine could be a useful prognostic tool that could allow early therapeutic intervention. Disclosures: No relevant conflicts of interest to declare.

> Le deficit immunitaire combine severe (DICS) est une maladie rare, caracterisee par l’absence de lymphocytes T, associee, dans certains cas, a l’absence de cellules NK (natural killer) et/ou de lymphocytes B. Le deficit immunitaire peut etre corrige par une transplantation de cellules souches hematopoietiques (TCSH) allogenique [1, 2] ou par therapie genique [3]. L’observation, apres greffe, d’une maladie cutanee grave et chronique liee a une infection par papillomavirus humains (HPV, human papillomavirus) chez certains patients greffes pour DICS pose un probleme intrigant. Les HPV ont un tropisme specifique pour les keratinocytes. Ils sont la cause des verrues planes ou vulgaires, des condylomes genitaux et des papillomes larynges. Habituellement, les lesions regressent spontanement ou apres traitement. Une susceptibilite genetique elective au developpement d’une maladie chronique et severe a HPV a ete mise en evidence chez des patients qui developpent un tableau clinique particulier, l’epidermodysplasie verruciforme (EV) (Figure 1). Ce syndrome grave, de transmission autosomique recessive, est genetiquement heterogene. Il est caracterise par la survenue de verrues disseminees, associees a des lesions mimant le pityriasis versicolor. La progression vers la malignite se produit dans 30% a 50% des cas [4]. Deux locus de susceptibilite ont ete decrits sur le chromosome 2p21-p24 et sur le chromosome 17q25 et des mutations ont ete recemment identifiees dans deux genes adjacents, EVER1 et EVER2 [5]. Nous avons observe une incidence elevee (9/41) de maladie a HPV chez des patients greffes pour traiter un DICS [6]. Chez ces patients, la maladie a HPV apparait entre 3 et 15 ans apres la TCSH (mediane: 8 ans), en l’absence de traitement immunosuppresseur (Figure 2). Elle est severe, progressive et refractaire aux therapeutiques usuelles. En outre, on ne peut exclure le developpement de cette maladie severe a HPV chez d’autres patients dans les annees a venir. Cette maladie a HPV provoque non seulement un prejudice esthetique, mais aussi un handicap fonctionnel important. De plus, les lesions mimant l’EV, associees a HPV-5 ou HPV-14, telles qu’elles sont observees chez quatre patients, pourraient evoluer vers la malignite. Le traitement de cette maladie est decevant, inefficace ou suivi de recidives. Neanmoins, on a observe une remission complete chez deux des neuf patients apres une seconde TCSH chez un patient ou un traitement local chez le second. Nous avons donc recherche quel(s) pourrai(en)t etre le(s) facteur(s) de risque de survenue de maladie grave a HPV dans ce groupe de patients. Le seul facteur de risque identifie est le type moleculaire a l’origine du DICS. Seuls les patients presentant un DICS avec lymphocytes B presents et cellules NK absentes (DICS B+ NK-) developpent une maladie a HPV. Les caracteristiques de la TCSH (compatibilite entre le donneur et le receveur, type de conditionnement) et les evenements secondaires a la TCSH (reaction du greffon contre l’hote, infections) ne semblent pas intervenir sur le risque de survenue de la maladie a HPV. Par ailleurs, a distance de la greffe, ni le type de chimerisme ni la qualite de la reconstitution des immunites T et B ne different chez les NOUVELLE

Une réponse immune exagérée, incontrôlée et le plus souvent fatale, connue sous le nom de syndrome hémophagocytaire (SH), est associée à un défaut de la fonction cytotoxique des lymphocytes T et natural killer (NK). Les anomalies moléculaires responsables, qui sont multiples, mettent en cause dans la plupart des cas un effecteur indispensable au fonctionnement de la machinerie lytique des lymphocytes. L’étude des lymphocytes cytotoxiques déficients en l’un ou l’autre de ces effecteurs apporte des éléments nouveaux quant à l’agencement des étapes clés de la sécrétion du contenu des granules lytiques au contact de la cellule cible. Des mécanismes moléculaires proches semblent contrôler la sécrétion vésiculaire au niveau des synapses immunologique et neurologique. D’autres effecteurs de la cytotoxicité ou du contrôle de l’homéostasie lymphocytaire à l’origine de SH doivent encore être caractérisés. Quant aux mécanismes précis de l’intervention de cette voie cytotoxique dans le maintien de l’homéostasie lymphocytaire (terminaison d’une réponse immune), ils demeurent à élucider., An in vivo disturbance of lymphocyte homeostasis occurs during the course of the hemophagocytic syndrome (HS). HS is a severe and often fatal syndrome resulting from potent and uncontrolled activation and proliferation of T-lymphocytes, mainly polyclonal CD8 lymphocytes, leading to excessive macrophage activation, high level of proinflammatory cytokine production and multiple deleterious effects. The onset of HS characterizes several inherited disorders in humans. In most of these conditions, the molecular defect impairs the granule-dependent cytotoxic activity of lymphocytes, thus highlighting the determinant role of this function in driving back the immune system to a state of equilibrium following infection. Several lines of evidence suggest that an increase in the expansion phase rather than a decrease in the contraction phase of the CD8+ T cells population characterizes the HS. Failure to kill antigen presenting cells through a transaction mechanism of cytotoxic cells should favor a sustained response, although the mechanism may be more complex than simple decrease of antigen load. Defect in the granule dependent cytotoxic function of lymphocytes result from perforin mutation in familial hemophagocytic lymphohistiocytosis type 2, from Munc13-4 (UNC13D) mutation in familial hemophagocytic lymphohistiocytosis type 3, from Rab27a mutation in Griscelli syndrome type 2, and from CHS/LYST mutation in Chediak-Higashi syndrome. The characterization of the molecular causes leading to these conditions identified Rab27a and Munc13-4 as two critical effectors of the exocytic machinery, required for the terminal transport/docking or priming of the cytotoxic granules, respectively. Different members of the Rab and Munc13 family of proteins are also used in neurotransmitter release at the neurological synapse, highlighting the similarity of the mechanisms regulating both secretory pathways. Future investigations regarding HS will continue to elucidate this exocytic pathway machinery and improve our understanding of how it finely regulates the immune response, an area that is likely to be useful for therapeutic intervention.

The cytokines controlling the development of human interleukin (IL) 17--producing T helper cells in vitro have been difficult to identify. We addressed the question of the development of human IL-17--producing T helper cells in vivo by quantifying the production and secretion of IL-17 by fresh T cells ex vivo, and by T cell blasts expanded in vitro from patients with particular genetic traits affecting transforming growth factor (TGF) beta, IL-1, IL-6, or IL-23 responses. Activating mutations in TGFB1, TGFBR1, and TGFBR2 (Camurati-Engelmann disease and Marfan-like syndromes) and loss-of-function mutations in IRAK4 and MYD88 (Mendelian predisposition to pyogenic bacterial infections) had no detectable impact. In contrast, dominant-negative mutations in STAT3 (autosomal-dominant hyperimmunoglobulin E syndrome) and, to a lesser extent, null mutations in IL12B and IL12RB1 (Mendelian susceptibility to mycobacterial diseases) impaired the development of IL-17--producing T cells. These data suggest that IL-12Rbeta1- and STAT-3--dependent signals play a key role in the differentiation and/or expansion of human IL-17-producing T cell populations in vivo. The Laboratory of Human Genetics of Infectious Diseases is supported by the Agence Nationale de la Recherche, the Programme Hospitalier de Recherche Clinique, the European Union (grant LHSP-CT-2005-018736), the BNP Paribas Foundation, the March of Dimes, the Dana Foundation, and the Candi’Oser Association. L. de Beaucoudrey is supported by the Fondation pour la Recherche Medicale as part of the PhD program of Pierre et Marie Curie University. J.L. Casanova is an International Scholar of the Howard Hughes Medical Institute.

V(D)J recombination, accountable for the diversity of T cell receptor- and immunoglobulin-encoding genes, is initiated by a lymphoid-specific DNA double-strand break. The general DNA repair machinery is responsible for the resolution of this break. Any defect in one of the known components of the DNA repair/V(D)J recombination machinery (Ku70, Ku80, DNA-PKcs, XRCC4 and DNA ligase IV) leads to abortion of the V(D)J rearrangement process, early block in both T and B cell maturation, and ultimately to severe combined immune deficiency (SCID) in several animal models. A human SCID condition is also characterized by an absence of mature T and B lymphocytes, and is associated with an increase in sensitivity to DNA-damaging agents (RS-SCID). None of the above-mentioned genes are defective in these patients, arguing for the likelihood of the existence of yet another unknown component of the V(D)J recombination/DNA repair apparatus. Athabascan-speaking (SCIDA) Navajo and Apache Native Americans have a very high incidence of T – B – SCID. The SCIDA locus is highly linked with markers on chromosome 10p, although the exact molecular defect has not been recognized in these patients. We show here that cells with the SCIDA defect are impaired in the DNA repair phase of V(D)J recombination similarly to RS-SCID, precisely an absence of V(D)J coding joint formation. Moreover, genotyping analysis in several RS-SCID families corroborates a linkage of the RS-SCID locus to the SCIDA region on chromosome 10p. These results demonstrate the presence of a new essential DNA repair/V(D)J recombination gene in this region, the mutation of which causes RS-SCID in humans.

The group of immune disorders which leads to the occurrence of hemophagocytic lymphohistiocytosis (HLH) syndrome presents a strange paradox in that patients with these conditions associate a dramatic immune response to infection with the failure to establish an effective immune response. During the last few years, significant progress was made in the characterization and the understanding of the molecular basis involved in these inherited immune disorders. The hemophagocytic lymphohistiocytosis syndrome which characterized the evolution of the Chediak-Higashi syndrome and the Griscelli disease results from defects affecting intracellular trafficking. A defective SH2 protein interacting with T lymphocyte intracellular signaling pathways is the cause of the X-linked lymphoproliferative disease, whereas at least three distinct genetic defects can lead to the familial hemophagocytic lymphohistiocytosis. The molecular characterization of these latter defects is in progress. This review summarizes the recent advances as well as their implications in the diagnosis and the understanding of the physiopathology of these disorders.

Multiple carboxylase deficiency could be associated with defects of the immunoregulatory system. A boy with documented multiple carboxylase deficiency was found to display a fatty acid and biotin-responsive impairment of his lymphocyte suppressive activityin vitro. Furthermore, prostaglandin E2 (PGE2), a major product of unsaturated fatty acid elongation required forin vitro activation of the immunoregulatory system, was found to be very low in the patient's monocytes. Finally, PGE2 monocytic production responded well to biotin therapy. PGE2 deficiency could result from the combination of an essential fatty acid deficiency with an impaired elongation of unsaturated fatty acids possibly due to a defect of the biotin-dependent acetyl-CoA carboxylase reaction.

We showed that an excess of autologous or allogeneic monocytes, isolated from adult blood, added to mitogen or antigen-induced lymphocyte cultures inhibited their proliferative responses. Similarly, adult monocytes reduced the generation of Pokeweed mitogen-induced plasma cells. These suppressive effects were, both, mostly reverted by adjunction of Indomethacin (5.10−7M). In contrast, newborn (NB) monocytes did not exert any effect on proliferation and generation of plasma cells. Adherent NB monocytes were shown to secrete 10 times less prostaglandin E2 (PGE2) during a 24hrs incubation than adult monocytes. (2.300 vs 24 000 pg/m/106 monocytes). After activation by zymosan, NB monocytes secreted 6 to 10 times more PGE2. The same effect was obtained by incubation in supernatant of allogeneic leukocyte cultures and NB monocytes became able to suppress T and B in vitro functions. Also monocytes from late pregnant women did not suppress T lymphocyte proliferation. Our observations suggest that absence of suppressive effect of NB monocytes is due to an inhibition of PGE2 secretion rather than to an intrinsic immaturity. This phenomenon, which seems correlated with an excess of suppressor activity exerted by NB T lymphocytes on B lymphocyte maturation might play a role on immune responses in neonatal period.

GROWTH AND ENDOCRINE FUNCTION AFTER BONE MARROW TRANSPLANTATION (BMT) WITH OR WITHOUT TOTAL BODY IRRADIATION (TBI)

A 9 month-old boy, the fourth and the only surviving child of consanguineous parents of tunisian origin, was investigated for severe immunodeficiency associated with autoimmune anemia and thrombocytopenia. Increased serum immunoglobulin levels contrasted with an absence of antibody response after sensitization and blood T lymphocytes increased in number were incapable of proliferating normally in the presence of mitogens, antigens or of OKT3 antibody. Defective secretion of IL2 and of IFNrby patient's leukocytes was observed whatever the inducer used. Proliferative response as well as IFNr secretion were restored by incubation in the presence of exogenous IL2. Enzyme study disclosed a profound deficiency in cytidine deaminase (CDA) in the patient's lymphocytes (10-21-23 pmol.min−1.10−6 cells, normal value 130 ± 30, n = 39). Low values were found in the father (20-21) and the mother (18-55). Cytidine (100 μM) did reverse the inhibitory effect of pyrazofurin on parents' lymphocytes PHA-induced proliferation but not that of patient's lymphocytes. During a 6-months survey lymphocyte CDA activity of the patient, his father and mother rose to 63, 140 and 130 pmol.min−1.10−6 cells, respectively. The fact that CDA deficiency has been transitory in the parents rules out an hereditary enzymatic defect. Nevertheless, our results suggest that CDA activity should be systematically determined in immunodeficient patients.

Background Pulmonary arterial hypertension (PAH) is a rare but devastating complication of bone marrow transplantation (BMT). It has been reported after BMT for malignant disease or immune deficiencies. Methods Over a period of 10 years, all children with PAH after BMT were reviewed for underlying disease, pre-BMT conditioning, associated hepatic veno-occlusive disease, characteristics of PAH, management of PAH and outcome. Results 14 patients were diagnosed to have PAH after BMT (mean age 8.5 months, mean age at BMT 5.7 months). The underlying disease was an immune deficiency in 11 cases and a malignant disease in 3 cases. Eleven patients received chemoprophylaxis before BMT and conditioning was the same in all cases. Seven patients had a hepatic veno-occlusive disease before the onset of PAH. PAH appeared 40 days after BMT (4–96 days). The presenting symptom was dyspnea in 13 cases and syncope in 1 case. Extensive microbiological investigations were negative in all patients. PAH was diagnosed on echocardiography in all cases and confirmed by right heart catheterization in 7 cases. All patients had normal wedge pressure. Acute vasodilation testing with NO˚ was positive in 4/14. Histology was performed in 1/14 patient and showed arteriolar lesions. Six/14 patients needed mechanical ventilation, 12/14 received specific therapies for PAH. Five patients died. PAH resolved in the remaining 9 patients after a follow-up of 3 months. All the survivors had a favourable haematological issue. Conclusion PAH after BMT is a rare complication that occurs mainly in infants less than 2 years of age. It is inconstantly preceded by hepatic veno-occlusive disease. Mortality is high but reversibility is constant in survivors suggesting that an aggressive treatment should be early proposed.