Consensus statements on vaccination in patients with haemophilia-Results from the Italian haemophilia and vaccinations (HEVA) project.

Vaccination against communicable diseases is crucial for disease prevention, but this practice poses challenges to healthcare professionals in patients with haemophilia. Poor knowledge of the vaccination requirements for these patients and safety concerns often result in vaccination delay or avoidance. In order to address this issue, a panel of 11 Italian haemophilia and immunization experts conducted a Delphi consensus process to identify the main concerns regarding the safe use of vaccines in patients with haemophilia. The consensus was based on a literature search of the available evidence, which was used by the experts to design 27 consensus statements. A group of clinicians then rated these statements using the 5-point Likert-type scale (1 = strongly disagree; 5 = strongly agree). The main issues identified by the expert panel included vaccination schedule for haemophilic patients; protocol and optimal route of vaccine administration; vaccination of haemophilic patients with antibodies inhibiting coagulation factor VIII (inhibitors); and vaccination and risk of inhibitor development. This manuscript discusses these controversial areas in detail supported by the available literature evidence and provides evidence- and consensus-based recommendations. Overall, participants agreed on most statements, except those addressing the potential role of vaccination in inhibitor formation. Participants agreed that patients with haemophilia should receive vaccinations according to the institutional schedule for individuals without bleeding disorders; however, vaccination of patients with haemophilia requires comprehensive planning, taking into account disease severity, type and route of vaccination, and bleeding risk. Data also suggest vaccination timing does not need to take into consideration when the patient received factor VIII replacement.

Live from the liver: hepatocyte IL-7.

Interleukin 7 (IL-7), which is required for T cell survival, was previously found in lymphoid tissues. In this issue of Immunity, Sawa et al. (2009) have identified the liver as a new source of IL-7.

Interleukin-7 receptor expression: intelligent design.

Interleukin-7 (IL-7) is produced by stromal cells in lymphoid tissues and is required for the development of T cells and for their persistence in the periphery. Unlike many other cytokines that act on lymphocytes, IL-7 production by stromal cells is not substantially affected by extrinsic stimuli. So, the amount of available IL-7 protein is thought to be regulated by the rate that it is scavenged by T cells. As we review here, there is mounting evidence indicating that the amount of IL-7 receptor expressed on a cell not only determines how vigorously the cell responds to IL-7, but it can also determine how efficiently the cell consumes IL-7 and, therefore, affect the supply of this limiting resource in the niche.

The human IL-7 receptor gene: deletions, polymorphisms and mutations.

Most T cell subsets depend on IL-7 for survival. IL-7 binds to IL-7Rα and γc, initiating the signaling cascade. Deletion of IL-7Ra in humans has, for some time, been known to cause severe combined immunodeficiency. More recently, polymorphisms in IL-7R have been shown be a risk factor for a number of diseases that are autoimmune or involve excess immune and inflammatory responses including multiple sclerosis, type 1 diabetes, rheumatoid arthritis, primary biliary cirrhosis, inflammatory bowel disease, atopic dermatitis, inhalation allergy, sarcoidosis and graft-versus host disease. The polymorphism that affects risk to most of these immunopathologies is T244I at the border of the extracellular domain and the transmembrane region. The same region has recently been shown to harbor gain-of-function mutations in acute lymphoblastic leukemia. These studies have suggested new therapies that target the IL-7 pathway.

Development of regulatory T cells requires IL-7Ralpha stimulation by IL-7 or TSLP.

Interleukin-7 (IL-7), a cytokine produced by stromal cells, is required for thymic development and peripheral homeostasis of most major subsets of T cells. We examined whether regulatory T (Treg) cells also required the IL-7 pathway by analyzing IL-7Ralpha(-/-) mice. We observed a striking reduction in cells with the Treg surface phenotype (CD4, CD25, GITR (glucocorticoid-induced tumor necrosis factor [TNF]-like receptor), CD45RB, CD62L, CD103) or intracellular markers (cytotoxic T-lymphocyte-associated antigen-4, CTLA-4, and forkhead box transcription factor 3, Foxp3). Foxp3 transcripts were virtually absent in IL-7Ralpha(-/-) lymphoid tissues, and no Treg cell suppressive activity could be detected. There are 2 known ligands for IL-7Ralpha: IL-7 itself and thymic stromal lymphopoietin (TSLP). Surprisingly, mice deficient in IL-7 or the other chain of the TSLP receptor (TSLPR) developed relatively normal numbers of Treg cells. Combined deletion of IL-7 and TSLP receptor greatly reduced Treg cell development in the thymus but was not required for survival of mature peripheral Treg cells. We conclude that Treg cells, like other T cells, require signals from the IL-7 receptor, but unlike other T cells, do not require IL-7 itself because of at least partially overlapping actions of IL-7 and TSLP for development of Treg cells.

Cell biology of IL-7, a key lymphotrophin.

IL-7 is essential for the development and survival of T lymphocytes. This review is primarily from the perspective of the cell biology of the responding T cell. Beginning with IL-7 receptor structure and regulation, the major signaling pathways appear to be via PI3K and Stat5, although the requirement for either has yet to be verified by published knockout experiments. The proliferation pathway induced by IL-7 differs from conventional growth factors and is primarily through posttranslational regulation of p27, a Cdk inhibitor, and Cdc25a, a Cdk-activating phosphatase. The survival function of IL-7 is largely through maintaining a favorable balance of bcl-2 family members including Bcl-2 itself and Mcl-1 on the positive side, and Bax, Bad and Bim on the negative side. There are also some remarkable metabolic effects of IL-7 withdrawal. Studies of IL-7 receptor signaling have yet to turn up unique pathways, despite the unique requirement for IL-7 in T cell biology. There remain significant questions regarding IL-7 production and the major producing cells have yet to be fully characterized.

A point mutation affecting an SP1 binding site in the promoter of the ferrochelatase gene impairs gene transcription and causes erythropoietic protoporphyria.

OBJECTIVE: Clinical manifestation of erythropoietic protoporphyria (EPP) results from coinheritance of a mutated allele and a wild-type low-expressed allele of the ferrochelatase (FECH) gene. Currently, up to 90 different mutations affecting the coding region or splicing junctions of the FECH gene have been identified. Despite the high molecular heterogeneity, no functional mutations have been previously reported in the promoter region. The weaker allele expression has been controversially associated to the presence of different intragenic polymorphisms. METHODS: We applied a two-step screening strategy using denaturing gradient gel electrophoresis followed by direct sequencing in order to rapidly identify FECH gene mutations in Italian EPP patients. We identified two unrelated subjects showing a normal FECH coding region but a single G>C base substitution at position -250 in the FECH promoter and the -251G, IVS1-23T, and IVS3-48C polymorphisms in trans to the substitution. To investigate the effect of the -250G>C mutation on protein binding to the FECH promoter, we conducted electro mobility shift assay (EMSA) and supershift analysis. To determine its effect on the transcriptional activity, K562 and Jurkat cell lines were transiently transfected. RESULTS: EMSA showed that the -250G>C mutation results in the loss of an SP1 binding site, and transient transfection assays demonstrated that such mutation strongly impairs promoter activity. Moreover, we showed that the -251A>G polymorphism, although unable to affect SP1 binding, displays a significant reduction in the transcriptional activity of the promoter. CONCLUSION: This is the first report of a mutation in the FECH promoter affecting binding of a transcription factor and causing EPP phenotype.

Establishment of an ex vivo model of monocytes-derived macrophages differentiated from peripheral blood mononuclear cells (PBMCs) from HIV-1 transgenic rats.

It has been developed an HIV type 1 transgenic rat model (HIV-1 Tg Rat) which contains a gag-pol-deleted HIV-1 provirus regulated by the viral LTR promoter. Although it harbors a non infectious provirus, efficient viral expression occurs in different tissues and disease manifestations as well as immune-response alterations and pathologies similar to humans can be observed. Regulation of HIV-1 expression is influenced by various cellular factors and it is well known that macrophages are one of the major reservoir of HIV-1 infection and a vehicle for virus spread to other tissues. Purpose of our work was to establish an antigen presenting cells (monocyte-derived macrophages, MDM) ex vivo model from these HIV-1 transgenic rats. This model can be used to study function of HIV-infected MDM and their behavior like HIV-1 reservoir. Ultimately, these studies may be helpful in defining approaches to control HIV-1 spread.

Expression of interleukin-15 and interleukin-15Rα in monocytes of HIV type 1-infected patients with different courses of disease progression.

Interleukin-15 (IL-15) enhances the effector mechanisms of anti-HIV immune responses and thus is considered a potential adjuvant of HIV-1 vaccine. However, there are a lack of data concerning the relationships between IL-15 expression and regulation in HIV-1-infected patients and the course of disease progression. We found that IL-15, but not IL-15Rα, is expressed at significantly higher levels in the CD14(+) monocytes [stimulated or not with interferon (IFN)-γ] of long-term nonprogressors (LTNP) than in those of HIV-1 progressors or healthy controls. There was no between-group difference in the amounts of soluble IL-15 released from the cells. We also found that like the healthy controls, the LTNP expressed the IL-15 and IL-15Rα genes in a more coordinated manner than the progressors. Our findings show that there are significant differences in IL-15 expression between patients with different courses of HIV infection, and that the coordinated expression of the IL-15 and IL-15Rα genes is dysregulated in patients with progressive disease. They also provide important information concerning the mechanisms of infection and the potential use of IL-15 as a therapeutic agent.

Visualization and identification of IL-7 producing cells in reporter mice.

Interleukin-7 (IL-7) is required for lymphocyte development and homeostasis although the actual sites of IL-7 production have never been clearly identified. We produced a bacterial artificial chromosome (BAC) transgenic mouse expressing ECFP in the Il7 locus. The construct lacked a signal peptide and ECFP (enhanced cyan fluorescent protein) accumulated inside IL-7-producing stromal cells in thoracic thymus, cervical thymus and bone marrow. In thymus, an extensive reticular network of IL-7-containing processes extended from cortical and medullary epithelial cells, closely contacting thymocytes. Central memory CD8 T cells, which require IL-7 and home to bone marrow, physically associated with IL-7-producing cells as we demonstrate by intravital imaging.

IL-7 induces myelopoiesis and erythropoiesis.

IL-7 administration to mice was previously reported to increase the mobilization of progenitor cells from marrow to peripheral sites. We now report that IL-7 increases the number of mature myeloid and monocytic cells in spleen and peripheral blood. This effect required T cells, and we show that IL-7 treatment in vivo induced GM-CSF and IL-3 production by T cells with memory phenotype. However, additional myelopoietic cytokines were shown to be involved because mice deficient in both GM-CSF and IL-3 also responded to IL-7 with increased myelopoiesis. Candidate cytokines included IFN-gamma and Flt3 ligand, which were also produced in response to IL-7. Because IFN-gamma-deficient mice also increased myelopoiesis, it was suggested that IL-7 induced production of redundant myelopoietic cytokines. In support of this hypothesis, we found that the supernatant from IL-7-treated, purified T cells contained myelopoietic activity that required a combination of Abs against GM-CSF, IL-3, and anti-Flt3 ligand to achieve maximum neutralization. IL-7 administration increased the number of splenic erythroid cells in either normal, Rag1 or GM-CSF-IL-3-deficient mice, suggesting that IL-7 might directly act on erythroid progenitors. In support of this theory, we detected a percentage of TER-119(+) erythroid cells that expressed the IL-7Ralpha-chain and common gamma-chain. Bone marrow cells expressing IL-7R and B220 generated erythroid colonies in vitro in response to IL-7, erythropoietin, and stem cell factor. This study demonstrates that IL-7 can promote nonlymphoid hemopoiesis and production of cytokines active in the host defense system in vivo, supporting its possible clinical utility.