<Emphasis Type="Italic">RABGAP1L</Emphasis> gene rearrangement resulting from a der(Y)t(Y;1)(q12;q25) in acute myeloid leukemia arising in a child with Klinefelter syndrome

In this study, we report the molecular cytogenetic characterization of an acute myeloid leukemia with a der(Y)t(Y;1)(q12;q25) in bone marrow cells in a child with Klinefelter syndrome. Conventional cytogenetics demonstrated the unbalanced translocation, i.e., a trisomic 1q25-qter juxtaposed to Yq12 replaced the terminal segment of chromosome Y was acquired and present only on bone marrow cells. Fluorescence in situ hybridization showed that the breakpoint at 1q25 disrupted RABGAP1L, a strongly expressed gene in CFU-GEMM, erythroid cells, and megakaryocytes, while the Yq12 breakpoint fell within the heterochromatic region. As der(Y)t(Y;1)(q12;q25) was an isolated cytogenetic change, RABGAP1L rearrangement as well as gene(s) dosage effects correlated to 1q25-qter trisomy, and Yq12-qter loss may make a major contribution to leukemogenesis and/or disease progression. Maria Cristina Roberti and Roberta La Starza should be regarded as joint first authors.     

Similarity and Divergence among Adherent-Invasive Escherichia coli and Extraintestinal Pathogenic E. coli Strains

Adherent-invasive Escherichia coli (AIEC) pathovar strains, which are associated with Crohn''s disease, share many genetic and phenotypic features with extraintestinal pathogenic E. coli (ExPEC) strains, but little is known about the level of genetic similarity between the two pathovars. We aimed to determine the frequency of strains with the “AIEC phenotype” among a collection of ExPEC strains and to further search for a common phylogenetic origin for the intestinal and extraintestinal AIEC strains. The adhesion, invasion, and intramacrophage replication capabilities (AIEC phenotype) of 63 ExPEC strains were determined. Correlations between virulence genotype and AIEC phenotype and between intestinal/extraintestinal origin, serotype, and phylogroup were evaluated for the 63 ExPEC and 23 intestinal AIEC strains. Phylogenetic relationships between extraintestinal and intestinal AIEC strains were determined using multilocus sequence typing (MLST) and pulsed-field gel electrophoresis. Only four (6.35%) ExPEC strains, belonging to the O6:H1, O83:H1, and O25:H4 serotypes, were classified as having an AIEC phenotype. These strains were found to be genetically related to some intestinal AIEC strains of the same serotypes as revealed by MLST. No particular virulence gene sets correlated with the intestinal/extraintestinal origin of the strains or with the AIEC phenotype, whereas the gene sets did correlate with the serogroup. We identified two intestinal AIEC strains and one extraintestinal AIEC strain belonging to the O25:H4 serotype that also belonged to the emerging and virulent clonal group ST131. In conclusion, the ExPEC and AIEC pathovars share similar virulence gene sets, and certain strains are phylogenetically related. However, the majority of ExPEC strains did not behave like AIEC strains, thus confirming that the AIEC pathovar possesses virulence-specific features that, to date, are detectable only phenotypically.Members of the Enterobacteriaceae family, especially Escherichia coli, have been repeatedly suggested to play a role in the origin and/or perpetuation of Crohn''s disease (CD). In part, this suggestion was based on the higher abundance of this bacterium in CD patients than in control subjects (4, 10, 20, 23, 28, 29, 32, 41, 48, 51). Although considerable effort has been devoted to the search for intestinal pathogenic E. coli strains associated with CD, to date none of the six previously described pathovars (27) has been implicated in this condition. Darfeuille-Michaud et al. (18) observed that E. coli strains with adhesion and invasion properties colonized the ileal mucosae of CD patients more frequently than those of control subjects. Darfeuille-Michaud et al. further characterized these strains and proposed a new potential E. coli pathovar associated with CD, which was designated adherent-invasive E. coli (AIEC) (10). The implication of AIEC in CD is becoming increasingly relevant because several independent studies from different countries have reported a higher prevalence of invasive E. coli in CD patients (4, 17, 33, 34, 47).The main characteristics of AIEC are (i) the ability to adhere to and invade intestinal epithelial cells, (ii) the ability to survive and replicate expansively within macrophages without triggering host cell death and inducing the release of tumor necrosis factor alpha (21), and (iii) the lack of known invasive determinants (17). Recently, Glasser and Darfeuille-Michaud (22) proposed a model explaining the mechanism of pathogenesis for AIEC strains. The AIEC strains isolated to date are clonally diverse and belong to distinct serotypes. Moreover, despite the fact that they fall primarily into the B2 phylogroup, AIEC strains belonging to the A, B1, and D phylogroups have also been isolated (4, 33-35, 47). Although no specific virulence factors have been described for this pathovar, AIEC strains carry many virulence-associated genes characteristic of extraintestinal pathogenic E. coli (ExPEC) strains, which suggests that the AIEC pathovar could be closely related to the ExPEC pathovar (4, 17, 34).The aim of this work was to determine the frequency of strains with the “AIEC phenotype” among E. coli strains that cause extraintestinal infections, including uropathogenic E. coli (UPEC), septicemic E. coli, and neonatal meningitis E. coli strains. To achieve this objective, we determined the ability of a collection of ExPEC strains to adhere to and invade intestinal epithelial cells, as well as their capacity to survive and replicate within macrophages. In parallel, we compared the distributions of virulence-associated genes among ExPEC and AIEC strains. Furthermore, we searched for a common phylogenetic origin of the ExPEC strains that had an AIEC phenotype (referred to in this study as extraintestinal AIEC) and a collection of AIEC strains isolated mainly from the intestinal mucosae of CD patients (intestinal AIEC).     

Molecular characterization and phylogenetic relationships of Desmodium leaf distortion virus (DeLDV): a new begomovirus infecting <Emphasis Type="Italic">Desmodium glabrum</Emphasis> in Yucatan,Mexico

The complete DNA-A component sequence of Desmodium leaf distortion virus (DeLDV, Begomovirus) isolated in Yucatan was determined to be 2569 nucleotides (nt) in length, and it was most closely related to Cotton leaf crumple virus-California (CLCrV-[Cal]), at 76%. The complete DNA-B component sequence was 2514 nt in length, and shared its highest nucleotide identity (60%) with Potato yellow mosaic Trinidad virus (PYMTV). Phylogenetic analyses group the DeLDV DNA-A component in the SLCV clade, whereas, the DeLDV DNA-B was grouped with the Abutilon mosaic virus clade, which also contains PYMV, suggesting that the DeLDV components have distinct evolutionary histories, possibly as the result of recombination and reassortment.     

A new CYP21A1P/CYP21A2 chimeric gene identified in an Italian woman suffering from classical congenital adrenal hyperplasia form

Background  

More than 90% of Congenital Adrenal Hyperplasia (CAH) cases are associated with mutations in the 21-hydroxylase gene (CYP21A2) in the HLA class III area on the short arm of chromosome 6p21.3. In this region, a 30 kb deletion produces a non functional chimeric gene with its 5' and 3' ends corresponding to CYP21A1P pseudogene and CYP21A2, respectively. To date, five different CYP21A1P/CYP21A2 chimeric genes have been found and characterized in recent studies. In this paper, we describe a new CYP21A1P/CYP21A2 chimera (CH-6) found in an Italian CAH patient.     

Caveolin-1 Expression Determines the Route of Neutrophil Extravasation through Skin Microvasculature

Interleukin-8 plays a key role in the acute inflammatory response by mediating recruitment of neutrophils through vessel walls into affected tissues. During this process, molecular signals guide circulating blood neutrophils to target specific vessels for extravasation and to migrate through such vessels via particular routes. Our results show that levels of endothelial caveolin-1, the protein responsible for the induction of the membrane domains known as caveolae, are critical to each of these processes. We demonstrate that, in response to the intradermal injection of interleukin-8, neutrophils are preferentially recruited to a unique subset of venules that express high levels of intercellular adhesion molecule-1 and low levels of caveolin-1. Our results show that neutrophils traverse human dermal microvascular endothelial cells using one of two pathways: a transcellular route directly through the cell or a paracellular route through cellular junctions. Caveolin-1 expression appears to favor the transcellular path while down-regulation of caveolin-1 promotes the paracellular route.Wounding of the epithelium and entry of a foreign body elicit a series of responses from the innate immune system. One of the main hallmarks of acute inflammation is neutrophil infiltration at the affected site.^1,2 In response to injury or infection, resident phagocytic cells become activated and release inflammatory cytokines such as tumor necrosis factor (TNF)-α and interleukin (IL)-8. TNF-α activates the vascular endothelium causing vasodilation and cellular infiltration.³ IL-8 functions as a critical chemotactic factor attracting neutrophils from the blood to the affected area.^1,4It is currently thought that leukocyte recruitment and migration through the vasculature is an active process not only for migrating blood cells but also for endothelial cells lining the vessels. Initially, inflammatory cytokines or bacterial endotoxins induce expression of P- and E-selectin on the surface of microvascular endothelial cells.^5,6 These molecules recognize carbohydrate counterligands on the surface of circulating leukocytes and mediate the tethering and rolling of these cells along vessel walls.^5,6,7 Firm adhesion is then initiated through the upregulation of endothelial adhesion molecules such as intercellular adhesion molecule (ICAM)-1 and vascular adhesion molecule (VCAM)-1, which bind to integrins expressed on the leukocyte surface.^5,7,8 Finally, the leukocyte is induced to migrate through the vessel in a process known as diapedesis.^5,6,7Among the many proteins implicated in the process of diapedesis, the adhesion molecule ICAM-1, which is up-regulated on activated endothelium, and caveolin-1, which is expressed on most terminally differentiated cell types but is largely undetectable in white blood cells, have been most closely associated with the route of transendothelial migration in in vitro systems.^7,9,10 A recent study by Millan et al clearly demonstrates that ICAM-1 and caveolin-1 are involved in directing the path of T lymphoblast migration through human umbilical vein endothelial cells (HUVECs).⁷Although both caveolin-1 and ICAM-1 have been associated with leukocyte transendothelial migration in vitro, the distribution of these proteins in vessels used by migrating leukocytes in vivo remain unclear. While all endothelial cells (ECs) share common features, the vascular tree is known to be extremely heterogeneous. As a result, the precise molecular profile of selectins and adhesion molecules defining vessels targeted for extravasation by circulating leukocytes is unknown. Furthermore, since the phenotype of vessel ECs is determined in large part by their unique in vivo microenvironment, site specific and regional differences in the expression of molecules contributing to the regulation of leukocyte transmigration have yet to be thoroughly characterized.^11,12 In this study, we have examined the in vivo molecular profile of vessels targeted by circulating neutrophils in response to IL-8 in the skin and have determined the effect of the expression of these factors on the route of neutrophil transmigration in vitro.     

Analysis of Immunity to Febrile Malaria in Children That Distinguishes Immunity from Lack of Exposure

In studies of immunity to malaria, the absence of febrile malaria is commonly considered evidence of “protection.” However, apparent “protection” may be due to a lack of exposure to infective mosquito bites or due to immunity. We studied a cohort that was given curative antimalarials before monitoring began and documented newly acquired asymptomatic parasitemia and febrile malaria episodes during 3 months of surveillance. With increasing age, there was a shift away from febrile malaria to acquiring asymptomatic parasitemia, with no change in the overall incidence of infection. Antibodies to the infected red cell surface were associated with acquiring asymptomatic infection rather than febrile malaria or remaining uninfected. Bed net use was associated with remaining uninfected rather than acquiring asymptomatic infection or febrile malaria. These observations suggest that most uninfected children were unexposed rather than “immune.” Had they been immune, we would have expected the proportion of uninfected children to rise with age and that the uninfected children would have been distinguished from children with febrile malaria by the protective antibody response. We show that removing the less exposed children from conventional analyses clarifies the effects of immunity, transmission intensity, bed nets, and age. Observational studies and vaccine trials will have increased power if they differentiate between unexposed and immune children.Malaria is a pressing global health problem (36). The correlates of immunity in observational field-based studies are often used to guide vaccine design (22), in which the chosen definition of immunity to malaria is usually the absence of febrile malaria. However, the findings obtained with this approach are often inconsistent, and responses to a specific antigen are associated with protection in some studies but not in others (4, 6, 7, 9-12, 23, 29). This may be because of parasite polymorphism (38), because of a confounding association between protective and nonprotective responses, because the endpoint of mild febrile malaria is not specific (26), or because rapidly waning antibody responses are not a stable predictive measure for the follow-up period (15).In studies in Kilifi, Kenya, associations between specific antibody responses and protection were stronger in children who had asymptomatic parasitemia at the start of monitoring (5, 16, 20, 28, 30, 31). This might imply that there is premunition, where a chronic low-level infection is required to provide immunity against further infection (35), and that antibody responses are more long lived in the presence of asymptomatic parasitemia (1). Alternatively, antibody responses measured in the presence of a challenge with asymptomatic parasitemia may be more informative than antibody responses measured without current exposure. For instance, protection against hepatitis B is predicted by the antibody titer shortly after vaccination, even when antibody titers subsequently become undetectable (32). However, it may simply be that parasitemia reflects greater exposure to malaria and hence a greater power to detect associations.In this study, we cleared asymptomatic parasitemia with highly effective antimalarials in order to identify newly acquired parasitemia during follow-up. We compared children who acquired asymptomatic parasitemia with children who developed febrile malaria by examining the associations with known markers of exposure and immunity. We then examined what impact excluding “unexposed” children had on conventional survival analyses in order to determine whether such analyses should be more widely used to study outcomes in observational studies or clinical trials.     

α7 and non-α7 nicotinic acetylcholine receptors modulate dopamine release in vitro and in vivo in the rat prefrontal cortex

Nicotine enhances attentional and working memory aspects of executive function in the prefrontal cortex (PFC) where dopamine plays a major role. Here, we have determined the nicotinic acetylcholine receptor (nAChR) subtypes that can modulate dopamine release in rat PFC using subtype-selective drugs. Nicotine and 5-Iodo-A-85380 (β2* selective) elicited [³H]dopamine release from both PFC and striatal prisms in vitro and dopamine overflow from medial PFC in vivo . Blockade by dihydro-β-erythroidine supports the participation of β2* nAChRs. However, insensitivity of nicotine-evoked [³H]dopamine release to α-conotoxin-MII in PFC prisms suggests no involvement of α6β2* nAChRs, in contrast to the striatum, and this distinction is supported by immunoprecipitation of nAChR subunits from these tissues. The α7 nAChR-selective agonists choline and Compound A also promoted dopamine release from PFC in vitro and in vivo , and their effects were enhanced by the α7 nAChR-selective allosteric potentiator PNU-120596 and blocked by specific antagonists. DNQX and MK801 inhibited [³H]dopamine release evoked by choline and PNU-120596, suggesting crosstalk between α7 nAChRs, glutamate and dopamine in the PFC. In vivo, systemic (but not local) administration of PNU-120596, in the absence of agonist, facilitated dopamine overflow in the medial PFC, consistent with the activation of extracortical α7 nAChRs by endogenous acetylcholine or choline. These data establish that both β2* and α7 nAChRs can modulate dopamine release in the PFC in vitro and in vivo . Through their distinct actions on dopamine release, these nAChR subtypes could contribute to executive function, making them specific therapeutic targets for conditions such as schizophrenia and attention deficit hyperactivity disorder.