Adenosine A2A receptor gene: Evidence for association of risk variants with panic disorder and anxious personality

Adenosine A_2A receptors are suggested to play an important role in different brain circuits and pathways involved in anxiety reactions. A variant within the corresponding ADORA2A gene (rs5751876) increased the risk for panic disorder (PD), for elevated anxiety during challenge tests in healthy probands and for anxiety-related arousal in blood-injury phobia. These multiple effects may mirror a more general effect of the SNP on basic personality traits. In the present study we therefore aimed to replicate the original finding in a large PD sample and extend it by investigating an additional proband sample characterized for different anxiety-related personality scores. In addition, as rs5751876 is assumed not to be the disease variant itself but to be in linkage disequilibrium (LD) with the true functional polymorphism other SNPs of potentially functional relevance were identified by re-sequencing the whole gene including several newly identified regions of putative regulatory potential and analysed for their impact on PD and anxious personality. We were indeed able to replicate rs5751876 as risk factor for PD, particularly PD with agoraphobia. Rs5751876 and several other variants in high LD (rs5751862, rs2298383 and rs3761422) as well as the corresponding haplotypes were also associated with different anxiety-related personality scores (Bonferroni corrected P_all < 0.05). Of these variants, rs2298383 shows functional potential based on in silico analyses and might therefore represent the true underlying causal variant. Our data provide further support for an important role of ADORA2A variants in the pathogenesis of anxiety disorders and anxious personality reflecting their potential as basic susceptibility factors.     

Acceptance of female urologists among patients with suspected prostate disease

BackgroundsThe number of practicing female urologists is rising. The aim of this study is to evaluate the acceptance of female urologists by male patients and their partners.MethodsMen who underwent a prostate MRI or a prostate biopsy between January and December 2018 and their partners, were sent questionnaires prior to the examination. Two types of questionnaires were used. One questionnaire asked “I want to be seen by: (I) a male urologist or (II) a female urologist or (III) no preference” (Group_np), the other questionnaire only offered two possible answers: “I want to be seen by: (I) a male urologist or (II) a female urologist” (Group_m,f). All other questions were on prostate MRI and prostate biopsies.ResultsOverall, 377 questionnaires were sent to patients. One hundred and ninety-six questionnaires (52.0%) were returned. In Group_np, 34.7% wanted to be seen by a male urologist, 60.8% of patients chose “no preference”. The answers of the patients’ female partners in Group_np did not differ statistically significant (57.3% chose “no preference”, 0% chose a female urologist). In Group_m,f, 54.5% of patients preferred a male urologist, one patient wanted to be seen by a female urologist, 44.3% did not answer the question. In Group_m,f, there was no statistically significant difference in preference in regard to the doctor’s gender between the patients and their female partners (57% of partners wanted a male urologist, 0% wanted a female urologist).ConclusionsA large number of patients with prostate disease and their partners prefer male urologists rather than female urologists.     

Bone morphogenetic protein 4 is induced in hepatocellular carcinoma by hypoxia and promotes tumour progression

Striking similarities exist between molecular mechanisms driving embryonic liver development and progression of hepatocellular carcinoma (HCC). Bone morphogenetic proteins (BMPs), particularly BMP4, have been proposed to regulate embryonic hepatic development. BMP expression has been observed in neoplasia but the expression and biological role of BMP4 in human HCC are unknown. We found increased BMP4 mRNA and protein in HCC cell lines and tissue samples compared to primary human hepatocytes and corresponding non‐tumourous tissue. Hypoxia further induced BMP4 expression in HCC cells, which was abolished by transfection of a dominant negative form of HIF‐1alpha (dnHIF‐1alpha). However, gel shift assays revealed only minor binding activity in nuclear extracts from (hypoxic) HCC cells to a putative hypoxia‐response element in the BMP4 promoter. Sequence analysis of the BMP4 promoter revealed two Ets‐1 binding sites, and Ets‐1 activity was increased in HCC cells under hypoxic conditions. Transfection of dnHIF‐1alpha completely abrogated hypoxia‐induced Ets‐1 activity as well as BMP4 expression. Overexpression of Ets‐1 markedly enhanced BMP4 promoter activity, while antisense Ets‐1 almost completely abolished basal as well as hypoxia‐induced BMP4 expression. These data demonstrate that Ets‐1 activity contributes to baseline expression of the BMP4 gene and is the predominant mediator of the HIF‐dependent BMP4 induction under hypoxic conditions. To determine the functional relevance of BMP4 expression, HCC cell lines were treated with antisense BMP4 constructs or siRNA against BMP4. BMP4 suppression resulted in a strong reduction of the migratory and invasive potential and anchorage‐independent growth. Furthermore, tube formation assays indicated that BMP4 expressed by HCC cells promotes vasculogenesis. Our findings demonstrate that BMP4 is increased in HCC and promotes HCC progression. Therefore, BMP4 expression may have clinical relevance, and interfering with BMP4 signalling appears as an attractive therapeutic target for this highly aggressive tumour. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Divergent modulation of Chlamydia pneumoniae infection cycle in human monocytic and endothelial cells by iron,tryptophan availability and interferon gamma

Chlamydia pneumoniae is an obligatory intracellular bacterium causing chronic inflammatory diseases in humans. We studied the role of the nutritive factors, iron and tryptophan, towards the course of infection and immune response pathways in C. pneumoniae infected endothelial cells and monocytes.Human endothelial (EA.hy923) and monocytic cells (THP-1) were infected with C. pneumoniae, supplemented with iron or 1-methyltryptophan (1-MT), an inhibitor of the tryptophan degrading enzyme indoleamine 2,3-dioxygenase (IDO), and subsequently stimulated with IFN-γ or left untreated. The number of infected cells, the morphology and quantity of C. pneumoniae inclusion bodies, IDO activity and innate immune effector pathways were analysed.While neither iron challenge, IDO inhibition or IFN-γ treatment had a significant effect on C. pneumoniae morphology or numbers within THP-1 monocytic cells, iron supplementation to EA.hy926 cells resulted in promotion of C. pneumoniae proliferation and differentiation while IFN-γ had an inhibitory effect. Furthermore, the number of infected endothelial cells was significantly decreased upon 1-MT treatment. C. pneumoniae infection induced a pro-inflammatory immune response as evidenced by increased IDO activity, neopterin formation or TNF-α production in THP-1 but not in endothelial cells. These pathways were superinduced upon IFN-γ treatment and partly modulated by iron supplementation.Our results demonstrate that the infectious cycle of C. pneumoniae behaves differently between monocytic and endothelial cells. While the intracellular pathogen remains in a persistent form within monocytes, it can differentiate and proliferate within endothelial cells indicating that endothelial cells are a preferred environment for Chlamydia. Nutritive factors such as iron have subtle effects on C. pneumoniae biology in endothelial, but not monocytic cells. Our results contribute to a better understanding of C. pneumoniae infection and its role in chronic inflammatory diseases such as atherosclerosis.     

Interoceptive sensitivity in anxiety and anxiety disorders: An overview and integration of neurobiological findings

Interoceptive sensitivity, particularly regarding heartbeat, has been suggested to play a pivotal role in the pathogenesis of anxiety and anxiety disorders. This review provides an overview of methods which are frequently used to assess heartbeat perception in clinical studies and summarizes presently available results referring to interoceptive sensitivity with respect to heartbeat in anxiety-related traits (anxiety sensitivity, state/trait anxiety), panic disorder and other anxiety disorders. In addition, recent neurobiological studies of neuronal activation correlates of heartbeat perception using positron emission tomography (PET), functional magnetic resonance imaging (fMRI) or electroencephalographic (EEG) techniques are presented. Finally, possible clinical and therapeutic implications (e.g., beta-blockers, biofeedback therapy, cognitive interventions and interoceptive exposure) of the effects of heartbeat perception on anxiety and the anxiety disorders and the potential use of interoceptive sensitivity as an intermediate phenotype of anxiety disorders in future neurobiological and genetic studies are discussed.     

MYC High Level Gene Amplification Is a Distinctive Feature of Angiosarcomas after Irradiation or Chronic Lymphedema

Angiosarcomas (AS) are rare vascular malignancies that arise either de novo as primary tumors or secondary to irradiation or chronic lymphedema. The cytogenetics of angiosarcomas are poorly characterized. We applied array-comparative genomic hybridization as a screening method to identify recurrent alterations in 22 cases. Recurrent genetic alterations were identified only in secondary but not in primary AS. The most frequent recurrent alterations were high level amplifications on chromosome 8q24.21 (50%), followed by 10p12.33 (33%) and 5q35.3 (11%). Fluorescence in situ hybridization analysis in 28 primary and 33 secondary angiosarcomas (31 tumors secondary to irradiation, 2 tumors secondary to chronic lymphedema) confirmed high level amplification of MYC on chromosome 8q24.21 as a recurrent genetic alteration found exclusively in 55% of AS secondary to irradiation or chronic lymphedema, but not in primary AS. Amplification of MYC did not predispose to high grade morphology or increased cell turnover. In conclusion, despite their identical morphology, secondary AS are genetically different from primary AS and are characterized by a high frequency of high level amplifications of MYC. This finding may have implications both for the diagnosis and treatment of these tumors.Angiosarcomas (AS) are rare sarcomas with morphological and functional properties of endothelial cells.¹ AS represent <1% of all sarcomas.² Roughly 35% of cases arise in the skin, 25% in soft tissue and the rest in various other locations including breast, liver, spleen, and bone.¹ The prognosis of AS has generally been considered to be poor with unpredictable clinical behavior. However, several publications clearly showed that prognosis depends on the primary site with a particularly poor prognosis for tumors arising in liver, spleen, heart, and bone with a 5-year-survival rate of 0%, as compared with approximately 50% for skin and soft tissue AS.^3,4 Other factors with an adverse impact on prognosis are older age, tumor necrosis, and epitheloid features.⁵ Tumors can arise either de novo (primary AS, pAS) or as secondary conditions in patients with long standing lymphedema^6,7 or after irradiation⁸ (secondary AS; sAS), especially in female patients irradiated for breast cancer. In fact, sAS of the breast is by far the most frequent radiation-induced sarcoma in women treated with radiation therapy as part of their initial treatment,⁹ with a more than 1000-fold increased relative risk¹⁰ as compared with the general population. It has been calculated that the standardized incidence ratio in irradiated breast cancer patients to develop AS is 26 compared with 3.8 to develop any other sarcoma⁹ over a latency period of 5 to 10 years. Of note, the latency period of sAS has been reported to be much shorter than in other radiation-induced sarcomas, which is generally >10 years.^11–14 Relatively little is known about the genetic changes in postradiation sarcomas in general^15–18 and in sAS in particular, where published genetic studies are limited to case reports or small case collections.^19–28 Probably due to the small case numbers, available data have so far failed to show consistent recurrent chromosomal abnormalities. In this report, we studied clinicopathologic and molecular genetic features in 61 primary and secondary AS from multiple international institutions.     

Novel role of regulatory T cells in limiting early neutrophil responses in skin

It is clear that CD4⁺ CD25⁺ Foxp3⁺ regulatory T (Treg) cells inhibit chronic inflammatory responses as well as adaptive immune responses. Among the CD4⁺ T‐cell population in the skin, at least one‐fifth express Foxp3. As the skin is constantly exposed to antigenic challenge and is a common site of vaccination, understanding the role of these skin‐resident Treg cells is important. Although the suppressive effect of Treg cells on T cells is well documented, less is known about the types of innate immune cells influenced by Treg cells and whether the Treg cells suppress acute innate immune responses in vivo. To address this we used a mouse melanoma cell line expressing Fas ligand (B16FasL), which induces an inflammatory response following subcutaneous injection of mice. We demonstrate that Treg cells limit this response by inhibiting neutrophil accumulation and survival within hours of tumour cell inoculation. This effect, which was associated with decreased expression of the neutrophil chemoattractants CXCL1 and CXCL2, promoted survival of the inoculated tumour cells. Overall, these data imply that Treg cells in the skin are rapidly mobilized and that this activity serves to limit the amplification of inflammatory responses at this site.     

Embryonated Eggs as an Alternative Infection Model To Investigate Aspergillus fumigatus Virulence

Infection models are essential tools for studying microbial pathogenesis. Murine models are considered the “gold standard” for studying in vivo infections caused by Aspergillus species, such as A. fumigatus. Recently developed molecular protocols allow rapid construction of high numbers of fungal deletion mutants, and alternative infection models based on cell culture or invertebrates are widely used for screening such mutants to reduce the number of rodents in animal experiments. To bridge the gap between invertebrate models and mice, we have developed an alternative, low-cost, and easy-to-use infection model for Aspergillus species based on embryonated eggs. The outcome of infections in the egg model is dose and age dependent and highly reproducible. We show that the age of the embryos affects the susceptibility to A. fumigatus and that increased resistance coincides with altered chemokine production after infection. The progress of disease in the model can be monitored by using egg survival and histology. Based on pathological analyses, we hypothesize that invasion of embryonic membranes and blood vessels leads to embryonic death. Defined deletion mutant strains previously shown to be fully virulent or partially or strongly attenuated in a mouse model of bronchopulmonary aspergillosis showed comparable degrees of attenuation in the egg model. Addition of nutrients restored the reduced virulence of a mutant lacking a biosynthetic gene, and variations of the infectious route can be used to further analyze the role of distinct genes in our model. Our results suggest that embryonated eggs can be a very useful alternative infection model to study A. fumigatus virulence and pathogenicity.Aspergillus fumigatus is a ubiquitous mold with the ability to cause life-threatening disease in immunocompromised human patients (19). Despite the development of novel antimycotics, the lethality of invasive aspergillosis in human patients is still high and can exceed 50% (14, 57). Thus, a better understanding of the infectious process and the fungal factors involved in pathogenesis is essential for development of more effective treatments. One approach to identify virulence factors is to study fungal gene deletion mutants in appropriate infection models. The most commonly used infection models for A. fumigatus are laboratory rodents, especially mice. These models are well characterized and have been critical for understanding host-pathogen interactions, as well as for developing better therapeutic approaches (11). However, ethical considerations and legal restrictions limit the use of mammals for infection studies. Additionally, animal experiments are costly and require specialized facilities and specially trained personal, which further limits the availability of mammalian models to researchers. Therefore, several alternative infection models for fungal pathogens have been developed in recent years, ranging from cell lines and tissue cultures to invertebrate hosts. Insect models, like the fruit fly (Drosophila melanogaster) (28, 29) and larvae of the greater wax moth (Galleria mellonella) (21, 30, 37, 42, 43), have been successfully used to investigate virulence traits of A. fumigatus and the efficacy of antimycotic treatment. However, these models are limited compared to mammalian hosts; the limitations include, for example, the low temperature required for experiments, the route of infection, and certain aspects of the immune response. One possible way to bridge the gap between insect and mammalian hosts is to use embryonated eggs as an alternative infection model.Birds of all ages are susceptible to aspergillosis, and this disease has been described for a variety of avian species, both birds in captivity and wild birds (54). Aspergillosis outbreaks can cause significant economic losses in the poultry industry (41) and have prompted the development of vaccination strategies to prevent Aspergillus infections (46). The avian immune response to Aspergillus infections is similar to the mammalian response with respect to the beneficial effect of a type 1 response, the role of macrophages and heterophils (avian immune cells similar to mammalian neutrophils), and the production of specific antibodies (10, 27, 33). Although these features suggest that birds could be an interesting and potential animal model for studying aspergillosis (11, 47), the same ethical, legal, financial, and logistical restrictions that apply to experiments with mammals apply to experiments with birds. However, embryonated bird eggs could provide an alternative model, which would fulfill the demand for refinement of animal models. Fertilized eggs are readily available from commercial breeders at low cost, are easy to handle, and require little specialized equipment and no specialized facilities or personal. Since embryonated eggs have been successfully used to study virulence features of various bacteria (2, 55) and the fungal pathogen Candida albicans (16, 17), we investigated the potential of embryonated eggs as an alternative model for studying the virulence of A. fumigatus.     

Extended runs of homozygosity at 17q11.2: an association with type‐2 NF1 deletions?

Large deletions in the NF1 gene region at 17q11.2 are caused by nonallelic homologous recombination (NAHR). The recurrent type‐2 NF1 deletions span 1.2 Mb, with breakpoints in the SUZ12 gene and SUZ12P. Type‐2 NF1 deletions occur preferentially during mitosis and are associated with somatic mosaicism. A panel of 16 type‐2 NF1 deletions was used as a model system in which to investigate whether extended homozygosity across 17q11.2 might be associated with somatic deletion. Using SNP arrays, a 3.2 Mb interval encompassing the NF1 deletion region was found to harbor runs of homozygosity (ROHs) in different human populations. However, ROHs ≥500 kb directly flanking the NF1 deletion region on both sides were not found to occur disproportionately in NF1 patients harboring type‐2 deletions compared to controls. Although low allelic diversity in 17q11.2 is unlikely to be a key factor in promoting NAHR‐mediated somatic type‐2 deletions, a specific ROH of 588 kb (roh1), located some 525 kb proximal to the deletion interval, was found to occur more frequently (P=0.012) in the type‐2 deletion patients compared with controls. We postulate that roh1 may act remotely, via an as yet unknown mechanism, to increase the frequency of somatic recombination between the distally duplicated SUZ12 sequences. Hum Mutat 30:1–10, 2010. © 2010 Wiley‐Liss, Inc.