Adenosine A2A receptor gene (ADORA2A) variants may increase autistic symptoms and anxiety in autism spectrum disorder

Autism spectrum disorders (ASDs) are heterogeneous disorders presenting with increased rates of anxiety. The adenosine A2A receptor gene (ADORA2A) is associated with panic disorder and is located on chromosome 22q11.23. Its gene product, the adenosine A2A receptor, is strongly expressed in the caudate nucleus, which also is involved in ASD. As autistic symptoms are increased in individuals with 22q11.2 deletion syndrome, and large 22q11.2 deletions and duplications have been observed in ASD individuals, in this study, 98 individuals with ASD and 234 control individuals were genotyped for eight single-nucleotide polymorphisms in ADORA2A. Nominal association with the disorder was observed for rs2236624-CC, and phenotypic variability in ASD symptoms was influenced by rs3761422, rs5751876 and rs35320474. In addition, association of ADORA2A variants with anxiety was replicated for individuals with ASD. Findings point toward a possible mediating role of ADORA2A variants on phenotypic expression in ASD that need to be replicated in a larger sample

Scratching increases epidermal neuronal branching and alters psychophysical testing responses in atopic dermatitis and brachioradial pruritus

BackgroundChronic scratching imposes a major stress on the skin and can lead to itch intensity worsening, and consequently, patients may enter an itch–scratch cycle. This repetitive mechanical stress can result in lichenification, worsening of epidermal barrier function, and enhanced cutaneous inflammation. Furthermore, a reduction of intraepidermal nerve fibers was previously described in lichenification.AimThe aim of this study was to investigate the influence of chronic scratching on the epidermal neuroanatomy and on sensory changes, in particular the prevalence of hyperknesis and alloknesis in patients after mechanical, chemical, and electrical stimuli.MethodsAnalyses were performed on pruritic lichenified (chronically scratched), pruritic non-lichenified (not chronically scratched), and non-pruritic non-lesional (unaffected) skin areas of patients with inflammatory pruritus, i.e., atopic dermatitis (n = 35), and neuropathic pruritus, i.e., brachioradial pruritus (n = 34) vs. healthy matched controls (n = 64). Our fine-grained spatial skin characterization enabled specifically studying the differential effects of chronic scratching in inflammatory and neuropathic itch.ResultsAnalysis of intraepidermal nerve fiber density showed rarefaction of fibers in all three skin areas of patients compared with healthy controls in both diagnoses. Even more, the two pruritic areas had significantly less nerve fibers than the unaffected skin, whereas electrically induced itch was massively increased. Epidermal branching of the remaining nerve fibers in lichenified/chronically scratched skin was increased, particularly in patients with brachioradial pruritus, which may contribute to the pronounced local neuronal sensitivity. Hyperknesis and alloknesis were found to increase independently of lichenification.ConclusionOur results indicate that chronic scratching may not affect intraepidermal nerve fiber density but leads to a stronger branching pattern of intraepidermal nerve fibers, which may contribute to local hypersensitivity. The increased sensitivity in the pruritic areas suggests mechanisms of peripheral sensitization, whereas the increased sensation of electrically and chemically induced itch in unaffected skin indicates central sensitization for itch

Site-specific chromatin immunoprecipitation: a selective method to individually analyze neighboring transcription factor binding sites in vivo

<p>Abstract</p> <p>Background</p> <p>Transcription factors (TFs) and their binding sites (TFBSs) play a central role in the regulation of gene expression. It is therefore vital to know how the allocation pattern of TFBSs affects the functioning of any particular gene in vivo. A widely used method to analyze TFBSs in vivo is the chromatin immunoprecipitation (ChIP). However, this method in its present state does not enable the individual investigation of densely arranged TFBSs due to the underlying unspecific DNA fragmentation technique. This study describes a site-specific ChIP which aggregates the benefits of both EMSA and in vivo footprinting in only one assay, thereby allowing the individual detection and analysis of single binding motifs.</p> <p>Findings</p> <p>The standard ChIP protocol was modified by replacing the conventional DNA fragmentation, i. e. via sonication or undirected enzymatic digestion (by MNase), through a sequence specific enzymatic digestion step. This alteration enables the specific immunoprecipitation and individual examination of occupied sites, even in a complex system of adjacent binding motifs in vivo. Immunoprecipitated chromatin was analyzed by PCR using two primer sets - one for the specific detection of precipitated TFBSs and one for the validation of completeness of the enzyme digestion step. The method was established exemplary for Sp1 TFBSs within the <it>egfr </it>promoter region. Using this site-specific ChIP, we were able to confirm four previously described Sp1 binding sites within <it>egfr </it>promoter region to be occupied by Sp1 in vivo. Despite the dense arrangement of the Sp1 TFBSs the improved ChIP method was able to individually examine the allocation of all adjacent Sp1 TFBS at once. The broad applicability of this site-specific ChIP could be demonstrated by analyzing these SP1 motifs in both osteosarcoma cells and kidney carcinoma tissue.</p> <p>Conclusions</p> <p>The ChIP technology is a powerful tool for investigating transcription factors in vivo, especially in cancer biology. The established site-specific enzyme digestion enables a reliable and individual detection option for densely arranged binding motifs in vivo not provided by e.g. EMSA or in vivo footprinting. Given the important function of transcription factors in neoplastic mechanism, our method enables a broad diversity of application options for clinical studies.</p

Morphologic characterization of osteosarcoma growth on the chick chorioallantoic membrane

<p>Abstract</p> <p>Background</p> <p>The chick chorio-allantoic membrane (CAM) assay is a commonly used method for studying angiogenic or anti-angiogenic activities <it>in vivo</it>. The ease of access allows direct monitoring of tumour growth by biomicroscopy and the possibility to screen many samples in an inexpensive way. The CAM model provides a powerful tool to study effects of molecules, which interfere with physiological angiogenesis, or experimental tumours derived from cancer cell lines. We therefore screened eight osteosarcoma cell lines for their ability to form vascularized tumours on the CAM.</p> <p>Findings</p> <p>We implanted 3-5 million cells of human osteosarcoma lines (HOS, MG63, MNNG-HOS, OST, SAOS, SJSA1, U2OS, ZK58) on the CAM at day 10 of embryonic development. Tumour growth was monitored by <it>in vivo </it>biomicroscopy at different time points and tumours were fixed in paraformaldehyde seven days after cell grafting. The tissue was observed, photographed and selected cases were further analyzed using standard histology.</p> <p>From the eight cell lines the MNNG-HOS, U2OS and SAOS were able to form solid tumours when grafted on the CAM. The MNNG-HOS tumours showed the most reliable and consistent growth and were able to penetrate the chorionic epithelium, grow in the CAM stroma and induce a strong angiogenic response.</p> <p>Conclusions</p> <p>Our results show that the CAM assay is a useful tool for studying osteosarcoma growth. The model provides an excellent alternative to current rodent models and could serve as a preclinical screening assay for anticancer molecules. It might increase the speed and efficacy of the development of new drugs for the treatment of osteosarcoma.</p

A short-term in vivo model for giant cell tumor of bone

<p>Abstract</p> <p>Background</p> <p>Because of the lack of suitable <it>in vivo </it>models of giant cell tumor of bone (GCT), little is known about its underlying fundamental pro-tumoral events, such as tumor growth, invasion, angiogenesis and metastasis. There is no existing cell line that contains all the cell and tissue tumor components of GCT and thus <it>in vitro </it>testing of anti-tumor agents on GCT is not possible. In this study we have characterized a new method of growing a GCT tumor on a chick chorio-allantoic membrane (CAM) for this purpose.</p> <p>Methods</p> <p>Fresh tumor tissue was obtained from 10 patients and homogenized. The suspension was grafted onto the CAM at day 10 of development. The growth process was monitored by daily observation and photo documentation using <it>in vivo </it>biomicroscopy. After 6 days, samples were fixed and further analyzed using standard histology (hematoxylin and eosin stains), Ki67 staining and fluorescence <it>in situ </it>hybridization (FISH).</p> <p>Results</p> <p>The suspension of all 10 patients formed solid tumors when grafted on the CAM. <it>In vivo </it>microscopy and standard histology revealed a rich vascularization of the tumors. The tumors were composed of the typical components of GCT, including (CD51+/CD68+) multinucleated giant cells whichwere generally less numerous and contained fewer nuclei than in the original tumors. Ki67 staining revealed a very low proliferation rate. The FISH demonstrated that the tumors were composed of human cells interspersed with chick-derived capillaries.</p> <p>Conclusions</p> <p>A reliable protocol for grafting of human GCT onto the chick chorio-allantoic membrane is established. This is the first <it>in vivo </it>model for giant cell tumors of bone which opens new perspectives to study this disease and to test new therapeutical agents.</p

Analyzing the basic principles of tissue microarray data measuring the cooperative phenomena of marker proteins in invasive breast cancer

Background: The analysis of a protein-expression pattern from tissue microarray (TMA) data will not immediately give an answer on synergistic or antagonistic effects between the observed proteins. But contrary to apparent first impression, it is possible to reveal those cooperative phenomena from TMA data. The data is (1) preserving a lot of the original physiological information content and (2) because of minor variances between the tumor samples, contains several related slightly different biological states. We present here a largely assumption-free combinatorial analysis, related to correlation networks but with much less arbitrary constraints. A strong focus was put on the analysis of the basic data to analyze how the cooperative phenomena might be imprinted in the TMA data structure. Results: The study design was based on two independent panels of 589 and 366 invasive breast cancer cases from different institutions, assembled on tissue microarrays. The combinatorial analysis generates an optimal rank ordering of protein-expression coherence. The outcome of the analysis corresponds to all the single observations scattered over several publications and integrates them in one context. This means all these scattered observations can also be deduced from one TMA experiment. A comprehensive statistical meta-analysis of the TMA data suggests the existence of a superposition of three basic coherence situations, and offers the opportunity to analyze these data properties with additional real-world data and synthetic data in more detail. Conclusion: The presented algorithm gives molecular pathologists a tool to extract dependency information from TMA data. Beyond this practical benefit, some light was shed on how dependency aspects might be imprinted into expression data. This will certainly foster the refinement of algorithms to reconstruct dependency networks. The implementation of the algorithm is at the moment not end-user suitable, but available on request

Intraepidermal Nerve Fiber Density: Diagnostic and Therapeutic Relevance in the Management of Chronic Pruritus: a Review

<p><b>Article full text</b></p> <p><br></p> <p>The full text of this article can be found here<b>. </b><a href="https://link.springer.com/article/10.1007/s13555-016-0146-1">https://link.springer.com/article/10.1007/s13555-016-0146-1</a></p><p></p> <p><br></p> <p><b>Provide enhanced content for this article</b></p> <p><br></p> <p>If you are an author of this publication and would like to provide additional enhanced content for your article then please contact <a href="http://www.medengine.com/Redeem/”mailto:[email protected]”"><b>[email protected]</b></a>.</p> <p><br></p> <p>The journal offers a range of additional features designed to increase visibility and readership. All features will be thoroughly peer reviewed to ensure the content is of the highest scientific standard and all features are marked as ‘peer reviewed’ to ensure readers are aware that the content has been reviewed to the same level as the articles they are being presented alongside. Moreover, all sponsorship and disclosure information is included to provide complete transparency and adherence to good publication practices. This ensures that however the content is reached the reader has a full understanding of its origin. No fees are charged for hosting additional open access content.</p> <p><br></p> <p>Other enhanced features include, but are not limited to:</p> <p><br></p> <p>• Slide decks</p> <p>• Videos and animations</p> <p>• Audio abstracts</p> <p>• Audio slides</p