Functional expression of the ATP-gated P2X7 receptor in human iPSC-derived astrocytes.

Activation of the ATP-gated P2X7 receptor (P2X7R), implicated in numerous diseases of the brain, can trigger diverse responses such as the release of pro-inflammatory cytokines, modulation of neurotransmission, cell proliferation or cell death. However, despite the known species-specific differences in its pharmacological properties, to date, most functional studies on P2X7R responses have been analyzed in cells from rodents or immortalised cell lines. To assess the endogenous and functional expression of P2X7Rs in human astrocytes, we differentiated human-induced pluripotent stem cells (hiPSCs) into GFAP and S100 ß-expressing astrocytes. Immunostaining revealed prominent punctate P2X7R staining. P2X7R protein expression was also confirmed by Western blot. Importantly, stimulation with the potent non-selective P2X7R agonist 2',3'-O-(benzoyl-4-benzoyl)-adenosine 5'- triphosphate (BzATP) or endogenous agonist ATP induced robust calcium rises in hiPSC-derived astrocytes which were blocked by the selective P2X7R antagonists AFC-5128 or JNJ-47965567. Our findings provide evidence for the functional expression of P2X7Rs in hiPSC-derived astrocytes and support their in vitro utility in investigating the role of the P2X7R and drug screening in disorders of the central nervous system (CNS).

Cheminformatics at the interface of medicinal chemistry and proteomics.

Multiple factors have to be optimized in the course of a drug discovery project. Traditionally this includes potency on a single target, eventually specificity as well as the pharmacokinetic, physicochemical and the safety profile. Recently an additional dimension has been added by realizing that the therapeutic outcome of a drug is often determined not only by its activity on a single target but also by its activity profile across a variety of biological targets. To address the polypharmacology of drug candidates many compounds are tested on a set of targets or in phenotypic screens generating a tremendous amount of data. To extract useful information computational methods at the interface of proteomics and cheminformatics are indispensable. This review will focus on some recent developments in this field. This article is part of a Special Issue entitled: Computational Proteomics in the Post-Identification Era. Guest Editors: Martin Eisenacher and Christian Stephan.

Increased expression of the ATP-gated P2X7 receptor reduces responsiveness to anti-convulsants during status epilepticus in mice.

BACKGROUND AND PURPOSE: Refractory status epilepticus is a clinical emergency associated with high mortality and morbidity. Increasing evidence suggests neuroinflammation contributes to the development of drug-refractoriness during status epilepticus. Here, we have determined the contribution of the ATP-gated P2X7 receptor, previously linked to inflammation and increased hyperexcitability, to drug-refractory status epilepticus and its therapeutic potential. EXPERIMENTAL APPROACH: Status epilepticus was induced via a unilateral microinjection of kainic acid into the amygdala in adult mice. Severity of status epilepticus was compared in animals with overexpressing or knock-out of the P2X7 receptor, after inflammatory priming by pre-injection of bacterial lipopolysaccharide (LPS) and in mice treated with P2X7 receptor-targeting and anti-inflammatory drugs. KEY RESULTS: Mice overexpressing P2X7 receptors were unresponsive to several anticonvulsants (lorazepam, midazolam, phenytoin and carbamazepine) during status epilepticus. P2X7 receptor expression increased in microglia during status epilepticus, at times when responses to anticonvulsants were reduced. Overexpression of P2X7 receptors induced a pro-inflammatory phenotype in microglia during status epilepticus and the anti-inflammatory drug minocycline restored normal responses to anticonvulsants in mice overexpressing P2X7 receptors. Pretreatment of wild-type mice with LPS increased P2X7 receptor levels in the brain and reduced responsiveness to anticonvulsants during status epilepticus, which was overcome by either genetic deletion of P2X7 receptors or treatment with the P2X7 receptor antagonists, AFC-5128 or ITH15004. CONCLUSION AND IMPLICATIONS: Our results demonstrate that P2X7 receptor-induced pro-inflammatory effects contribute to resistance to pharmacotherapy during status epilepticus. Therapies targeting P2X7 receptors could be novel adjunctive treatments for drug-refractory status epilepticus.

Cell-based proteome analysis: the first stage in the pipeline for biomarker discovery.

The early detection of a distinct disease is crucial for a successful treatment and depends on a sensitive as well as specific diagnosis. In last years tremendous attempts were undertaken to identify new biomarker applying proteomics, but no relevant candidate has been identified for clinical application. Although proteomics is enabling quantitative and qualitative analysis of proteins within complex mixtures it could not significantly contribute to this field. Therefore, different proteomics approaches have been established focusing on the direct analysis of cell populations involved in pathogenic processes to identify candidate biomarkers even for in vitro diagnosis. Here, we will outline approaches applying cell- and tissue based proteome analysis as the first decisive step in the pipeline for the discovery of new diagnostic biomarkers. We will show examples for analysing precursor lesions of the pancreatic ductal adenocarcinoma (PDAC), nephron glomeruli and fibrotic and non-fibrotic liver tissue. This article provides also an overview about currently available techniques in the field of cell enrichment and quantitative proteome analysis of lowest amounts of sample.

Chaos-on-a-chip secures data transmission in optical fiber links.

Security in information exchange plays a central role in the deployment of modern communication systems. Besides algorithms, chaos is exploited as a real-time high-speed data encryption technique which enhances the security at the hardware level of optical networks. In this work, compact, fully controllable and stably operating monolithic photonic integrated circuits (PICs) that generate broadband chaotic optical signals are incorporated in chaos-encoded optical transmission systems. Data sequences with rates up to 2.5 Gb/s with small amplitudes are completely encrypted within these chaotic carriers. Only authorized counterparts, supplied with identical chaos generating PICs that are able to synchronize and reproduce the same carriers, can benefit from data exchange with bit-rates up to 2.5Gb/s with error rates below 10(-12). Eavesdroppers with access to the communication link experience a 0.5 probability to detect correctly each bit by direct signal detection, while eavesdroppers supplied with even slightly unmatched hardware receivers are restricted to data extraction error rates well above 10(-3).

Inside SMP proteomics: six years German Human Brain Proteome Project (HBPP) - a summary.

In 2001, the German Federal Ministry of Education and Research (BMBF) initiated the National Genome Research Network (NGFN; www.ngfn.de) as a nation-wide multidisciplinary networking platform aiming at the analysis of common human diseases and aging. Within the NGFN the Human Brain Proteome Project (HBPP; www.smp-proteomics.de) focuses on the analysis of the human brain in health and disease. The concept is based on two consecutive steps: (i) Elaborating and establishing the necessary technology platforms. (ii) Application of the established technologies for research in Alzheimer's disease and Parkinson's disease. In the first funding period, HBPP1, running from 2001 to 2004, necessary technologies were established and optimized. In HBPP2, which started 2004 and will end in May 2008, the developed technologies are used for large-scale experiments, offering new links for disease related research and therapies. The following overview describes structure, aims and outcome of this unique German Brain Proteome Project.

Proteomics Data Collection - 2nd ProDaC Workshop 5 October 2007, Seoul, Korea.

Proteomics Data Collection (ProDaC) is an EU-funded "Coordination Action" within the 6th framework programme. It aims to simplify the publication, dissemination and utilization of proteomics data by establishing standards that will support broad data collection from the research community. As a part of ProDaC, regular workshops are organized on a half-yearly basis to enable communication and discussion of the involved partners and to report on project progress. After the kick-off meeting (October 2006) in Long Beach, CA, USA and the 1st workshop in Lyon, France (April 2007), the 2nd ProDaC workshop took place at the COEX InterContinental Hotel in Seoul, Korea, on 5th October 2007, shortly before the HUPO World Congress. The progress achieved within the first year was presented by the leaders of the work packages. Additionally, a Journal's representative talked about his experiences and future plans concerning Proteomics standards; and two further external speakers presented their research related to data handling and Proteomics repositories.

Proteomics Data Collection - 3rd ProDaC workshop April 22nd 2008, Toledo, Spain.

The "Coordination Action" ProDaC (Proteomics Data Collection) - funded by the EU within the 6th framework programme - was created to support the dissemination, utilization and publication of proteomics data. Within this international consortium, standards are developed and maintained to support extensive data collection by the proteomics community. An important part of ProDaC are workshops organized on a regular basis (two per year) to allow discussions and communication between the ProDaC partners and to report on the progress of the project. The kick-off meeting took place in October 2006 in Long Beach, CA, USA. The 1st ProDaC workshop was held in Lyon, France (April 2007) and the 2nd in Seoul, Korea in October 2007. ProDaC organized the 3rd ProDaC workshop at the Beatriz Hotel, Toledo, on 22nd April, 2008, directly before the HUPO - PSI spring meeting (Human Proteome Organisation - Proteomics Standards Initiative). The work package coordinators presented talks about the progress achieved during the past six months. Additionally four external speakers presented their work on data conversion and data repositories. The concluding discussion session was chaired by the Journal's representative.

Applications in brain proteomics: 8(th) HUPO Brain Proteome Project Workshop 7 October 2007, Seoul, Korea.

What are the current approaches in brain proteomics? Can we combine different, but complementary study designs to obtain better results concerning brain diseases? What are the neuro-hotspots, especially in Korea? These were some of the questions the participants of the 8(th) HUPO Brain Proteome Project Workshop tried to answer prior to the 6(th) HUPO World Congress in Seoul, Korea. Around 100 scientists came together during the afternoon of 7 October, 2007, to discuss and to catch up on the latest results and strategies concerning Huntington's disease, glioblastoma and standardization.

The HUPO Brain Proteome project wish list--summary of the 9(th) HUPO BPP Workshop 9-10 January 2008, Barbados.

The Human Brain Proteome Project (HUPO BPP) aims at advancing knowledge and the understanding of neurodiseases and aging with the purpose of identifying prognostic and diagnostic biomarkers, as well as to push new diagnostic approaches and medications. The participating groups meet in semi-annual workshops to discuss the progress, as well as the needs, within the field of proteomics. The 9(th) HUPO BPP workshop took place in Barbados from 9-10 January, 2008. Discussing the future HUPO BPP Roadmap, the attendees drafted the so called HUPO BPP wish list containing timelines, suggestions and missions. This wish list will be updated regularly and will serve as a guideline for the next phase.

Maintaining standardization: an update of the HUPO Brain Proteome Project.

The Human Proteome Organisation was launched in February 2001 as a result of the need of an international proteomic forum to improve the understanding of human diseases. The initiative dealing with the brain is the Human Proteome Organisation Brain Proteome Project chaired in Germany and Korea. In order to estimate the existing approaches in brain proteomics, as well as to establish a standardized data reprocessing pipeline, pilot studies were initiated including both mouse and human samples. Data had to be submitted to a Data Collection Center for central re-processing and are now publicly accessible at the PRIDE database serving as reference data for future analysis. It became clear that heterogeneity, for example, different analysis strategies and data formats, are a real challenge when comparing results and when working in a consortium. Therefore, standardization, the organisation of data management and the synergistic effects of a consortium of collaborators are of outstanding importance to any big proteome analysis. The following manuscript will highlight these activities and aims of the Human Proteome Organisation Brain Proteome Project, summarizing its historical timeline and its two pilot studies.

How proteomics reveals potential biomarkers in brain diseases.

The brain is complex, and so are the proteomics studies of brain tissue and its diseases, including Alzheimer's Disease, Parkinson's Disease and schizophrenia. In this review, general considerations and strategies of proteomics technologies, the advantages and challenges as well as the special needs for brain tissue are described and summarized. In addition, the results of the first studies are presented including a quality evaluation of the candidate proteins for these diseases. A paragraph is dedicated to the efforts of standardization in this field.

Inter-lab proteomics: data mining in collaborative projects on the basis of the HUPO brain proteome project's pilot studies.

Several projects were initiated by the Human Proteome Organisation (HUPO) focusing on the proteome analysis of distinct human organs. The initiative dedicated to the brain, its development and correlated diseases is the HUPO Brain Proteome Project (HUPO BPP). An objective data submission, storage, and reprocessing strategy have been established with the help of the results gained in a pilot study phase and within subsequent studies. The bioinformatic relevance of the data is drawn from the inter-laboratory comparisons as well as from the recalculation of all data sets submitted by the different groups. In the following, results of the single groups as well as the centralised reprocessing effort are summarised, demonstrating the added-value of this concerted work.