GLOBAL PROTEOME AND PHOSPHOPROTEOME ANALYSIS OF MENINGIOMAS.

INTRODUCTIONMeningiomas are common brain tumours arising from meningeal tissue. Despite the majority of them displaying benign features, they can cause mild to severe morbidity. The current main therapeutic approach is complete tumour resection commonly with adjunct radiation therapy. However, tumour location can hamper complete resection and chemotherapies are ineffective. In this study we aim to elucidate dysregulated pathways in meningioma pathogenesis and identify novel molecular targets by deciphering the proteome and phosphoproteome of different grades of meningiomas.METHODSTumour lysates were collected from grade I, II and III frozen meningioma specimens and three normal healthy human meninges. Phosphoprotein purification was performed using Qiagen® PhosphoProtein Purification Kit. Proteins were separated by SDS-PAGE followed by in-gel tryptic digestion. Extracted peptides were purified and analysed by electrospray ionization LC-MS/MS. Raw mass spectrometry files were analysed using MaxQuantTM. Expression data was validated by Western blot and functionally annotated using Ingenuity® Pathway Analysis (IPA®) and DAVID 6.8.RESULTSWe have quantified 3888 proteins and 3074 phosphoproteins across all grades of meningioma and normal meninges. Comparative analysis confirmed 181 proteins and 338 phosphoproteins to be commonly significantly upregulated (log2 fold-change≥1.5; p\u3c0.05) among all grades vs. normal meninges. We have successfully validated the expression data of several upregulated proteins and phosphoproteins. Gene Ontology revealed biological processes including EFGR and VEGFR signalling to be enriched in the phosphoproteome. Grade-wise comparisons identified 667 proteins and 769 phosphoproteins to be differentially expressed (p\u3c0.05) between grade III meningiomas compared to grade II and I.CONCLUSIONWe have performed a comparative proteomic analysis across all meningioma grades and identified changes in proteomic profiles between these tumours and normal healthy meninges. We will use this data to define novel targets common to all grades and specific to a grade of meningioma for future therapies

Constitutive activation of the EGFR-STAT1 axis increases proliferation of meningioma tumor cells.

Background: Meningiomas are the most frequent primary brain tumors of the central nervous system. The standard of treatment is surgery and radiotherapy, but effective pharmacological options are not available yet. The well-characterized genetic background stratifies these tumors in several subgroups, thus increasing diversification. We identified epidermal growth factor receptor-signal transducer and activator of transcription 1 (EGFR-STAT1) overexpression and activation as a common identifier of these tumors. Methods: We analyzed STAT1 overexpression and phosphorylation in 131 meningiomas of different grades and locations by utilizing several techniques, including Western blots, qPCR, and immunocytochemistry. We also silenced and overexpressed wild-type and mutant forms of the gene to assess its biological function and its network. Results were further validated by drug testing. Results: STAT1 was found widely overexpressed in meningioma but not in the corresponding healthy controls. The protein showed constitutive phosphorylation not dependent on the JAK-STAT pathway. STAT1 knockdown resulted in a significant reduction of cellular proliferation and deactivation of AKT and ERK1/2. STAT1 is known to be activated by EGFR, so we investigated the tyrosine kinase and found that EGFR was also constitutively phosphorylated in meningioma and was responsible for the aberrant phosphorylation of STAT1. The pharmaceutical inhibition of EGFR caused a significant reduction in cellular proliferation and of overall levels of cyclin D1, pAKT, and pERK1/2. Conclusions: STAT1-EGFR-dependent constitutive phosphorylation is responsible for a positive feedback loop that causes its own overexpression and consequently an increased proliferation of the tumor cells. These findings provide the rationale for further studies aiming to identify effective therapeutic options in meningioma

Potential influences of complement factor H in autoimmune inflammatory and thrombotic disorders

Complement system homeostasis is important for host self-protection and anti-microbial immune surveillance, and recent research indicates roles in tissue development and remodelling. Complement also appears to have several points of interaction with the blood coagulation system. Deficiency and altered function due to gene mutations and polymorphisms in complement effectors and regulators, including Factor H, has been associated with familial and sporadic autoimmune inflammatory - thrombotic disorders, in which autoantibodies play a part. These include systemic lupus erythematosus, rheumatoid arthritis, atypical haemolytic uremic syndrome, anti-phospholipid syndrome and age-related macular degeneration. Such diseases are generally complex – multigenic and heterogeneous in their symptoms and predisposition/susceptibility. They usually need to be triggered by vascular trauma, drugs or infection and non-complement genetic factors also play a part. Underlying events seem to include decline in peripheral regulatory T cells, dendritic cell, and B cell tolerance, associated with alterations in lymphoid organ microenvironment. Factor H is an abundant protein, synthesised in many cell types, and its reported binding to many different ligands, even if not of high affinity, may influence a large number of molecular interactions, outwith the accepted role of Factor H within the complement system. Factor H is involved in mesenchymal stem cell mediated tolerance and also contributes to self tolerance by augmenting iC3b production and opsonisation of apoptotic cells for their silent dendritic cell engulfment via complement receptor CR3, which mediates anti-inflammatory tolerogenic effects in the apoptotic cell context. There may be co-operation with other phagocytic receptors, such as complement C1q receptors, and the Tim glycoprotein family, which specifically bind phosphatidylserine expressed on the apoptotic cell surface. Factor H is able to discriminate between self and nonself surfaces for self-protection and anti-microbe defence. Factor H, particularly as an abundant platelet protein, may also modulate blood coagulation, having an anti-thrombotic role. Here we review a number of interaction pathways in coagulation and in immunity, together with associated diseases, and indicate where Factor H may be expected to exert an influence, based on reports of the diversity of ligands for Factor H

Hantavirus Infection in the Republic of Georgia

We describe a laboratory-confirmed case of hantavirus infection in the Republic of Georgia. Limited information is available about hantavirus infections in the Caucasus, although the infection has been reported throughout Europe and Russia. Increasing awareness and active disease surveillance contribute to our improved understanding of the geographic range of this pathogen

Effects of dust on light-curves of \epsilon Aur type stars

\epsilon Auriga is one of the most mysterious objects on the sky. Prior modeling of its light-curve assumed a dark, inclined, non-transparent or semi-transparent, dusty disk with a central hole. The hole was necessary to explain the light-curve with a sharp mid-eclipse brightening. The aim of the present paper is to study the effects of dust on the light-curves of eclipsing binary stars and to develop an alternative physical model for $\epsilon$ Aur type objects which is based on the optical properties of dust grains. The code Shellspec has been modified to calculate the light-curves and spectra of such objects. The code solves the radiative transfer along the line of sight in interacting binaries. Dust and angle dependent Mie scattering were introduced into the code for this purpose. Our model of $\epsilon$ Aur consists of two geometrically thick flared disks: an internal optically thick disk and an external optically thin disk which absorbs and scatters radiation. Disks are in the orbital plane and are almost edge-on. We argue that there is no need for an inclined disk with a hole to explain the current eclipse of $\epsilon$ Aur not even if there is a possible shallow mid-eclipse brightening. It was demonstrated that phase dependent light scattering and the optical properties of the dust can have an important effect on the light-curves of such stars and can even produce a mid-eclipse brightening. This is a natural consequence of the strong forward scattering. It was also demonstrated that shallow mid-eclipse brightening might result from eclipses by nearly edge-on flared (dusty or gaseous) disks.Comment: A&A Letter, accepted 4 pages, 3 figure