A novel RhoA/ROCK-CPI-17-MEF2C signaling pathway regulates vascular smooth muscle cell gene expression

Differentiation of vascular smooth muscle cells (VSMC) is a fundamental aspect of normal development and vascular disease. During contraction, VSMCs modulate calcium sensitivity through RhoA/ROCK-mediated inhibition of the myosin light chain phosphatase complex (MLCP). Previous studies have demonstrated that this signaling pathway functions in parallel to increase the expression of smooth muscle genes through the myocardin-family of co-activators. MEF2C fulfills a critical role in VSMC differentiation and regulates myocardin expression, leading us to investigate whether the RhoA/ROCK signaling cascade might regulate MEF2 activity. Depolarization-induced calcium signaling increased the expression of myocardin, which was sensitive to ROCK and p38 MAPK inhibition. We previously identified protein phosphatase 1α (PP1α), a known catalytic subunit of the MLCP in VSMCs, as a potent repressor of MEF2 activity. PP1α inhibition resulted in increased expression of myocardin, while ectopic expression of PP1α inhibited the induction of myocardin by MEF2C. Consistent with these data, shRNA-mediated suppression of a PP1α inhibitor, CPI-17, reduced myocardin expression and inhibited VSMC differentiation, suggesting a pivotal role for CPI-17 in regulating MEF2 activity. These data constitute evidence of a novel signaling cascade that links RhoA-mediated calcium sensitivity to MEF2-dependent myocardin expression in VSMCs through a mechanism involving p38 MAPK, PP1α, and CPI-17

Neighbours of cancer-related proteins have key influence on pathogenesis and could increase the drug target space for anticancer therapies

Even targeted chemotherapies against solid cancers show a moderate success increasing the need to novel targeting strategies. To address this problem, we designed a systems-level approach investigating the neighbourhood of mutated or differentially expressed cancer-related proteins in four major solid cancers (colon, breast, liver and lung). Using signalling and protein–protein interaction network resources integrated with mutational and expression datasets, we analysed the properties of the direct and indirect interactors (first and second neighbours) of cancer-related proteins, not found previously related to the given cancer type. We found that first neighbours have at least as high degree, betweenness centrality and clustering coefficient as cancer-related proteins themselves, indicating a previously unknown central network position. We identified a complementary strategy for mutated and differentially expressed proteins, where the affect of differentially expressed proteins having smaller network centrality is compensated with high centrality first neighbours. These first neighbours can be considered as key, so far hidden, components in cancer rewiring, with similar importance as mutated proteins. These observations strikingly suggest targeting first neighbours as a novel strategy for disrupting cancer-specific networks. Remarkably, our survey revealed 223 marketed drugs already targeting first neighbour proteins but applied mostly outside oncology, providing a potential list for drug repurposing against solid cancers. For the very central first neighbours, whose direct targeting would cause several side effects, we suggest a cancer-mimicking strategy by targeting their interactors (second neighbours of cancer-related proteins, having a central protein affecting position, similarly to the cancer-related proteins). Hence, we propose to include first neighbours to network medicine based approaches for (but not limited to) anticancer therapies

Overview of diagnosis and management of paediatric headache. Part I: diagnosis

Headache is the most common somatic complaint in children and adolescents. The evaluation should include detailed history of children and adolescents completed by detailed general and neurological examinations. Moreover, the possible role of psychological factors, life events and excessively stressful lifestyle in influencing recurrent headache need to be checked. The choice of laboratory tests rests on the differential diagnosis suggested by the history, the character and temporal pattern of the headache, and the physical and neurological examinations. Subjects who have any signs or symptoms of focal/progressive neurological disturbances should be investigated by neuroimaging techniques. The electroencephalogram and other neurophysiological examinations are of limited value in the routine evaluation of headaches. In a primary headache disorder, headache itself is the illness and headache is not attributed to any other disorder (e.g. migraine, tension-type headache, cluster headache and other trigeminal autonomic cephalgias). In secondary headache disorders, headache is the symptom of identifiable structural, metabolic or other abnormality. Red flags include the first or worst headache ever in the life, recent headache onset, increasing severity or frequency, occipital location, awakening from sleep because of headache, headache occurring exclusively in the morning associated with severe vomiting and headache associated with straining. Thus, the differential diagnosis between primary and secondary headaches rests mainly on clinical criteria. A thorough evaluation of headache in children and adolescents is necessary to make the correct diagnosis and initiate treatment, bearing in mind that children with headache are more likely to experience psychosocial adversity and to grow up with an excess of both headache and other physical and psychiatric symptoms and this creates an important healthcare problem for their future life

The Regulation of Beta-catenin Function in Vascular Smooth Muscle Cells

Vascular smooth muscle cells (VSMCs) maintain the ability to switch between a quiescent and proliferative phenotype based on external stimuli; this is an important feature for vascular repair. However, it leaves VSMCs predisposed to be affected by aberrant stimuli which results in improper cell proliferation, contributing to cardiovascular disease. Therefore, understanding the molecular mechanisms involved in activating VSMCs is of great clinical and therapeutic importance. -catenin is a multifunctional protein that plays an essential role in cell adhesion at the membrane and is required for Wnt dependent target gene expression in the nucleus. -catenin has been shown to play a significant role in VSMCs during both development and wound repair, and -catenins role is wholly dependent on specific protein-protein interactions in the cell, thus the aim of this project was to identify novel interacting partners and determine their effect on -catenin function in VSMCs. Our lab has previously identified the myocyte enhancer factor 2 (MEF2) family of transcription factors as potential -catenin interacting partners. The MEF2s are involved in vascular development during embryogenesis, but are also upregulated in the neointima following carotid artery injury in mice. In the first study, a direct interaction between MEF2 and -catenin was documented. This interaction was enhanced by p38 phosphorylation of MEF2, which resulted in increased nuclear retention of -catenin. Active p38 in primary VSMCs was able to stimulate Wnt signalling in a MEF2-dependent manner, and the MEF2--catenin interaction also synergistically increased cell proliferation. The purpose of the second study was to identify novel -catenin interacting partners in VSMCs using mass spectrometry. The investigation revealed 131 interactions, and gene ontology enrichment analysis emphasized translation as a significant biological process. Examination of one potential interactor, the fragile X mental retardation protein (FMRP), revealed a direct interaction with -catenin. siRNA silencing of -catenin increased global protein synthesis, and biochemical analysis determined that -catenin associates at the pre-initiation complex with FMRP. Wnt-dependent nuclear shuttling of -catenin de-repressed protein translation, and we concluded that cytoplasmic -catenin can inhibit translation. Thus, we established a completely novel role for -catenin function in the regulation of translation by interaction with FMRP