Uncovering distinct protein-network topologies in heterogeneous cell populations

Background: Cell biology research is fundamentally limited by the number of intracellular components, particularly proteins, that can be co-measured in the same cell. Therefore, cell-to-cell heterogeneity in unmeasured proteins can lead to completely different observed relations between the same measured proteins. Attempts to infer such relations in a heterogeneous cell population can yield uninformative average relations if only one underlying biochemical network is assumed. To address this, we developed a method that recursively couples an iterative unmixing process with a Bayesian analysis of each unmixed subpopulation. Results: Our approach enables to identify the number of distinct cell subpopulations, unmix their corresponding observations and resolve the network structure of each subpopulation. Using simulations of the MAPK pathway upon EGF and NGF stimulations we assess the performance of the method. We demonstrate that the presented method can identify better than clustering approaches the number of subpopulations within a mixture of observations, thus resolving correctly the statistical relations between the proteins. Conclusions: Coupling the unmixing of multiplexed observations with the inference of statistical relations between the measured parameters is essential for the success of both of these processes. Here we present a conceptual and algorithmic solution to achieve such coupling and hence to analyze data obtained from a natural mixture of cell populations. As the technologies and necessity for multiplexed measurements are rising in the systems biology era, this work addresses an important current challenge in the analysis of the derived data.Fil: Wieczorek, Jakob. Universitat Dortmund; AlemaniaFil: Malik Sheriff, Rahuman S.. Institut Max Planck fur Molekulare Physiologie; Alemania. Imperial College London; Reino Unido. European Bioinformatics Institute. European Molecular Biology Laboratory; Reino UnidoFil: Fermin, Yessica. Universitat Dortmund; AlemaniaFil: Grecco, Hernan Edgardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Física de Buenos Aires. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Física de Buenos Aires; Argentina. Institut Max Planck fur Molekulare Physiologie; AlemaniaFil: Zamir, Eli. Institut Max Planck fur Molekulare Physiologie; AlemaniaFil: Ickstadt, Katja. Universitat Dortmund; Alemani

On the tightness of Marton's regions for semi-additive broadcast channels

We study cost constrained side-information channels, where the cost function depends on a state which is known only to the encoder. In the additive noise case, we bound the capacity loss due to not knowing the cost state at the decoder and show that it is small under various assumptions, and goes to zero in the limit of weak noise. This model plays an important role in the (non-degraded) broadcast channel. In the semi-additive noise case, we bound the gap between the best known single letter achievable region and the true capacity region, using tools developed for the first problem. In the limit of weak noise, we show that the bounds coincide, thus we get the complete characterization of the capacity region

Pulmonary function correlates with arterial stiffness in asthmatic patients

SummaryBackgroundAt the population level, asthma has been associated with chronic systemic inflammation as well as adverse cardiovascular outcomes.ObjectivesThe aim of this study was to investigate peripheral vascular hemodynamic variables of arterial stiffness (AS) and their relationship to pulmonary function tests in asthmatic patients.MethodsYoung asthmatic patients from the tertiary center for pulmonary diseases at the Barzilai Medical Center underwent pulmonary function evaluation and non-invasive radial artery hemodynamic profiling, pre- and post-exercise. Results were compared to age matched, non-asthmatic controls.Results23 young asthmatics and 41 controls, completed all evaluation points. Pulmonary flow parameters were significantly reduced in the asthma group at all points. There were no differences between groups in BMI, blood pressure, pulse rate or measurements of AS at baseline or after bronchodilation. The % predicted forced expiratory volume in the first second at baseline (FEV1%) in asthmatics was positively correlated with the small arteries elasticity index (SAEI) and negatively correlated with the systemic vascular resistance (SVR) in these patients. These correlations were not observed in non-asthmatic controls. In multifactorial regression FEV1 remained the major factor associated with measurements of AS in asthmatic patients, while gender was the only significant factor in non-asthmatic controls.ConclusionsSignificant correlations between measurements of AS and FEV1 in young asthmatics, suggest the presence of a common systemic, most likely inflammatory pathway involving both the cardiovascular and respiratory systems

Symmetric exchange of multi-protein building blocks between stationary focal adhesions and the cytosol

How can the integrin adhesome get self-assembled locally, rapidly, and correctly as diverse cell-matrix adhesion sites? Here, we investigate this question by exploring the cytosolic state of integrin-adhesome components and their dynamic exchange between adhesion sites and cytosol. Using fluorescence cross-correlation spectroscopy (FCCS) and fluorescence recovery after photobleaching (FRAP) we found that the integrin adhesome is extensively pre-assembled already in the cytosol as multi-protein building blocks for adhesion sites. Stationary focal adhesions release symmetrically the same types of protein complexes that they recruit, thereby keeping the cytosolic pool of building blocks spatiotemporally uniform. We conclude a model in which multi-protein building blocks enable rapid and modular self-assembly of adhesion sites and symmetric exchange of these building blocks preserves their specifications and thus the assembly logic of the system

Utilization of Murine Laparoscopy for Continuous In-Vivo Assessment of the Liver in Multiple Disease Models

BACKGROUND: Current strategies for follow up of murine models of liver disease are flawed by inability to continuously monitor disease progression in the tissue level, and necessitate sacrifice of animals for tissue sampling. AIMS: In this study we aimed at developing a safe repetitive tool for sampling livers in vivo, by utilization of a miniaturized endoscopy system for laparoscopic liver biopsies and for injection of tumor cells into livers. RESULTS: We report the development of a protocol for murine laparoscopy that allows repeated visualization of murine intra-abdominal organs. The system enables safe and repeated liver biopsies in mice and rats, yielding adequate tissue for histological staining and RNA extraction. In addition, injection of tumor cells into livers facilitates under-vision implantation of hepatic tumors in liver, followed by visualization of tumor growth. CONCLUSIONS: Murine laparoscopy may be employed as a novel imaging modality for continuous assessment and manipulation of chronic liver disease models

Focal Contacts as Mechanosensors: Externally Applied Local Mechanical Force Induces Growth of Focal Contacts by an Mdia1-Dependent and Rock-Independent Mechanism

The transition of cell–matrix adhesions from the initial punctate focal complexes into the mature elongated form, known as focal contacts, requires GTPase Rho activity. In particular, activation of myosin II–driven contractility by a Rho target known as Rho-associated kinase (ROCK) was shown to be essential for focal contact formation. To dissect the mechanism of Rho-dependent induction of focal contacts and to elucidate the role of cell contractility, we applied mechanical force to vinculin-containing dot-like adhesions at the cell edge using a micropipette. Local centripetal pulling led to local assembly and elongation of these structures and to their development into streak-like focal contacts, as revealed by the dynamics of green fluorescent protein–tagged vinculin or paxillin and interference reflection microscopy. Inhibition of Rho activity by C3 transferase suppressed this force-induced focal contact formation. However, constitutively active mutants of another Rho target, the formin homology protein mDia1 (Watanabe, N., T. Kato, A. Fujita, T. Ishizaki, and S. Narumiya. 1999. Nat. Cell Biol. 1:136–143), were sufficient to restore force-induced focal contact formation in C3 transferase-treated cells. Force-induced formation of the focal contacts still occurred in cells subjected to myosin II and ROCK inhibition. Thus, as long as mDia1 is active, external tension force bypasses the requirement for ROCK-mediated myosin II contractility in the induction of focal contacts. Our experiments show that integrin-containing focal complexes behave as individual mechanosensors exhibiting directional assembly in response to local force

Quantitative Multicolor Compositional Imaging Resolves Molecular Domains in Cell-Matrix Adhesions

Background: Cellular processes occur within dynamic and multi-molecular compartments whose characterization requires analysis at high spatio-temporal resolution. Notable examples for such complexes are cell-matrix adhesion sites, consisting of numerous cytoskeletal and signaling proteins. These adhesions are highly variable in their morphology, dynamics, and apparent function, yet their molecular diversity is poorly defined. Methodology/Principal Findings: We present here a compositional imaging approach for the analysis and display of multicomponent compositions. This methodology is based on microscopy-acquired multicolor data, multi-dimensional clustering of pixels according to their composition similarity and display of the cellular distribution of these composition clusters. We apply this approach for resolving the molecular complexes associated with focal-adhesions, and the time-dependent effects of Rho-kinase inhibition. We show here compositional variations between adhesion sites, as well as ordered variations along the axis of individual focal-adhesions. The multicolor clustering approach also reveals distinct sensitivities of different focaladhesion-associated complexes to Rho-kinase inhibition. Conclusions/Significance: Multicolor compositional imaging resolves ‘‘molecular signatures’ ’ characteristic to focaladhesions and related structures, as well as sub-domains within these adhesion sites. This analysis enhances the spatial information with additional ‘‘contents-resolved’ ’ dimensions. We propose that compositional imaging can serve as

The Critical Role of Chemokine (C–C Motif) Receptor 2-Positive Monocytes in Autoimmune Cholangitis

The therapy of primary biliary cholangitis (PBC) has lagged behind other autoimmune diseases despite significant improvements in our understanding of both immunological and molecular events that lead to loss of tolerance to the E2 component of pyruvate dehydrogenase, the immunodominant autoepitope of PBC. It is well known that Ly6Chi monocytes are innate immune cells infiltrating inflammatory sites that are dependent on the expression of C–C motif chemokine receptor 2 (CCR2) for emigration from bone marrow. Importantly, humans with PBC have a circulating monocyte pro-inflammatory phenotype with macrophage accumulation in portal tracts. We have taken advantage of an inducible chemical xenobiotic model of PBC and recapitulated the massive infiltration of monocytes to portal areas. To determine the clinical significance, we immunized both CCR2-deficient mice and controls with 2OA-BSA and noted that CCR2 deficiency is protective for the development of autoimmune cholangitis. Importantly, because of the therapeutic potential, we focused on inhibiting monocyte infiltration through the use of cenicriviroc (CVC), a dual chemokine receptor CCR2/CCR5 antagonist shown to be safe in human trials. Importantly, treatment with CVC resulted in amelioration of all aspects of disease severity including serum total bile acids, histological severity score, and fibrosis stage. In conclusion, our results indicate a major role for Ly6Chi monocytes and for CCR2 in PBC pathogenesis and suggest that inhibition of this axis by CVC should be explored in humans through the use of clinical trials