Stochastic Simulation of Process Calculi for Biology

Biological systems typically involve large numbers of components with complex, highly parallel interactions and intrinsic stochasticity. To model this complexity, numerous programming languages based on process calculi have been developed, many of which are expressive enough to generate unbounded numbers of molecular species and reactions. As a result of this expressiveness, such calculi cannot rely on standard reaction-based simulation methods, which require fixed numbers of species and reactions. Rather than implementing custom stochastic simulation algorithms for each process calculus, we propose to use a generic abstract machine that can be instantiated to a range of process calculi and a range of reaction-based simulation algorithms. The abstract machine functions as a just-in-time compiler, which dynamically updates the set of possible reactions and chooses the next reaction in an iterative cycle. In this short paper we give a brief summary of the generic abstract machine, and show how it can be instantiated with the stochastic simulation algorithm known as Gillespie's Direct Method. We also discuss the wider implications of such an abstract machine, and outline how it can be used to simulate multiple calculi simultaneously within a common framework.Comment: In Proceedings MeCBIC 2010, arXiv:1011.005

Modelling the Dynamics of an Aedes albopictus Population

We present a methodology for modelling population dynamics with formal means of computer science. This allows unambiguous description of systems and application of analysis tools such as simulators and model checkers. In particular, the dynamics of a population of Aedes albopictus (a species of mosquito) and its modelling with the Stochastic Calculus of Looping Sequences (Stochastic CLS) are considered. The use of Stochastic CLS to model population dynamics requires an extension which allows environmental events (such as changes in the temperature and rainfalls) to be taken into account. A simulator for the constructed model is developed via translation into the specification language Maude, and used to compare the dynamics obtained from the model with real data.Comment: In Proceedings AMCA-POP 2010, arXiv:1008.314

Perspectives of Statistician, Microbiologist, and Clinician Stakeholders on the Use of Microbiological Outcomes in Randomised Trials of Antimicrobial Stewardship Interventions

Microbiological data are used as indicators of infection, for diagnosis, and the identification of antimicrobial resistance in trials of antimicrobial stewardship interventions. However, several problems have been identified in a recently conducted systematic review (e.g., inconsistency in reporting and oversimplified outcomes), which motivates the need to understand and improve the use of these data including analysis and reporting. We engaged key stakeholders including statisticians, clinicians from both primary and secondary care, and microbiologists. Discussions included issues identified in the systematic review and questions about the value of using microbiological data in clinical trials, perspectives on current microbiological outcomes reported in trials, and alternative statistical approaches to analyse these data. Various factors (such as unclear sample collection process, dichotomising or categorising complex microbiological data, and unclear methods of handling missing data) were identified that contributed to the low quality of the microbiological outcomes and the analysis of these outcomes in trials. Whilst not all of these factors would be easy to overcome, there is room for improvement and a need to encourage researchers to understand the impact of misusing these data. This paper discusses the experience and challenges of using microbiological outcomes in clinical trials

Efficient generation of vesicular stomatitis virus (VSV)-pseudotypes bearing morbilliviral glycoproteins and their use in quantifying virus neutralising antibodies

Morbillivirus neutralising antibodies are traditionally measured using either plaque reduction neutralisation tests (PRNTs) or live virus microneutralisation tests (micro-NTs). While both test formats provide a reliable assessment of the strength and specificity of the humoral response, they are restricted by the limited number of viral strains that can be studied and often present significant biological safety concerns to the operator. In this study, we describe the adaptation of a replication-defective vesicular stomatitis virus (VSVΔG) based pseudotyping system for the measurement of morbillivirus neutralising antibodies. By expressing the haemagglutinin (H) and fusion (F) proteins of canine distemper virus (CDV) on VSVΔG pseudotypes bearing a luciferase marker gene, neutralising antibody titres could be measured rapidly and with high sensitivity. Further, by exchanging the glycoprotein expression construct, responses against distinct viral strains or species may be measured. Using this technique, we demonstrate cross neutralisation between CDV and peste des petits ruminants virus (PPRV). As an example of the value of the technique, we demonstrate that UK dogs vary in the breadth of immunity induced by CDV vaccination; in some dogs the neutralising response is CDV-specific while, in others, the neutralising response extends to the ruminant morbillivirus PPRV. This technique will facilitate a comprehensive comparison of cross-neutralisation to be conducted across the morbilliviruses

Statistical Model Checking for Stochastic Hybrid Systems

This paper presents novel extensions and applications of the UPPAAL-SMC model checker. The extensions allow for statistical model checking of stochastic hybrid systems. We show how our race-based stochastic semantics extends to networks of hybrid systems, and indicate the integration technique applied for implementing this semantics in the UPPAAL-SMC simulation engine. We report on two applications of the resulting tool-set coming from systems biology and energy aware buildings.Comment: In Proceedings HSB 2012, arXiv:1208.315

Regulation of pH by Carbonic Anhydrase 9 Mediates Survival of Pancreatic Cancer Cells With Activated KRAS in Response to Hypoxia.

Background & Aims Most pancreatic ductal adenocarcinomas (PDACs) express an activated form of KRAS, become hypoxic and dysplastic, and are refractory to chemo and radiation therapies. To survive in the hypoxic environment, PDAC cells upregulate enzymes and transporters involved in pH regulation, including the extracellular facing carbonic anhydrase 9 (CA9). We evaluated the effect of blocking CA9, in combination with administration of gemcitabine, in mouse models of pancreatic cancer. Methods We knocked down expression of KRAS in human (PK-8 and PK-1) PDAC cells with small hairpin RNAs. Human and mouse (KrasG12D/Pdx1-Cre/Tp53/RosaYFP) PDAC cells were incubated with inhibitors of MEK (trametinib) or extracellular signal-regulated kinase (ERK), and some cells were cultured under hypoxic conditions. We measured levels and stability of the hypoxia-inducible factor 1 subunit alpha (HIF1A), endothelial PAS domain 1 protein (EPAS1, also called HIF2A), CA9, solute carrier family 16 member 4 (SLC16A4, also called MCT4), and SLC2A1 (also called GLUT1) by immunoblot analyses. We analyzed intracellular pH (pHi) and extracellular metabolic flux. We knocked down expression of CA9 in PDAC cells, or inhibited CA9 with SLC-0111, incubated them with gemcitabine, and assessed pHi, metabolic flux, and cytotoxicity under normoxic and hypoxic conditions. Cells were also injected into either immune-compromised or immune-competent mice and growth of xenograft tumors was assessed. Tumor fragments derived from patients with PDAC were surgically ligated to the pancreas of mice and the growth of tumors was assessed. We performed tissue microarray analyses of 205 human PDAC samples to measure levels of CA9 and associated expression of genes that regulate hypoxia with outcomes of patients using the Cancer Genome Atlas database. Results Under hypoxic conditions, PDAC cells had increased levels of HIF1A and HIF2A, upregulated expression of CA9, and activated glycolysis. Knockdown of KRAS in PDAC cells, or incubation with trametinib, reduced the posttranscriptional stabilization of HIF1A and HIF2A, upregulation of CA9, pHi, and glycolysis in response to hypoxia. CA9 was expressed by 66% of PDAC samples analyzed; high expression of genes associated with metabolic adaptation to hypoxia, including CA9, correlated with significantly reduced survival times of patients. Knockdown or pharmacologic inhibition of CA9 in PDAC cells significantly reduced pHi in cells under hypoxic conditions, decreased gemcitabine-induced glycolysis, and increased their sensitivity to gemcitabine. PDAC cells with knockdown of CA9 formed smaller xenograft tumors in mice, and injection of gemcitabine inhibited tumor growth and significantly increased survival times of mice. In mice with xenograft tumors grown from human PDAC cells, oral administration of SLC-0111 and injection of gemcitabine increased intratumor acidosis and increased cell death. These tumors, and tumors grown from PDAC patient-derived tumor fragments, grew more slowly than xenograft tumors in mice given control agents, resulting in longer survival times. In KrasG12D/Pdx1-Cre/Tp53/RosaYFP genetically modified mice, oral administration of SLC-0111 and injection of gemcitabine reduced numbers of B cells in tumors. Conclusions In response to hypoxia, PDAC cells that express activated KRAS increase expression of CA9, via stabilization of HIF1A and HIF2A, to regulate pH and glycolysis. Disruption of this pathway slows growth of PDAC xenograft tumors in mice and might be developed for treatment of pancreatic cancer

Sloan Digital Sky Survey Imaging of Low Galactic Latitude Fields: Technical Summary and Data Release

The Sloan Digital Sky Survey (SDSS) mosaic camera and telescope have obtained five-band optical-wavelength imaging near the Galactic plane outside of the nominal survey boundaries. These additional data were obtained during commissioning and subsequent testing of the SDSS observing system, and they provide unique wide-area imaging data in regions of high obscuration and star formation, including numerous young stellar objects, Herbig-Haro objects and young star clusters. Because these data are outside the Survey regions in the Galactic caps, they are not part of the standard SDSS data releases. This paper presents imaging data for 832 square degrees of sky (including repeats), in the star-forming regions of Orion, Taurus, and Cygnus. About 470 square degrees are now released to the public, with the remainder to follow at the time of SDSS Data Release 4. The public data in Orion include the star-forming region NGC 2068/NGC 2071/HH24 and a large part of Barnard's loop.Comment: 31 pages, 9 figures (3 missing to save space), accepted by AJ, in press, see http://photo.astro.princeton.edu/oriondatarelease for data and paper with all figure

Effect of promoter architecture on the cell-to-cell variability in gene expression

According to recent experimental evidence, the architecture of a promoter, defined as the number, strength and regulatory role of the operators that control the promoter, plays a major role in determining the level of cell-to-cell variability in gene expression. These quantitative experiments call for a corresponding modeling effort that addresses the question of how changes in promoter architecture affect noise in gene expression in a systematic rather than case-by-case fashion. In this article, we make such a systematic investigation, based on a simple microscopic model of gene regulation that incorporates stochastic effects. In particular, we show how operator strength and operator multiplicity affect this variability. We examine different modes of transcription factor binding to complex promoters (cooperative, independent, simultaneous) and how each of these affects the level of variability in transcription product from cell-to-cell. We propose that direct comparison between in vivo single-cell experiments and theoretical predictions for the moments of the probability distribution of mRNA number per cell can discriminate between different kinetic models of gene regulation.Comment: 35 pages, 6 figures, Submitte