Dissecting cell identity via network inference and in silico gene perturbation

Cell identity is governed by the complex regulation of gene expression, represented as gene-regulatory network

Gene regulatory network reconfiguration in direct lineage reprogramming

In direct lineage conversion, transcription factor (TF) overexpression reconfigures gene regulatory networks (GRNs) to reprogram cell identity. We previously developed CellOracle, a computational method to infer GRNs from single-cell transcriptome and epigenome data. Using inferred GRNs, CellOracle simulates gene expression changes in response to TF perturbation, enabling in silico interrogation of network reconfiguration. Here, we combine CellOracle analysis with lineage tracing of fibroblast to induced endoderm progenitor (iEP) conversion, a prototypical direct reprogramming paradigm. By linking early network state to reprogramming outcome, we reveal distinct network configurations underlying successful and failed fate conversion. Via in silico simulation of TF perturbation, we identify new factors to coax cells into successfully converting their identity, uncovering a central role for the AP-1 subunit Fos with the Hippo signaling effector, Yap1. Together, these results demonstrate the efficacy of CellOracle to infer and interpret cell-type-specific GRN configurations, providing new mechanistic insights into lineage reprogramming

NHS Greater Glasgow & Clyde Acute Pharmacy Redesign Program : Report for the Pharmacy and Prescribing Support Unit, NHS Greater Glasgow and Clyde

NHS Greater Glasgow and Clyde are in the midst of a major pharmacy redesign programme that aims to maximise the application of technology in the medicines supply chain and to release staff to deliver improved patient care through the Making the Most of your Medicines (MMyM) service. This report discusses the findings from a study undertaken by a team of researchers from the University of Strathclyde and the Pharmacy and Prescribing Support Unit (PPSU) in support of this redesign programme. In particular, the study focuses on the implementation of a new approach to in-patient medicines management designed by the PPSU. This approach takes the form of a robotic pharmacy distribution system, installed in a newly-built centrally-located Pharmacy Distribution Centre (PDC). The study was conducted from January to September 2010. The aims of the study were, first, to develop a suitable metrics framework for the new pharmacy distribution system and, second, to capture the organisational learning gained from the implementation phase of the PDC. However, as the project progressed it became clear that the primary focus would be on capturing organisational learning and providing expert advice to support implementation before a more effective performance measurement system could be designed. The report makes three sets of recommendations relating to: (1) standardising processes, improving quality and sharing best practice; (2) improving staff morale; (3) analysing and improving inventory management procedures. These recommendations are complemented by a proposal for a new, multi-layered performance measurement framework. This would consist of a Balanced Scorecard for strategic control (quarterly, monthly), into which feeds an Operational Dashboard (weekly, daily, all of which would be underpinned by a Lean Six Sigma improvement framework (incorporating FMEA and HACCP techniques where possible)

Barcoded oligonucleotides ligated on RNA amplified for multiplexed and parallel in situ analyses.

We present barcoded oligonucleotides ligated on RNA amplified for multiplexed and parallel insitu analyses (BOLORAMIS), a reverse transcription-free method for spatially-resolved, targeted, in situ RNA identification of single or multiple targets. BOLORAMIS was demonstrated on a range of cell types and human cerebral organoids. Singleplex experiments to detect coding and non-coding RNAs in human iPSCs showed a stem-cell signature pattern. Specificity of BOLORAMIS was found to be 92% as illustrated by a clear distinction between human and mouse housekeeping genes in a co-culture system, as well as by recapitulation of subcellular localization of lncRNA MALAT1. Sensitivity of BOLORAMIS was quantified by comparing with single molecule FISH experiments and found to be 11%, 12% and 35% for GAPDH, TFRC and POLR2A, respectively. To demonstrate BOLORAMIS for multiplexed gene analysis, we targeted 96 mRNAs within a co-culture of iNGN neurons and HMC3 human microglial cells. We used fluorescence in situ sequencing to detect error-robust 8-base barcodes associated with each of these genes. We then used this data to uncover the spatial relationship among cells and transcripts by performing single-cell clustering and gene-gene proximity analyses. We anticipate the BOLORAMIS technology for in situ RNA detection to find applications in basic and translational research

BT-benzo-29 inhibits bacterial cell proliferation by perturbing FtsZ assembly

We have identified a potent antibacterial agent N-(4-sec-butylphenyl)-2-(thiophen-2-yl)-1H-benzod]imidazole-4-carboxa mide (BT-benzo-29) from a library of benzimidazole derivatives that stalled bacterial division by inhibiting FtsZ assembly. A short (5 min) exposure of BT-benzo-29 disassembled the cytokinetic Z-ring in Bacillus subtilis cells without affecting the cell length and nucleoids. BT-benzo-29 also perturbed the localization of early and late division proteins such as FtsA, ZapA and SepF at the mid-cell. Further, BT-benzo-29 bound to FtsZ with a dissociation constant of 24 +/- 3 m and inhibited the assembly and GTPase activity of purified FtsZ. A docking analysis suggested that BT-benzo-29 may bind to FtsZ at the C-terminal domain near the T7 loop. BT-benzo-29 displayed significantly weaker inhibitory effects on the assembly and GTPase activity of two mutants (L272A and V275A) of FtsZ supporting the prediction of the docking analysis. Further, BT-benzo-29 did not appear to inhibit DNA duplication and nucleoid segregation and it did not perturb the membrane potential of B. subtilis cells. The results suggested that BT-benzo-29 exerts its potent antibacterial activity by inhibiting FtsZ assembly. Interestingly, BT-benzo-29 did not affect the membrane integrity of mammalian red blood cells. BT-benzo-29 bound to tubulin with a much weaker affinity than FtsZ and exerted significantly weaker effects on mammalian cells than on the bacterial cells indicating that the compound may have a strong antibacterial potential

Comparative evaluation of the effectiveness of audio and audiovisual distraction aids in the management of anxious pediatric dental patients

Objective: The aim of this study was to evaluate and compare audio and audiovisual distraction aids in management of anxious pediatric dental patients of different age groups and to study children′s response to sequential dental visits with the use of distraction aids. Study Design: This study was conducted on two age groups, that is, 4-6 years and 6-8 years with 30 patients in each age group on their first dental visit. The children of both the age groups were divided into 3 subgroups, the control group, audio distraction group, audiovisual distraction group with 10 patients in each subgroup. Each child in all the subgroups had gone through three dental visits. Child anxiety level at each visit was assessed by using a combination of anxiety measuring parameters. The data collected was tabulated and subjected to statistical analysis. Results: Tukey honest significant difference post-hoc test at 0.05% level of significance revealed audiovisual group showed statistically highly significant difference from audio and control group, whereas audio group showed the statistically significant difference from the control group. Conclusion: Audiovisual distraction was found to be a more effective mode of distraction in the management of anxious children in both the age groups when compared to audio distraction. In both the age groups, a significant effect of the visit type was also observed

Robotics within the medicines supply chain in NHS Scotland – results from a national survey : report for the directors of pharmacy, NHS Scotland

This report provides the results from a national survey on robotics within the medicines supply chain in NHS Scotlan