Machine Learning of Stem Cell Identities From Single-Cell Expression Data via Regulatory Network Archetypes

The molecular regulatory network underlying stem cell pluripotency has been intensively studied, and we now have a reliable ensemble model for the “average” pluripotent cell. However, evidence of significant cell-to-cell variability suggests that the activity of this network varies within individual stem cells, leading to differential processing of environmental signals and variability in cell fates. Here, we adapt a method originally designed for face recognition to infer regulatory network patterns within individual cells from single-cell expression data. Using this method we identify three distinct network configurations in cultured mouse embryonic stem cells—corresponding to naïve and formative pluripotent states and an early primitive endoderm state—and associate these configurations with particular combinations of regulatory network activity archetypes that govern different aspects of the cell's response to environmental stimuli, cell cycle status and core information processing circuitry. These results show how variability in cell identities arise naturally from alterations in underlying regulatory network dynamics and demonstrate how methods from machine learning may be used to better understand single cell biology, and the collective dynamics of cell communities

Quantification of intracellular payload release from polymersome nanoparticles

Polymersome nanoparticles (PMs) are attractive candidates for spatio-temporal controlled delivery of therapeutic agents. Although many studies have addressed cellular uptake of solid nanoparticles, there is very little data available on intracellular release of molecules encapsulated in membranous carriers, such as polymersomes. Here, we addressed this by developing a quantitative assay based on the hydrophilic dye, fluorescein. Fluorescein was encapsulated stably in PMs of mean diameter 85 nm, with minimal leakage after sustained dialysis. No fluorescence was detectable from fluorescein PMs, indicating quenching. Following incubation of L929 cells with fluorescein PMs, there was a gradual increase in intracellular fluorescence, indicating PM disruption and cytosolic release of fluorescein. By combining absorbance measurements with flow cytometry, we quantified the real-time intracellular release of a fluorescein at a single-cell resolution. We found that 173 ± 38 polymersomes released their payload per cell, with significant heterogeneity in uptake, despite controlled synchronisation of cell cycle. This novel method for quantification of the release of compounds from nanoparticles provides fundamental information on cellular uptake of nanoparticle-encapsulated compounds. It also illustrates the stochastic nature of population distribution in homogeneous cell populations, a factor that must be taken into account in clinical use of this technology.</p

Single cell analyses and machine learning define hematopoietic progenitor and HSC-like cells derived from human PSCs

Haematopoietic stem and progenitor cells (HSPCs) develop through distinct waves at various anatomical sites during embryonic development. The in vitro differentiation of human pluripotent stem cells (hPSCs) is able to recapitulate some of these processes, however, it has proven difficult to generate functional haematopoietic stem cells (HSCs). To define the dynamics and heterogeneity of HSPCs that can be generated in vitro from hPSCs, we exploited single cell RNA sequencing (scRNAseq) in combination with single cell protein expression analysis. Bioinformatics analyses and functional validation defined the transcriptomes of naïve progenitors as well as erythroid, megakaryocyte and leukocyte-committed progenitors and we identified CD44, CD326, ICAM2/CD9 and CD18 as markers of these progenitors, respectively. Using an artificial neural network (ANN), that we trained on a scRNAseq derived from human fetal liver, we were able to identify a wide range of hPSCs-derived HPSC phenotypes, including a small group classified as HSCs. This transient HSC-like population decreased as differentiation proceeded and was completely missing in the dataset that had been generated using cells selected on the basis of CD43expression. By comparing the single cell transcriptome of in vitro-generated HSC-like cells with those generated within the fetal liver we identified transcription factors and molecular pathways that can be exploited in the future to improve the in vitro production of HSCs

Measurement of triple gauge boson couplings from WW production at LEP energies up to 189 GeV

A measurement of triple gauge boson couplings is presented, based on W-pair data recorded by the OPAL detector at LEP during 1998 at a centre-of-mass energy of 189 GeV with an integrated luminosity of 183 pb^-1. After combining with our previous measurements at centre-of-mass energies of 161-183 GeV we obtain k_g=0.97 +0.20 -0.16, g_1^z=0.991 +0.060 -0.057 and lambda_g=-0.110 +0.058 -0.055, where the errors include both statistical and systematic uncertainties and each coupling is determined by setting the other two couplings to their SM values. These results are consistent with the Standard Model expectations.Comment: 28 pages, 8 figures, submitted to Eur. Phys. J.

Search for Yukawa Production of a Light Neutral Higgs Boson at LEP

Within a Two-Higgs-Doublet Model (2HDM) a search for a light Higgs boson in the mass range of 4-12 GeV has been performed in the Yukawa process e+e- -> b bbar A/h -> b bbar tau+tau-, using the data collected by the OPAL detector at LEP between 1992 and 1995 in e+e- collisions at about 91 GeV centre-of-mass energy. A likelihood selection is applied to separate background and signal. The number of observed events is in good agreement with the expected background. Within a CP-conserving 2HDM type II model the cross-section for Yukawa production depends on xiAd = |tan beta| and xihd = |sin alpha/cos beta| for the production of the CP-odd A and the CP-even h, respectively, where tan beta is the ratio of the vacuum expectation values of the Higgs doublets and alpha is the mixing angle between the neutral CP-even Higgs bosons. From our data 95% C.L. upper limits are derived for xiAd within the range of 8.5 to 13.6 and for xihd between 8.2 to 13.7, depending on the mass of the Higgs boson, assuming a branching fraction into tau+tau- of 100%. An interpretation of the limits within a 2HDM type II model with Standard Model particle content is given. These results impose constraints on several models that have been proposed to explain the recent BNL measurement of the muon anomalous magnetic moment.Comment: 24 pages, 9 figures, Submitted to Euro. Phys. J.

Measurement of the Hadronic Cross-Section for the Scattering of Two Virtual Photons at LEP

The interaction of virtual photons is investigated using the reaction e+e- -> e+e- hadrons based on data taken by the OPAL experiment at e+e- centre-of-mass energies sqrt(s_ee)=189-209 GeV, for W>5 GeV and at an average Q^2 of 17.9 GeV^2. The measured cross-sections are compared to predictions of the Quark Parton Model (QPM), to the Leading Order QCD Monte Carlo model PHOJET to the NLO prediction for the reaction e+e- -> e+e-qqbar, and to BFKL calculations. PHOJET, NLO e+e- -> e+e-qqbar, and QPM describe the data reasonably well, whereas the cross-section predicted by a Leading Order BFKL calculation is too large.Comment: 30 pages, 10 figures, Submitted to Eur.Phys.J.

Measurement of the B0 Lifetime and Oscillation Frequency using B0->D*+l-v decays

The lifetime and oscillation frequency of the B0 meson has been measured using B0->D*+l-v decays recorded on the Z0 peak with the OPAL detector at LEP. The D*+ -> D0pi+ decays were reconstructed using an inclusive technique and the production flavour of the B0 mesons was determined using a combination of tags from the rest of the event. The results t_B0 = 1.541 +- 0.028 +- 0.023 ps, Dm_d = 0.497 +- 0.024 +- 0.025 ps-1 were obtained, where in each case the first error is statistical and the second systematic.Comment: 17 pages, 4 figures, submitted to Phys. Lett.

WW Production Cross Section and W Branching Fractions in e+e- Collisions at 189 GeV

From a data sample of 183 pb^-1 recorded at a center-of-mass energy of roots = 189 GeV with the OPAL detector at LEP, 3068 W-pair candidate events are selected. Assuming Standard Model W boson decay branching fractions, the W-pair production cross section is measured to be sigmaWW = 16.30 +- 0.34(stat.) +- 0.18(syst.) pb. When combined with previous OPAL measurements, the W boson branching fraction to hadrons is determined to be 68.32 +- 0.61(stat.) +- 0.28(syst.) % assuming lepton universality. These results are consistent with Standard Model expectations.Comment: 22 pages, 5 figures, submitted to Phys. Lett.

Genuine Correlations of Like-Sign Particles in Hadronic Z0 Decays

Correlations among hadrons with the same electric charge produced in Z0 decays are studied using the high statistics data collected from 1991 through 1995 with the OPAL detector at LEP. Normalized factorial cumulants up to fourth order are used to measure genuine particle correlations as a function of the size of phase space domains in rapidity, azimuthal angle and transverse momentum. Both all-charge and like-sign particle combinations show strong positive genuine correlations. One-dimensional cumulants initially increase rapidly with decreasing size of the phase space cells but saturate quickly. In contrast, cumulants in two- and three-dimensional domains continue to increase. The strong rise of the cumulants for all-charge multiplets is increasingly driven by that of like-sign multiplets. This points to the likely influence of Bose-Einstein correlations. Some of the recently proposed algorithms to simulate Bose-Einstein effects, implemented in the Monte Carlo model PYTHIA, are found to reproduce reasonably well the measured second- and higher-order correlations between particles with the same charge as well as those in all-charge particle multiplets.Comment: 26 pages, 6 figures, Submitted to Phys. Lett.