P. M. W. French

Imperial College London, Londinium, England, United Kingdom

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Publications (332)640.4 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Fully differentiated pancreatic β cells are essential for normal glucose homeostasis in mammals. Dedifferentiation of these cells has been suggested to occur in type 2 diabetes, impairing insulin production. Since chronic fuel excess ("glucotoxicity") is implicated in this process, we sought here to identify the potential roles in β-cell identity of the tumor suppressor liver kinase B1 (LKB1/STK11) and the downstream fuel-sensitive kinase, AMP-activated protein kinase (AMPK). Highly β-cell-restricted deletion of each kinase in mice, using an Ins1-controlled Cre, was therefore followed by physiological, morphometric, and massive parallel sequencing analysis. Loss of LKB1 strikingly (2.0-12-fold, E<0.01) increased the expression of subsets of hepatic (Alb, Iyd, Elovl2) and neuronal (Nptx2, Dlgap2, Cartpt, Pdyn) genes, enhancing glutamate signaling. These changes were partially recapitulated by the loss of AMPK, which also up-regulated β-cell "disallowed" genes (Slc16a1, Ldha, Mgst1, Pdgfra) 1.8- to 3.4-fold (E<0.01). Correspondingly, targeted promoters were enriched for neuronal (Zfp206; P=1.3×10(-33)) and hypoxia-regulated (HIF1; P=2.5×10(-16)) transcription factors. In summary, LKB1 and AMPK, through only partly overlapping mechanisms, maintain β-cell identity by suppressing alternate pathways leading to neuronal, hepatic, and other characteristics. Selective targeting of these enzymes may provide a new approach to maintaining β-cell function in some forms of diabetes.-Kone, M., Pullen, T. J., Sun, G., Ibberson, M., Martinez-Sanchez, A., Sayers, S., Nguyen-Tu, M.-S., Kantor, C., Swisa, A., Dor, Y., Gorman, T., Ferrer, J., Thorens, B., Reimann, F., Gribble, F., McGinty, J. A., Chen, L., French, P. M., Birzele, F., Hildebrandt, T., Uphues, I., Rutter, G. A. LKB1 and AMPK differentially regulate pancreatic β-cell identity.
    FASEB journal : official publication of the Federation of American Societies for Experimental Biology. 07/2014;
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    ABSTRACT: We image core-shell nanoparticles, consisting of a dye-doped silica core covered with a layer of gold, with a STED-FLIM microscope. Due to the field enhancement provided by the localised surface plasmon resonance of the gold shell, we demonstrate a reduction of the STED depletion power required to obtain resolution improvement by a factor of four. This validates the concept of nanoparticle-assisted STED (NP-STED), where hybrid dye-plasmonic nanoparticles are used as labels for STED in order to decrease the depletion powers required for sub-wavelength imaging.
    Nano letters. 07/2014;
  • Paul M. French
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    ABSTRACT: I will review our development and application of fluorescence lifetime imaging implemented in microscopy, tomography and endoscopy to provide molecular readouts across the scales from super-resolved microscopy through imaging of disease models to clinical applications.
    CLEO: Science and Innovations; 06/2014
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    ABSTRACT: : Fluorescence spectroscopy is a promising tool for the characterisation of biological tissues and shows the potential for in vivo clinical diagnosis in many diseases. It exploits the inherent photo-physical properties of a number of endogenous molecules. Fluorescence lifetime measurements of these fluorophores such as NADH and flavoproteins present an opportunity to discern functional information concerning myocardial energetics without issues of toxicity or systemic effects associated with the introduction of exogenous compounds. Additionally, autofluorescence from extra-cellular matrix molecules e.g. collagen may provide information on structural changes to the heart. Measurements of fluorescence lifetime are independent of fluorophore concentration, excitation intensity, sample attenuation and other experimental artefacts and can also report on changes to the fluorophore microenvironment e.g. pH and protein binding state. Thus we are interested to develop autofluorescence lifetime (AFL)-based techniques as a myocardial "optical biopsy". We report the application of a custom fibre-optic probe-based time-resolved spectrofluorometer utilising time-correlated single photon counting that we developed to characterise the autofluorescence signatures associated with the histological, morphological, metabolic and functional changes in myocardial tissue in health and disease states. We studied an in vivo rat left anterior descending coronary artery ligation heart failure (MI-HF) model 16 weeks post-infarction, which is a model well characterised in our institution. We observed stable AFL signals across age-matched control (AMC) hearts (n = 6) in different anatomical locations. We observed significant differences in AFL signals between MI-HF (n = 6) and AMC (n = 6) in infarcted regions: left ventricle (LV) anterior wall (p < 0.0001) and "border zone" region (p < 0.0001). We also observed significant differences between MI-HF and AMC in remodelled regions distant to the infarct: LV posterior wall (p < 0.01) and right ventricle (RV) (p < 0.001). Application of principal component analysis and linear discriminant analysis to the data facilitated development of a diagnostic algorithm to differentiate MI-HF and AMC in a given anatomical location with a high degree of accuracy both in infarcted regions (specificity 100%, sensitivity 100%) and even in remote regions e.g. RV (specificity 96%, sensitivity 89%). This represents, to the best of our knowledge, the first in vivo application of time-resolved fluorescence spectroscopy to the study of the heart. This technique shows promise as a diagnostic tool and could be readily and rapidly translatable into clinical cardiology practice.
    Heart (British Cardiac Society) 06/2014; 100(Suppl 3):A104. · 5.01 Impact Factor
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    ABSTRACT: Purpose: To correct for attenuation in two-photon fluorescence (TPF) measurements of riboflavin absorption in porcine corneas. Methods: TPF imaging of riboflavin was performed using excitation at a wavelength of 890nm, with fluorescence signal detected between 525-650nm. TPF signal attenuation was demonstrated by imaging from either side of a uniformly soaked corneo-scleral button. To overcome this attenuation, a reservoir of dextran-free 0.1% riboflavin 5'-monophosphate in saline and hydroxypropylmethylcellulose (HPMC) was placed on top of porcine corneas (globe intact-epithelium removed). TPF imaging was performed through this reservoir with image stacks acquired at 10µm steps through the cornea repeated at regular intervals for up to 60 minutes. A novel correction method was applied to achieve corneal riboflavin concentration measurements in whole eyes (n=4). Results: Significant attenuation of the TPF signal was observed in all eyes, with the signal decreasing approximately linearly with depth in uniformly soaked tissue. Cross-sectional TPF images taken of excised corneal strips confirmed the tissue was uniformly soaked so that the decrease in signal was not due to spatial variations in riboflavin concentration. After correcting for signal attenuation, we observed increased riboflavin concentrations with longer soak duration, with a mean (±SD) maximum tissue concentration of 0.094 (±0.001)% [1.36mg/ml]. Uniform riboflavin absorption was achieved after a minimum 50 minutes. Following a 30 minute soak, a mean stromal concentration of 0.086 (±0.001)% [1.25mg/ml] was achieved at a depth of 300µm. Conclusions: The accuracy of TPF measurements of corneal riboflavin absorption can be increased by applying a correction for depth-related signal attenuation.
    Investigative ophthalmology & visual science 03/2014; · 3.43 Impact Factor
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    ABSTRACT: Degradation of articular cartilage extracellular matrix (ECM) by proteolytic enzyme is the hallmark of arthritis that leads to joint destruction. Detection of early biochemical changes in cartilage before irreversible structural damages become apparent is highly desirable. Here we report that the autofluorescence decay profile of cartilage is significantly affected by proteolytic degradation of cartilage ECM and can be characterised by measurements of the autofluorescence lifetime (AFL). A multidimensional fluorometer utilizing ultraviolet excitation at 355 nm or 375 nm coupled to a fibreoptic probe was developed for single point time-resolved AFL measurements of porcine articular cartilage explants treated with different proteinases. Degradation of cartilage matrix components by treating with bacterial collagenase, matrix metalloproteinase 1, or trypsin resulted in significant reduction of AFL of the cartilage in both a dose and time dependent manner. Differences in cartilage AFL were also confirmed by fluorescence lifetime imaging microscopy (FLIM). Our data suggest that AFL of cartilage tissue is a potential non-invasive readout to monitor cartilage matrix integrity that may be utilized for diagnosis of arthritis as well as monitoring the efficacy of anti-arthritic therapeutic agents.
    02/2014;
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    ABSTRACT: Fluorescence lifetime imaging (FLIM) has previously been shown to provide contrast between normal and diseased tissue. Here we present progress towards clinical and preclinical FLIM endoscopy of tissue autofluorescence, demonstrating a flexible wide-field endoscope that utilised a low average power blue picosecond laser diode excitation source and was able to acquire ∼mm-scale spatial maps of autofluorescence lifetimes from fresh ex vivo diseased human larynx biopsies in ∼8 seconds using an average excitation power of ∼0.5 mW at the specimen. To illustrate its potential for FLIM at higher acquisition rates, a higher power mode-locked frequency doubled Ti:Sapphire laser was used to demonstrate FLIM of ex vivo mouse bowel at up to 2.5 Hz using 10 mW of average excitation power at the specimen. (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).
    Journal of Biophotonics 02/2014; · 3.10 Impact Factor
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    ABSTRACT: The editors introduce the Biomedical Optics Express feature issue "Optical Molecular Probes, Imaging, and Drug Delivery," which is associated with a Topical Meeting of the same name held at the 2013 Optical Society of America (OSA) Optics in the Life Sciences Congress in Waikoloa Beach, Hawaii, April 14-18, 2013. The international meeting focused on the convergence of optical physics, photonics technology, nanoscience, and photochemistry with drug discovery and clinical medicine. Papers in this feature issue are representative of meeting topics, including advances in microscopy, nanotechnology, and optics in cancer research.
    Biomedical Optics Express 02/2014; 5(2):643-4. · 3.18 Impact Factor
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    ABSTRACT: We present an ex vivo study of temporally and spectrally resolved autofluorescence in a total of 47 endoscopic excision biopsy/resection specimens from colon, using pulsed excitation laser sources operating at wavelengths of 375 nm and 435 nm. A paired analysis of normal and neoplastic (adenomatous polyp) tissue specimens obtained from the same patient yielded a significant difference in the mean spectrally averaged autofluorescence lifetime -570 ± 740 ps (p = 0.021, n = 12). We also investigated the fluorescence signature of non-neoplastic polyps (n = 6) and inflammatory bowel disease (n = 4) compared to normal tissue in a small number of specimens.
    Biomedical Optics Express 02/2014; 5(2):515-38. · 3.18 Impact Factor
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    ABSTRACT: The forkhead box transcription factor FOXM1 is an essential effector of G2/M-phase transition, mitosis and the DNA damage response. As such, it is frequently deregulated during tumorigenesis. Here we report that FOXM1 is dynamically modified by SUMO1 but not by SUMO2/3 at multiple sites. We show that FOXM1 SUMOylation is enhanced in MCF-7 breast cancer cells in response to treatment with epirubicin and mitotic inhibitors. Mutation of five consensus conjugation motifs yielded a SUMOylation-deficient mutant FOXM1. Conversely, fusion of the E2 ligase Ubc9 to FOXM1 generated an auto-SUMOylating mutant (FOXM1-Ubc9). Analysis of wild-type FOXM1 and mutants revealed that SUMOylation inhibits FOXM1 activity, promotes translocation to the cytoplasm and enhances APC/Cdh1-mediated ubiquitination and degradation. Further, expression of the SUMOylation-deficient mutant enhanced cell proliferation compared with wild-type FOXM1, whereas the FOXM1-Ubc9 fusion protein resulted in persistent cyclin B1 expression and slowed the time from mitotic entry to exit. In summary, our findings suggest that SUMOylation attenuates FOXM1 activity and causes mitotic delay in cytotoxic drug response.Oncogene advance online publication, 23 December 2013; doi:10.1038/onc.2013.546.
    Oncogene 12/2013; · 7.36 Impact Factor
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    ABSTRACT: We describe a multicore endoscope fibre with minimised group index variation between cores obtained at a V parameter of 3. A spun fibre design enables the effects of bending to be reduced.
    Workshop on Specialty Optical Fibers and their Applications; 08/2013
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    ABSTRACT: We present a stimulated emission depletion (STED) microscope that provides 3-D super resolution by simultaneous depletion using beams with both a helical phase profile for enhanced lateral resolution and an annular phase profile to enhance axial resolution. The 3-D depletion point spread function is realised using a single spatial light modulator that can also be programmed to compensate for aberrations in the microscope and the sample. We apply it to demonstrate the first 3-D super-resolved imaging of an immunological synapse between a Natural Killer cell and its target cell. (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).
    Journal of Biophotonics 06/2013; · 3.10 Impact Factor
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    ABSTRACT: We describe a multicore endoscope fibre with minimised group index variation between cores that is obtained at a V parameter of 3. Tapering the fibre input enables us to achieve single-mode propagation.
    CLEO: Science and Innovations; 06/2013
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    ABSTRACT: We demonstrate the application of Fluorescence Lifetime Imaging (FLIM) to read out Förster resonant energy transfer (FRET) based biosensors for studying the spatio-temporal dynamics of signalling pathways in cells undergoing chemotaxis.
    Optical Molecular Probes, Imaging and Drug Delivery; 04/2013
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    ABSTRACT: We report the development and application of instrumentation to measure and image tissue autofluorescence lifetime for the study and diagnosis of disease including cancer and osteoarthritis.
    Optical Molecular Probes, Imaging and Drug Delivery; 04/2013
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    ABSTRACT: We report an automated fluorescence lifetime imaging multiwell plate reader for high content analysis, capable of subcellular mapping of protein interactions. This instrument can acquire FLIM data from 96 wells in less than 15 minutes.
    Optical Molecular Probes, Imaging and Drug Delivery; 04/2013
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    ABSTRACT: We describe an angular multiplexing technique for optical projection tomography that improves resolution, signal-to-noise ratio, and imaging speed by ameliorating the trade-off between spatial resolution and depth of field and improving the light collection efficiency. Here we demonstrate that imaging at two orthogonal angular projections simultaneously, focused on shifted planes in the sample, improves the average spatial resolution by ∼20% and the light collection efficiency by a factor of ∼4, thereby enabling increased acquisition speed and reduced light dose.
    Optics Letters 03/2013; 38(6):851-3. · 3.39 Impact Factor
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    ABSTRACT: Multiphoton laser tomography (MPT) combined with fluorescence lifetime imaging (FLIM) is a non-invasive imaging technique, based on the study of fluorescence decay times of naturally occurring fluorescent molecules, enabling a non-invasive investigation of the skin with subcellular resolution. The aim of this retrospective observational ex vivo study, was to characterize melanoma both from a morphologic and a quantitative point of view, attaining an improvement in the diagnostic accuracy with respect to dermoscopy. In the training phase, thirty parameters, comprising both cytological descriptors and architectural aspects, were identified. The training set included 6 melanomas with a mean Breslow thickness±S.D. of 0.89±0.48 mm. In the test phase, these parameters were blindly evaluated on a test data set consisting of 25 melanomas, 50 nevi and 50 basal cell carcinomas. Melanomas in the test phase comprised 8 in situ lesions and had a mean thickness±S.D. of 0.77±1.2 mm. Moreover, quantitative FLIM data were calculated for special areas of interest. Melanoma was characterized by the presence of atypical short lifetime cells and architectural disorder, in contrast to nevi presenting typical cells and a regular histoarchitecture. Sensitivity and specificity values for melanoma diagnosis were 100% and 98%, respectively, whereas dermoscopy achieved the same sensitivity, but a lower specificity (82%). Mean fluorescence lifetime values of melanocytic cells did not vary between melanomas and nevi, but significantly differed from those referring to basal cell carcinoma enabling a differential diagnosis based on quantitative data. Data from prospective preoperative trials are needed to confirm if MPT/FLIM could increase diagnostic specificity and thus reduce unnecessary surgical excisions.
    PLoS ONE 01/2013; 8(7):e70682. · 3.73 Impact Factor
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    ABSTRACT: Fluorescence lifetime imaging (FLIM) is widely applied to obtain quantitative information from fluorescence signals, particularly using Förster Resonant Energy Transfer (FRET) measurements to map, for example, protein-protein interactions. Extracting FRET efficiencies or population fractions typically entails fitting data to complex fluorescence decay models but such experiments are frequently photon constrained, particularly for live cell or in vivo imaging, and this leads to unacceptable errors when analysing data on a pixel-wise basis. Lifetimes and population fractions may, however, be more robustly extracted using global analysis to simultaneously fit the fluorescence decay data of all pixels in an image or dataset to a multi-exponential model under the assumption that the lifetime components are invariant across the image (dataset). This approach is often considered to be prohibitively slow and/or computationally expensive but we present here a computationally efficient global analysis algorithm for the analysis of time-correlated single photon counting (TCSPC) or time-gated FLIM data based on variable projection. It makes efficient use of both computer processor and memory resources, requiring less than a minute to analyse time series and multiwell plate datasets with hundreds of FLIM images on standard personal computers. This lifetime analysis takes account of repetitive excitation, including fluorescence photons excited by earlier pulses contributing to the fit, and is able to accommodate time-varying backgrounds and instrument response functions. We demonstrate that this global approach allows us to readily fit time-resolved fluorescence data to complex models including a four-exponential model of a FRET system, for which the FRET efficiencies of the two species of a bi-exponential donor are linked, and polarisation-resolved lifetime data, where a fluorescence intensity and bi-exponential anisotropy decay model is applied to the analysis of live cell homo-FRET data. A software package implementing this algorithm, FLIMfit, is available under an open source licence through the Open Microscopy Environment.
    PLoS ONE 01/2013; 8(8):e70687. · 3.73 Impact Factor
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    ABSTRACT: More than 65 loci, encoding up to 500 different genes, have been implicated by genome-wide association studies (GWAS) as conferring an increased risk of developing type 2 diabetes (T2D). Whilst mouse models have in the past been central to understanding the mechanisms through which more penetrant risk genes for T2D, for example, those responsible for neonatal or maturity-onset diabetes of the young, only a few of those identified by GWAS, notably TCF7L2 and ZnT8/SLC30A8, have to date been examined in mouse models. We discuss here the animal models available for the latter genes and provide perspectives for future, higher throughput approaches towards efficiently mining the information provided by human genetics.
    Journal of Diabetes Research 01/2013; 2013:906590. · 3.54 Impact Factor

Publication Stats

4k Citations
640.40 Total Impact Points

Institutions

  • 1985–2014
    • Imperial College London
      • • Section of Cell Biology
      • • Department of Physics
      Londinium, England, United Kingdom
  • 2012
    • Università degli Studi di Modena e Reggio Emilia
      • Department of Diagnostic Medicine, Clinical and Public Health
      Modène, Emilia-Romagna, Italy
  • 2007–2011
    • University College London
      • Department of Computer Science
      London, ENG, United Kingdom
  • 2006–2007
    • University of Strathclyde
      • Institute of Photonics
      Glasgow, SCT, United Kingdom
  • 2002
    • University of Oxford
      • Department of Engineering Science
      Oxford, ENG, United Kingdom
  • 2000
    • Purdue University
      • Department of Physics
      West Lafayette, IN, United States
  • 1986–1998
    • Imperial Valley College
      • Physics Department
      Imperial, California, United States