Publications by authors named "Jack Williams"

67 Publications

Strategies to expand peptide functionality through hybridisation with a small molecule component.

RSC Chem Biol 2021 Feb 8;2(1):151-165. Epub 2020 Dec 8.

Protein-Protein Interaction Laboratory, The Francis Crick Institute London UK

Combining different compound classes gives molecular hybrids that can offer access to novel chemical space and unique properties. Peptides provide ideal starting points for such molecular hybrids, which can be easily modified with a variety of molecular entities. The addition of small molecules can improve the potency, stability and cell permeability of therapeutically relevant peptides. Furthermore, they are often applied to create peptide-based tools in chemical biology. In this review, we discuss general methods that allow the discovery of this compound class and highlight key examples of peptide-small molecule hybrids categorised by the application and function of the small molecule entity.
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http://dx.doi.org/10.1039/d0cb00167hDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8341444PMC
February 2021

Growth Hormone Receptor (GHR) 6Ω Pseudoexon Activation: a Novel Cause of Severe Growth Hormone Insensitivity.

J Clin Endocrinol Metab 2021 Jul 28. Epub 2021 Jul 28.

Centre for Endocrinology, William Harvey Research Institute: Barts and The London School of Medicine and Dentistry William Harvey Research Institute, London EC1M 6BQ, UK.

Context: Severe forms of growth hormone insensitivity (GHI) are characterized by extreme short stature, dysmorphism, and metabolic anomalies.

Objective: This work aims to identify the genetic cause of growth failure in 3 "classical" GHI individuals.

Methods: A novel intronic growth hormone receptor gene (GHR) variant was identified, and in vitro splicing assays confirmed aberrant splicing. A 6Ω pseudoexon GHR vector and patient fibroblast analysis assessed the consequences of the novel pseudoexon inclusion and the impact on GHR function.

Results: We identified a novel homozygous intronic GHR variant (g.5:42700940T > G, c.618+836T > G), 44 bp downstream of the previously recognized intronic 6Ψ GHR pseudoexon mutation in the index patient. Two siblings also harbored the novel intronic 6Ω pseudoexon GHR variant in compound heterozygosity with the known GHR c.181C > T (R43X) mutation. In vitro splicing analysis confirmed inclusion of a 151-bp mutant 6Ω pseudoexon not identified in wild-type constructs. Inclusion of the 6Ω pseudoexon causes a frameshift resulting in a nonfunctional truncated GHR lacking the transmembrane and intracellular domains. The truncated 6Ω pseudoexon protein demonstrated extracellular accumulation and diminished activation of STAT5B signaling following GH stimulation.

Conclusion: Novel GHR 6Ω pseudoexon inclusion results in loss of GHR function consistent with a severe GHI phenotype. This represents a novel mechanism of Laron syndrome and is the first deep intronic variant identified causing severe postnatal growth failure. The 2 kindreds originate from the same town in Campania, Southern Italy, implying common ancestry. Our findings highlight the importance of studying variation in deep intronic regions as a cause of monogenic disorders.
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http://dx.doi.org/10.1210/clinem/dgab550DOI Listing
July 2021

Growth Hormone Receptor (Ghr) 6ω Pseudoexon Activation: A Novel Cause Of Severe Growth Hormone Insensitivity (Ghi).

J Clin Endocrinol Metab 2021 Jul 28. Epub 2021 Jul 28.

Centre for Endocrinology, William Harvey Research Institute: Barts and The London School of Medicine and Dentistry William Harvey Research Institute.

Context: Severe forms of Growth Hormone Insensitivity (GHI) are characterized by extreme short stature, dysmorphism and metabolic anomalies.

Objective: Identification of the genetic cause of growth failure in 3 'classical' GHI subjects.

Design: A novel intronic GHR variant was identified, and in vitro splicing assays confirmed aberrant splicing. A 6Ω pseudoexon GHR vector and patient fibroblast analysis assessed the consequences of the novel pseudoexon inclusion and the impact on GHR function.

Results: We identified a novel homozygous intronic GHR variant (g.5:42700940T>G, c.618 + 836T> G), 44bp downstream of the previously recognized intronic 6Ψ GHR pseudoexon mutation in the index patient. Two siblings also harbored the novel intronic 6Ω pseudoexon GHR variant in compound heterozygosity with the known GHR c.181C>T (R43X) mutation. In vitro splicing analysis confirmed inclusion of a 151bp mutant 6Ω pseudoexon not identified in wild-type constructs. Inclusion of the 6Ω pseudoexon causes a frameshift resulting in a non-functional truncated GHR lacking the transmembrane and intracellular domains. The truncated 6Ω pseudoexon protein demonstrated extracellular accumulation and diminished activation of STAT5B signaling following growth hormone stimulation.

Conclusion: Novel GHR 6Ω pseudoexon inclusion results in loss of GHR function consistent with a severe GHI phenotype. This represents a novel mechanism of Laron syndrome and is the first deep intronic variant identified causing severe postnatal growth failure. The 2 kindreds originate from the same town in Campania, Southern Italy, implying common ancestry. Our findings highlight the importance of studying variation in deep intronic regions as a cause of monogenic disorders.
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http://dx.doi.org/10.1210/clinem/dgab550DOI Listing
July 2021

Loss of Increases Expression of Oxidative Phosphorylation Complexes in C57BL/6J Hearts.

Int J Mol Sci 2021 Jun 5;22(11). Epub 2021 Jun 5.

Centre for Endocrinology, William Harvey Research Institute, Charterhouse Square, Barts and the London School of Medicine and Dentistry, Queen Mary University of London, London EC1M 6BQ, UK.

Nicotinamide nucleotide transhydrogenase (NNT) is a proton pump in the inner mitochondrial membrane that generates reducing equivalents in the form of NAPDH, which can be used for anabolic pathways or to remove reactive oxygen species (ROS). A number of studies have linked NNT dysfunction to cardiomyopathies and increased risk of atherosclerosis; however, biallelic mutations in humans commonly cause a phenotype of adrenal insufficiency, with rare occurrences of cardiac dysfunction and testicular tumours. Here, we compare the transcriptomes of the hearts, adrenals and testes from three mouse models: the C57BL/6N, which expresses NNT; the C57BL/6J, which lacks NNT; and a third mouse, expressing the wild-type NNT sequence on the C57BL/6J background. We saw enrichment of oxidative phosphorylation genes in the C57BL/B6J in the heart and adrenal, possibly indicative of an evolved response in this substrain to loss of However, differential gene expression was mainly driven by mouse background with some changes seen in all three tissues, perhaps reflecting underlying genetic differences between the C57BL/B6J and -6N substrains.
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http://dx.doi.org/10.3390/ijms22116101DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8201288PMC
June 2021

Tranexamic acid to reduce head injury death in people with traumatic brain injury: the CRASH-3 international RCT.

Health Technol Assess 2021 Apr;25(26):1-76

Department of Health Services Research and Policy, London School of Hygiene & Tropical Medicine, London, UK.

Background: Tranexamic acid safely reduces mortality in traumatic extracranial bleeding. Intracranial bleeding is common after traumatic brain injury and can cause brain herniation and death. We assessed the effects of tranexamic acid in traumatic brain injury patients.

Objective: To assess the effects of tranexamic acid on death, disability and vascular occlusive events in traumatic brain injury patients. We also assessed cost-effectiveness.

Design: Randomised trial and economic evaluation. Patients were assigned by selecting a numbered treatment pack from a box containing eight packs that were identical apart from the pack number. Patients, caregivers and those assessing outcomes were masked to allocation. All analyses were by intention to treat. We assessed the cost-effectiveness of tranexamic acid versus no treatment from a UK NHS perspective using the trial results and a Markov model.

Setting: 175 hospitals in 29 countries.

Participants: Adults with traumatic brain injury within 3 hours of injury with a Glasgow Coma Scale score of ≤ 12 or any intracranial bleeding on computerised tomography scan, and no major extracranial bleeding, were eligible.

Intervention: Tranexamic acid (loading dose 1 g over 10 minutes then infusion of 1 g over 8 hours) or matching placebo.

Main Outcome Measures: Head injury death in hospital within 28 days of injury in patients treated within 3 hours of injury. Secondary outcomes were early head injury deaths, all-cause and cause-specific mortality, disability, vascular occlusive events, seizures, complications and adverse events.

Results: Among patients treated within 3 hours of injury ( = 9127), the risk of head injury death was 18.5% in the tranexamic acid group versus 19.8% in the placebo group (855/4613 vs. 892/4514; risk ratio 0.94, 95% confidence interval 0.86 to 1.02). In a prespecified analysis excluding patients with a Glasgow Coma Scale score of 3 or bilateral unreactive pupils at baseline, the results were 12.5% in the tranexamic acid group versus 14.0% in the placebo group (485/3880 vs. 525/3757; risk ratio 0.89, 95% confidence interval 0.80 to 1.00). There was a reduction in the risk of head injury death with tranexamic acid in those with mild to moderate head injury (166/2846 vs. 207/2769; risk ratio 0.78, 95% confidence interval 0.64 to 0.95), but in those with severe head injury (689/1739 vs. 685/1710; risk ratio 0.99, 95% confidence interval 0.91 to 1.07) there was no apparent reduction (-value for heterogeneity = 0.030). Early treatment was more effective in mild and moderate head injury ( = 0.005), but there was no obvious impact of time to treatment in cases of severe head injury ( = 0.73). The risk of disability, vascular occlusive events and seizures was similar in both groups. Tranexamic acid is highly cost-effective for mild and moderate traumatic brain injury (base case of £4288 per quality-adjusted life-year gained).

Conclusion: Early tranexamic acid treatment reduces head injury deaths. Treatment is cost-effective for patients with mild or moderate traumatic brain injury, or those with both pupils reactive.

Future Work: Further trials should examine early tranexamic acid treatment in mild head injury. Research on alternative routes of administration is needed.

Limitations: Time to treatment may have been underestimated.

Trial Registration: Current Controlled Trials ISRCTN15088122, ClinicalTrials.gov NCT01402882, EudraCT 2011-003669-14, Pan African Clinical Trial Registry PACTR20121000441277.

Funding: The project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in ; Vol. 25, No. 26. See the NIHR Journals Library website for further project information. In addition, funding was provided by JP Moulton Charitable Trust, Joint Global Health Trials (Medical Research Council, Department for International Development and the Wellcome Trust). This project was funded by the NIHR Global Health Trials programme.
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http://dx.doi.org/10.3310/hta25260DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8107835PMC
April 2021

Clinical Study of the IOPTx™ System - an Electroceutical Wearable to Lower Intraocular Pressure.

Curr Eye Res 2021 Oct 7;46(10):1531-1538. Epub 2021 Apr 7.

Department of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana, USA.

: To investigate the safety and efficacy of the IOPTx™ system - a novel wearable, electroceutical treatment to lower intraocular pressure. : Patients wear the customized contact lens and spectacles of the IOPTx™ system and undergo three 15-minute randomized stimulation trials at different stimulus amplitudes with 15 minutes of rest in between. The parameters for the stimulation trials include a frequency of 50 Hz, a pulse width of 100 µs, and current amplitudes between 90-150 µA. The optometrist measures the intraocular pressure (IOP) before, immediately after, and 15 minutes after the trial, and performs topography, a slit eye examination, and specular microscopy before and after the entire study to check the health of the eye and confirm the safety of the system. : The IOPTx™ system successfully modulates a patient's IOP. By testing various currents, we create individual tuning curves examining the effect of the stimulation amplitude on the change in IOP. Each patient may have an optimal dose-response curve and by normalizing to this value, the IOPTx™ system decreased IOP by an average of 17.7% with fifteen minutes of therapy. No Adverse Events or Adverse Device Effects occurred.: The results of this clinical case series provide preliminary evidence of efficacy and safety of the IOPTx™ system and its potential usefulness to lower IOP in glaucoma and ocular hypertension.
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http://dx.doi.org/10.1080/02713683.2021.1904999DOI Listing
October 2021

Brain Activity Fluctuations Propagate as Waves Traversing the Cortical Hierarchy.

Cereb Cortex 2021 Jul;31(9):3986-4005

Department of Biomedical Engineering, The Pennsylvania State University, University Park, PA, 16802, USA.

The brain exhibits highly organized patterns of spontaneous activity as measured by resting-state functional magnetic resonance imaging (fMRI) fluctuations that are being widely used to assess the brain's functional connectivity. Some evidence suggests that spatiotemporally coherent waves are a core feature of spontaneous activity that shapes functional connectivity, although this has been difficult to establish using fMRI given the temporal constraints of the hemodynamic signal. Here, we investigated the structure of spontaneous waves in human fMRI and monkey electrocorticography. In both species, we found clear, repeatable, and directionally constrained activity waves coursed along a spatial axis approximately representing cortical hierarchical organization. These cortical propagations were closely associated with activity changes in distinct subcortical structures, particularly those related to arousal regulation, and modulated across different states of vigilance. The findings demonstrate a neural origin of spatiotemporal fMRI wave propagation at rest and link it to the principal gradient of resting-state fMRI connectivity.
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http://dx.doi.org/10.1093/cercor/bhab064DOI Listing
July 2021

Knee cartilage T relaxation times 3 months after ACL reconstruction are associated with knee gait variables linked to knee osteoarthritis.

J Orthop Res 2021 Mar 30. Epub 2021 Mar 30.

Department of Mechanical Engineering, University of Delaware, Newark, Delaware, USA.

Osteoarthritis development after ACL reconstruction (ACLR) is not well understood. Investigators have examined associations between knee biomechanical alterations and quantitative MRI (qMRI) variables, reflective of cartilage health, 12-60 months following ACLR; however, none have done so early after surgery. As part of an exploratory study, 45 individuals (age, 23 ± 7 years) underwent motion analysis during walking and qMRI 3 months after ACLR. For each limb, peak knee adduction moment (pKAM) and peak knee flexion moment (pKFM) were determined using inverse dynamics and peak medial compartment force was calculated using a neuromusculoskeletal model. T relaxation times in the medial compartment and linear regressions were used to determine the associations between gait variables and deep and superficial cartilage T relaxation times in six regions. pKAM was positively associated with deep layer T relaxation times within the femoral central and posterior regions when examined in the involved limb and from an interlimb difference perspective (involved limb - uninvolved limb). After adjusting for age, the association between interlimb difference of pKAM and interlimb difference of deep layer T relaxation times in the tibial central region became significant (p = .043). Interlimb difference of pKFM was negatively associated with interlimb difference of deep layer T relaxation times within the femoral central and posterior regions. These associations suggest that degenerative pathways leading to osteoarthritis may be detectable as early as 3 months after reconstruction. Preventative therapeutic techniques may need to be employed early in the rehabilitation process to prevent cartilage degradation.
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http://dx.doi.org/10.1002/jor.25043DOI Listing
March 2021

Experimental Determination of H and CO Diffusion Coefficients in a Wax Mixture Confined in a Porous Titania Catalyst Support.

J Phys Chem B 2020 Dec 20;124(48):10971-10982. Epub 2020 Nov 20.

Department of Chemical Engineering and Biotechnology, University of Cambridge, West Cambridge Site, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K.

The ability to measure and predict molecular diffusion coefficients in multicomponent mixtures is not only of fundamental scientific interest but also of significant relevance in understanding how catalytic processes proceed. In the present work, the direct measurement of the molecular diffusion of H and CO gas-phase species diffusing in -alkane mixtures using pulsed-field gradient (PFG) nuclear magnetic resonance (NMR) methods is reported. The work is of direct relevance to Fischer-Tropsch (FT) catalysis, with the measurements being made of the gas-wax system with the wax in both the bulk liquid state and when confined within a titania catalyst support, at temperatures and pressures typical of low-temperature FT synthesis. Molecular diffusion coefficients of H and CO within wax-saturated porous titania in the range (1.00-2.43) × 10 and (6.44-8.50) × 10 m s, respectively, were measured in the temperature range of 140-240 and 200-240 °C for H and CO, respectively, at a pressure of 40 bar. The wax mixture was typical of a wax produced during FT catalysis and had a molar average carbon number of 36. It is shown that the hydrogen diffusion coefficient within this wax mixture is consistent, to within experimental error, with the hydrogen diffusion coefficient measured in pure single-component -hexatriacontane (-C) wax; this result held with the waxes in the bulk liquid state and when confined within the porous titania. The tortuosity of the porous titania was also measured using PFG NMR and found to be 1.77; this value is independent of temperature. The ability of existing correlations to predict these experimentally determined data was then critically evaluated. Although the Wilke-Chang correlation was found to underestimate the molecular diffusion coefficients of both H and CO diffusing in the wax in both the bulk state and when confined within the porous titania, parameterized correlations based on the rough hard sphere model, having accounted for the experimentally determined tortuosity factor, predicted the H and CO diffusion within bulk and confined wax to within 3%.
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http://dx.doi.org/10.1021/acs.jpcb.0c07440DOI Listing
December 2020

Cost-effectiveness analysis of tranexamic acid for the treatment of traumatic brain injury, based on the results of the CRASH-3 randomised trial: a decision modelling approach.

BMJ Glob Health 2020 09;5(9)

Health Services Research and Policy, London School of Hygiene & Tropical Medicine, London, UK.

Introduction: An estimated 69 million traumatic brain injuries (TBI) occur each year worldwide, with most in low-income and middle-income countries. The CRASH-3 randomised trial found that intravenous administration of tranexamic acid within 3 hours of injury reduces head injury deaths in patients sustaining a mild or moderate TBI. We examined the cost-effectiveness of tranexamic acid treatment for TBI.

Methods: A Markov decision model was developed to assess the cost-effectiveness of treatment with and without tranexamic acid, in addition to current practice. We modelled the decision in the UK and Pakistan from a health service perspective, over a lifetime time horizon. We used data from the CRASH-3 trial for the risk of death during the trial period (28 days) and patient quality of life, and data from the literature to estimate costs and long-term outcomes post-TBI. We present outcomes as quality-adjusted life years (QALYs) and 2018 costs in pounds for the UK, and US dollars for Pakistan. Incremental cost-effectiveness ratios (ICER) per QALY gained were estimated, and compared with country specific cost-effective thresholds. Deterministic and probabilistic sensitivity analyses were also performed.

Results: Tranexamic acid was highly cost-effective for patients with mild TBI and intracranial bleeding or patients with moderate TBI, at £4288 per QALY in the UK, and US$24 per QALY in Pakistan. Tranexamic acid was 99% and 98% cost-effective at the cost-effectiveness thresholds for the UK and Pakistan, respectively, and remained cost-effective across all deterministic sensitivity analyses. Tranexamic acid was even more cost-effective with earlier treatment administration. The cost-effectiveness for those with severe TBI was uncertain.

Conclusion: Early administration of tranexamic acid is highly cost-effective for patients with mild or moderate TBI in the UK and Pakistan, relative to the cost-effectiveness thresholds used. The estimated ICERs suggest treatment is likely to be cost-effective across all income settings globally.
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http://dx.doi.org/10.1136/bmjgh-2020-002716DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7470492PMC
September 2020

An Economic Evaluation of the Cost-Effectiveness of Opt-Out Hepatitis B and Hepatitis C Testing in an Emergency Department Setting in the United Kingdom.

Value Health 2020 08 14;23(8):1003-1011. Epub 2020 Jul 14.

Department of Health Service Research and Policy, London School of Hygiene & Tropical Medicine, London, England, UK; The National Institute for Health Research Health Protection Research Unit (NIHR HPRU) in Blood Borne and Sexually Transmitted Infections at University College, London, England, UK.

Objectives: The prevalence of hepatitis is high in emergency department (ED) attendees in the United Kingdom, with a prevalence of up to 2% for hepatitis B (HBV) HBsAg, and 2.9% for hepatitis C (HCV) RNA. The aim of this paper is to perform an economic evaluation of opt-out ED-based HCV and HBV testing.

Methods: A Markov model was developed to analyze the cost-effectiveness of opt-out HCV and HBV testing in EDs in the UK. The model used data from UK studies of ED testing to parameterize the HCV and HBV prevalence (1.4% HCV RNA, 0.84% HBsAg), test costs, and intervention effects (contact rates and linkage to care). For HCV, we used an antibody test cost of £3.64 and RNA test cost of £68.38, and assumed direct-acting antiviral treatment costs of £10 000. For HBV, we used a combined HBsAg and confirmatory test cost of £5.79. We also modeled the minimum prevalence of HCV (RNA-positive) and HBV (HBsAg) required to make ED testing cost-effective at a £20 000 willingness to pay per quality-adjusted life-year threshold.

Results: In the base case, ED testing was highly cost-effective, with HCV and HBV testing costing £8019 and £9858 per quality-adjusted life-year gained, respectively. HCV and HBV ED testing remained cost-effective at 0.25% HCV RNA or HBsAg prevalence or higher.

Conclusions: Emergency department testing for HCV and HBV is highly likely to be cost-effective in many areas across the UK depending on their prevalence. Ongoing studies will help evaluate ED testing across different regions to inform testing guidelines.
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http://dx.doi.org/10.1016/j.jval.2020.03.014DOI Listing
August 2020

null C57BL/6N mice develop cardiomyopathy, whereas null C57BL/6J mice do not.

Life Sci Alliance 2020 04 25;3(4). Epub 2020 Mar 25.

Centre for Endocrinology, William Harvey Research Institute, Charterhouse Square, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London, UK

The C57BL/6J and C57BL/6N mice have well-documented phenotypic and genotypic differences, including the infamous nicotinamide nucleotide transhydrogenase () null mutation in the C57BL/6J substrain, which has been linked to cardiovascular traits in mice and cardiomyopathy in humans. To assess whether loss alone causes a cardiovascular phenotype, we investigated the C57BL/6N, C57BL/6J mice and a C57BL/6J-BAC transgenic rescuing NNT expression, at 3, 12, and 18 mo. We identified a modest dilated cardiomyopathy in the C57BL/6N mice, absent in the two B6J substrains. Immunofluorescent staining of cardiomyocytes revealed eccentric hypertrophy in these mice, with defects in sarcomere organisation. RNAseq analysis identified differential expression of a number of cardiac remodelling genes commonly associated with cardiac disease segregating with the phenotype. Variant calling from RNAseq data identified a myosin light chain kinase 3 () mutation in C57BL/6N mice, which abolishes MYLK3 protein expression. These results indicate the C57BL/6J -null mice do not develop cardiomyopathy; however, we identified a null mutation in as a credible cause of the cardiomyopathy phenotype in the C57BL/6N.
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http://dx.doi.org/10.26508/lsa.201900593DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7103425PMC
April 2020

Determination of Carbon Number Distributions of Mixtures of Linear Hydrocarbons Confined within Porous Media Using Pulsed Field Gradient NMR.

Anal Chem 2020 Apr 25;92(7):5125-5133. Epub 2020 Mar 25.

Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, United Kingdom.

Pulsed field gradient (PFG) NMR measurements, combined with a novel optimization method, are used to determine the composition of hydrocarbon mixtures of linear alkanes (C7-C16) in both the bulk liquid state and when imbibed within a porous medium of mean pore diameter 28.6 nm. The method predicts the average carbon number of a given mixture to an accuracy of ±1 carbon number and the mole fraction of a mixture component to within an average root-mean-square error of ±0.036 with just three calibration mixtures. Given that the method can be applied at any conditions of temperature and pressure at which the PFG NMR measurements are made, the method has the potential for application in characterizing hydrocarbon liquid mixtures inside porous media and at the operating conditions relevant to, for example, hydrocarbon recovery and heterogeneous catalysis.
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http://dx.doi.org/10.1021/acs.analchem.9b05600DOI Listing
April 2020

Cost effectiveness of an intervention to increase uptake of hepatitis C virus testing and treatment (HepCATT): cluster randomised controlled trial in primary care.

BMJ 2020 02 26;368:m322. Epub 2020 Feb 26.

Population Health Sciences, Bristol Medical School, University of Bristol, Bristol, UK

Objective: To evaluate the effectiveness and cost effectiveness of a complex intervention in primary care that aims to increase uptake of hepatitis C virus (HCV) case finding and treatment.

Design: Pragmatic, two armed, practice level, cluster randomised controlled trial and economic evaluation.

Setting And Participants: 45 general practices in South West England (22 randomised to intervention and 23 to control arm). Outcome data were collected from all intervention practices and 21/23 control practices. Total number of flagged patients was 24 473 (about 5% of practice list).

Intervention: Electronic algorithm and flag on practice systems identifying patients with HCV risk markers (such as history of opioid dependence or HCV tests with no evidence of referral to hepatology), staff educational training in HCV, and practice posters/leaflets to increase patients' awareness. Flagged patients were invited by letter for an HCV test (with one follow-up) and had on-screen pop-ups to encourage opportunistic testing. The intervention lasted one year, with practices recruited April to December 2016.

Main Outcome Measures: Primary outcome: uptake of HCV testing.

Secondary Outcomes: number of positive HCV tests and yield (proportion HCV positive); HCV treatment assessment at hepatology; cost effectiveness.

Results: Baseline HCV testing of flagged patients (six months before study start) was 608/13 097 (4.6%) in intervention practices and 380/11 376 (3.3%) in control practices. During the study 2071 (16%) of flagged patients in the intervention practices and 1163 (10%) in control practices were tested for HCV: overall intervention effect as an adjusted rate ratio of 1.59 (95% confidence interval 1.21 to 2.08; P<0.001). HCV antibodies were detected in 129 patients from intervention practices and 51 patients from control practices (adjusted rate ratio 2.24, 1.47 to 3.42) with weak evidence of an increase in yield (6.2% 4.4%; adjusted risk ratio 1.40, 0.99 to 1.95). Referral and assessment increased in intervention practices compared with control practices (adjusted rate ratio 5.78, 1.6 to 21.6) with a risk difference of 1.3 per 1000 and a "number needed to help" of one extra HCV diagnosis, referral, and assessment per 792 (95% confidence interval 558 to 1883) patients flagged. The average cost of HCV case finding was £4.03 (95% confidence interval £2.27 to £5.80) per at risk patient and £3165 per additional patient assessed at hepatology. The incremental cost effectiveness ratio was £6212 per quality adjusted life year (QALY), with 92.5% probability of being below £20 000 per QALY.

Conclusion: HepCATT had a modest impact but is a low cost intervention that merits optimisation and implementation as part of an NHS strategy to increase HCV testing and treatment.

Trial Registration: ISRCTN61788850.
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http://dx.doi.org/10.1136/bmj.m322DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7190058PMC
February 2020

GHR gene transcript heterogeneity may explain phenotypic variability in GHR pseudoexon (6Ψ) patients.

Endocr Connect 2020 Feb 1. Epub 2020 Feb 1.

H Storr, Centre for Endocrinology, William Harvey Research Institute, Barts and the London School of Medicine and Dentistry, London, EC1M 6BQ, United Kingdom of Great Britain and Northern Ireland.

Objectives: The homozygous GH receptor (GHR) pseudoexon (6Ψ) mutation leads to growth hormone insensitivity (GHI) with clinical and biochemical heterogeneity. We investigated whether transcript heterogeneity (6Ψ-GHR to WT-GHR transcript ratio) and/or concurrent defects in other short stature (SS) genes contribute to this.

Methods: 6Ψ-GHR and WT-GHR mRNA transcripts of 4 6Ψ patient (height SDS -4.2 to -3.1) and 1 control fibroblasts were investigated by RT-PCR. Transcripts were quantified by qRT-PCR and delta delta CT analysis and compared using ANOVA with Bonferroni correction. In eleven 6Ψ patients, 40 genes known to cause GHI/SS were analysed by targeted next generation sequencing.

Results: RT-PCR confirmed 6Ψ-GHR transcript in the 6Ψ patients but not control. 6Ψ-GHR transcript levels were comparable in patients 1 and 3 but significantly different among all other patients. The mean 6Ψ:WT transcript ratios ranged from 29-71:1 for patients 1-4 and correlated negatively with height SDS (R=-0.85; p<0.001). Eight deleterious variants in 6 genes were detected but the number of gene hits did not correlate with the degree of SS in individual 6Ψ patients.

Conclusion: Variable amounts of 6Ψ- and WT-GHR transcripts were identified in 6Ψ patients but no 6Ψ transcript was present in the control. Higher 6Ψ:WT GHR transcript ratio correlated with SS severity and may explain the phenotypic variability. Analysis of known SS genes suggested that phenotypic variation is independent of the genetic background. This is the first report of transcript heterogeneity producing a spectrum of clinical phenotypes in different individuals harbouring an identical homozygous genetic mutation.
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http://dx.doi.org/10.1530/EC-20-0026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7077524PMC
February 2020

A releasable disulfide-linked peptide tag facilitates the synthesis and purification of short peptides.

Chem Commun (Camb) 2020 Mar 10;56(19):2917-2920. Epub 2020 Feb 10.

Institute of Chemical Sciences and Engineering, Ecole Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland.

Combinatorial cyclization of hundreds to thousands of random linear peptides by structurally diverse chemical linkers offers access to large macrocyclic compound libraries. A bottleneck in the development of such libraries is the preparation of large numbers of short random linear peptides. Herein, we present a tag-based strategy that is not dependent on a throughput-limiting chromatographic purification step and thus enables parallel production of short peptides. In brief, peptides are synthesized on solid phase as conjugates with a disulfide-linked Cys-Gly-Arg-Trp tetra-peptide tag. The charged arginine residue in the tag allows for purification of the peptides by diethyl ether-precipitation and the tryptophan allows for quantification of the product by absorption measurement. Addition of a reducing agent releases the short peptides from the tag. The released sulfhydryl group in the peptide can readily be used for cyclization of the peptide library with bis-electrophilic linker reagents.
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http://dx.doi.org/10.1039/c9cc09247aDOI Listing
March 2020

Slower Walking Speed Is Related to Early Femoral Trochlear Cartilage Degradation After ACL Reconstruction.

J Orthop Res 2020 03 18;38(3):645-652. Epub 2019 Nov 18.

Department of Mechanical Engineering, University of Delaware, Newark, Delaware.

Post-traumatic patellofemoral osteoarthritis (OA) is prevalent after anterior cruciate ligament reconstruction (ACLR) and early cartilage degradation may be especially common in the femoral trochlear cartilage. Determining the presence of and factors associated with early femoral trochlear cartilage degradation, a precursor to OA, is a critical preliminary step in identifying those at risk for patellofemoral OA development and designing interventions to combat the disease. Early cartilage degradation can be detected using quantitative magnetic resonance imaging measures, such as tissue T relaxation time. The purposes of this study were to (i) compare involved (ACLR) versus uninvolved (contralateral) femoral trochlear cartilage T relaxation times 6 months after ACLR, and (ii) determine the relationship between walking speed and walking mechanics 3 months after ACLR and femoral trochlear cartilage T relaxation times 6 months after ACLR. Twenty-six individuals (age 23 ± 7 years) after primary, unilateral ACLR participated in detailed motion analyses 3.3 ± 0.6 months after ACLR and quantitative magnetic resonance imaging 6.3 ± 0.5 months after ACLR. There were no limb differences in femoral trochlear cartilage T relaxation times. Slower walking speed was related to higher (worse) femoral trochlear cartilage T relaxation times in the involved limb (Pearson's r: -0.583, p = 0.002) and greater interlimb differences in trochlear T relaxation times (Pearson's r: -0.349, p = 0.080). Walking mechanics were weakly related to trochlear T relaxation times. Statement of clinical significance: Slower walking speed was by far the strongest predictor of worse femoral trochlear cartilage health, suggesting slow walking speed may be an early clinical indicator of future patellofemoral OA after ACLR. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:645-652, 2020.
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http://dx.doi.org/10.1002/jor.24503DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7028512PMC
March 2020

Cost-Effectiveness of One-Time Birth Cohort Screening for Hepatitis C as Part of the National Health Service Health Check Program in England.

Value Health 2019 11 19;22(11):1248-1256. Epub 2019 Aug 19.

The National Institute for Health Research Health Protection Research Unit in Blood Borne and Sexually Transmitted Infections at University College London, England, UK; Population Health Sciences, Bristol Medical School, University of Bristol, England, UK; The National Institute for Health Research Health Protection Research Unit in Evaluation of Interventions, England, UK.

Background And Objectives: Birth cohort screening for the hepatitis C virus (HCV) has been implemented in the US, but there is little evidence of its cost-effectiveness in England. We aim to evaluate the cost-effectiveness of one-time HCV screening for individuals born between 1950 and 1979 as part of the National Health Service health check in England, a health check for adults aged 40 to 74 years in primary care.

Methods: A Markov model was developed to analyze add-on HCV testing to the National Health Service health check for individuals in birth cohorts between 1950 and 1979, versus current background HCV testing only, over a lifetime horizon. The model used data from a back-calculation model of the burden of HCV in England, sentinel surveillance of HCV testing, and published literature. Results are presented from a health service perspective in pounds in 2017, as incremental cost-effectiveness ratios per quality-adjusted life years gained.

Results: The base-case incremental cost-effectiveness ratios ranged from £7648 to £24 434, and £18 681 to £46 024, across birth cohorts when considering 2 sources of HCV transition probabilities. The intervention is most likely to be cost-effective for those born in the 1970s, and potentially cost-effective for those born from 1955 to 1969. The model results were most sensitive to the source of HCV transition probabilities, the probability of referral and receiving treatment, and the HCV prevalence among testers. The maximum value of future research across all birth cohorts was £11.3 million at £20 000 per quality-adjusted life years gained.

Conclusion: Birth cohort screening is likely to be cost-effective for younger birth cohorts, although considerable uncertainty exists for other birth cohorts. Further studies are warranted to reduce uncertainty in cost-effectiveness and consider the acceptability of the intervention.
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http://dx.doi.org/10.1016/j.jval.2019.06.006DOI Listing
November 2019

Chronic hepatitis B virus case-finding in UK populations born abroad in intermediate or high endemicity countries: an economic evaluation.

BMJ Open 2019 06 28;9(6):e030183. Epub 2019 Jun 28.

Public Health and Policy, London School of Hygiene and Tropical Medicine, London, UK.

Objectives: The majority (>90%) of new or undiagnosed cases of hepatitis B virus (HBV) in the UK are among individuals born in countries with intermediate or high prevalence levels (≥2%). We evaluate the cost-effectiveness of increased HBV case-finding among UK migrant populations, based on a one-time opt out case-finding approach in a primary care setting.

Design: Cost-effectiveness evaluation. A decision model based on a Markov approach was built to assess the progression of HBV infection with and without treatment as a result of case-finding. The model parameters, including the cost and effects of case-finding and treatment, were estimated from the literature. All costs were expressed in 2017/2018 British Pounds (GBPs) and health outcomes as quality-adjusted life-years (QALYs).

Intervention: Hepatitis B virus case-finding among UK migrant populations born in countries with intermediate or high prevalence levels (≥2%) in a primary care setting compared with no intervention (background testing).

Results: At a 2% hepatitis B surface antigen (HBsAg) prevalence, the case-finding intervention led to a mean incremental cost-effectiveness ratio of £13 625 per QALY gained which was 87% and 98% likely of being cost-effective at willingness to pay (WTP) thresholds of £20 000 and £30 000 per additional QALY, respectively. Sensitivity analyses indicated that the intervention would remain cost-effective under a £20 000 WTP threshold as long as HBsAg prevalence among the migrant population is at least 1%. However, the results were sensitive to a number of parameters, especially the time horizon and probability of treatment uptake.

Conclusions: HBV case-finding using a one-time opt out approach in primary care settings is very likely to be cost-effective among UK migrant populations with HBsAg prevalence ≥1% if the WTP for an additional QALY is around £20 000.
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http://dx.doi.org/10.1136/bmjopen-2019-030183DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6609059PMC
June 2019

Disrupted circadian clocks and altered tissue mechanics in primary human breast tumours.

Breast Cancer Res 2018 10 22;20(1):125. Epub 2018 Oct 22.

Wellcome Centre for Cell-Matrix Research and Manchester Breast Centre, Faculty of Biology, Medicine and Health, University of Manchester, Manchester, M13 9PT, UK.

Background: Circadian rhythms maintain tissue homeostasis during the 24-h day-night cycle. Cell-autonomous circadian clocks play fundamental roles in cell division, DNA damage responses and metabolism. Circadian disruptions have been proposed as a contributing factor for cancer initiation and progression, although definitive evidence for altered molecular circadian clocks in cancer is still lacking. In this study, we looked at circadian clocks in breast cancer.

Methods: We isolated primary tumours and normal tissues from the same individuals who had developed breast cancer with no metastases. We assessed circadian clocks within primary cells of the patients by lentiviral expression of circadian reporters, and the levels of clock genes in tissues by qPCR. We histologically examined collagen organisation within the normal and tumour tissue areas, and probed the stiffness of the stroma adjacent to normal and tumour epithelium using atomic force microscopy.

Results: Epithelial ducts were disorganised within the tumour areas. Circadian clocks were altered in cultured tumour cells. Tumour regions were surrounded by stroma with an altered collagen organisation and increased stiffness. Levels of Bmal1 messenger RNA (mRNA) were significantly altered in the tumours in comparison to normal epithelia.

Conclusion: Circadian rhythms are suppressed in breast tumour epithelia in comparison to the normal epithelia in paired patient samples. This correlates with increased tissue stiffness around the tumour region. We suggest possible involvement of altered circadian clocks in the development and progression of breast cancer.
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http://dx.doi.org/10.1186/s13058-018-1053-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6198506PMC
October 2018

Over ten-year insulin independence following single allogeneic islet transplant without T-cell depleting antibody induction.

Islets 2018 19;10(4):168-174. Epub 2018 Jul 19.

a Division of Transplant, Department of Surgery , University of Illinois at Chicago, Chicago , Illinois.

Islet cell transplantation is a promising functional cure for type 1 diabetes; however, maintaining long-term islet graft function and insulin independence is difficult to achieve. In this short report we present a patient with situs inversus, who at the time of islet transplantation had a 26-year history of type 1 diabetes, complicated by hypoglycemic unawareness and severe hypoglycemic events. After a single allogeneic islet transplant of a low islet mass, and despite developing de novo anti-insulin and anti-GAD65 autoantibodies, the patient has remarkably maintained insulin independence with tight glycemic control and normal metabolic profiles for 10 years, after receiving prolonged non-T-cell depleting immunosuppression.
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http://dx.doi.org/10.1080/19382014.2018.1451281DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6281363PMC
May 2019

Trout in hot water: A call for global action.

Science 2018 05;360(6391):866-867

Illinois Natural History Survey, University of Illinois, Urbana, IL 61801, USA.

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http://dx.doi.org/10.1126/science.aat8455DOI Listing
May 2018

Epithelial and stromal circadian clocks are inversely regulated by their mechano-matrix environment.

J Cell Sci 2018 03 6;131(5). Epub 2018 Mar 6.

Wellcome Centre for Cell-Matrix Research and Manchester Breast Centre, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PT, UK

The circadian clock is an autonomous molecular feedback loop inside almost every cell in the body. We have shown that the mammary epithelial circadian clock is regulated by the cellular microenvironment. Moreover, a stiff extracellular matrix dampens the oscillations of the epithelial molecular clock. Here, we extend this analysis to other tissues and cell types, and identify an inverse relationship between circadian clocks in epithelia and fibroblasts. Epithelial cells from mammary gland, lung and skin have significantly stronger oscillations of clock genes in soft 3D microenvironments, compared to stiff 2D environments. Fibroblasts isolated from the same tissues show the opposite response, exhibiting stronger oscillations and more prolonged rhythmicity in stiff microenvironments. RNA analysis identified that a subset of mammary epithelial clock genes, and their regulators, are upregulated in 3D microenvironments in soft compared to stiff gels. Furthermore, the same genes are inversely regulated in fibroblasts isolated from the same tissues. Thus, our data reveal for the first time an intrinsic difference in the regulation of circadian genes in epithelia and fibroblasts.
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http://dx.doi.org/10.1242/jcs.208223DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5897718PMC
March 2018

Extreme hydrothermal conditions at an active plate-bounding fault.

Nature 2017 06 17;546(7656):137-140. Epub 2017 May 17.

SGEES, Victoria University of Wellington, PO Box 600, Wellington, New Zealand.

Temperature and fluid pressure conditions control rock deformation and mineralization on geological faults, and hence the distribution of earthquakes. Typical intraplate continental crust has hydrostatic fluid pressure and a near-surface thermal gradient of 31 ± 15 degrees Celsius per kilometre. At temperatures above 300-450 degrees Celsius, usually found at depths greater than 10-15 kilometres, the intra-crystalline plasticity of quartz and feldspar relieves stress by aseismic creep and earthquakes are infrequent. Hydrothermal conditions control the stability of mineral phases and hence frictional-mechanical processes associated with earthquake rupture cycles, but there are few temperature and fluid pressure data from active plate-bounding faults. Here we report results from a borehole drilled into the upper part of the Alpine Fault, which is late in its cycle of stress accumulation and expected to rupture in a magnitude 8 earthquake in the coming decades. The borehole (depth 893 metres) revealed a pore fluid pressure gradient exceeding 9 ± 1 per cent above hydrostatic levels and an average geothermal gradient of 125 ± 55 degrees Celsius per kilometre within the hanging wall of the fault. These extreme hydrothermal conditions result from rapid fault movement, which transports rock and heat from depth, and topographically driven fluid movement that concentrates heat into valleys. Shear heating may occur within the fault but is not required to explain our observations. Our data and models show that highly anomalous fluid pressure and temperature gradients in the upper part of the seismogenic zone can be created by positive feedbacks between processes of fault slip, rock fracturing and alteration, and landscape development at plate-bounding faults.
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http://dx.doi.org/10.1038/nature22355DOI Listing
June 2017

Cellular mechano-environment regulates the mammary circadian clock.

Nat Commun 2017 01 30;8:14287. Epub 2017 Jan 30.

Faculty of Biology, Medicine and Health, University of Manchester, Oxford Road, Manchester M13 9PT, UK.

Circadian clocks drive ∼24 h rhythms in tissue physiology. They rely on transcriptional/translational feedback loops driven by interacting networks of clock complexes. However, little is known about how cell-intrinsic circadian clocks sense and respond to their microenvironment. Here, we reveal that the breast epithelial clock is regulated by the mechano-chemical stiffness of the cellular microenvironment in primary cell culture. Moreover, the mammary clock is controlled by the periductal extracellular matrix in vivo, which contributes to a dampened circadian rhythm during ageing. Mechanistically, the tension sensing cell-matrix adhesion molecule, vinculin, and the Rho/ROCK pathway, which transduces signals provided by extracellular stiffness into cells, regulate the activity of the core circadian clock complex. We also show that genetic perturbation, or age-associated disruption of self-sustained clocks, compromises the self-renewal capacity of mammary epithelia. Thus, circadian clocks are mechano-sensitive, providing a potential mechanism to explain how ageing influences their amplitude and function.
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http://dx.doi.org/10.1038/ncomms14287DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5290282PMC
January 2017

Circadian clocks and breast cancer.

Breast Cancer Res 2016 09 2;18(1):89. Epub 2016 Sep 2.

Faculty of Biology, Medicine and Health, and Wellcome Trust Centre for Cell-Matrix Research, University of Manchester, Oxford Road, Manchester, M13 9PT, UK.

Circadian clocks respond to environmental time cues to coordinate 24-hour oscillations in almost every tissue of the body. In the breast, circadian clocks regulate the rhythmic expression of numerous genes. Disrupted expression of circadian genes can alter breast biology and may promote cancer. Here we overview circadian mechanisms, and the connection between the molecular clock and breast biology. We describe how disruption of circadian genes contributes to cancer via multiple mechanisms, and link this to increased tumour risk in women who work irregular shift patterns. Understanding the influence of circadian rhythms on breast cancer could lead to more efficacious therapies, reformed public health policy and improved patient outcome.
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http://dx.doi.org/10.1186/s13058-016-0743-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5010688PMC
September 2016

The intervertebral disc contains intrinsic circadian clocks that are regulated by age and cytokines and linked to degeneration.

Ann Rheum Dis 2017 Mar 3;76(3):576-584. Epub 2016 Aug 3.

Faculty of Life Sciences, University of Manchester, Manchester, UK.

Objectives: The circadian clocks are internal timing mechanisms that drive ∼24-hour rhythms in a tissue-specific manner. Many aspects of the physiology of the intervertebral disc (IVD) show clear diurnal rhythms. However, it is unknown whether IVD tissue contains functional circadian clocks and if so, how their dysregulation is implicated in IVD degeneration.

Methods: Clock gene dynamics in ex vivo IVD explants (from PER2:: luciferase (LUC) reporter mice) and human disc cells (transduced with lentivirus containing ::luc reporters) were monitored in real time by bioluminescence photon counting and imaging. Temporal gene expression changes were studied by RNAseq and quantitative reverse transcription (qRT)-PCR. IVD pathology was evaluated by histology in a mouse model with tissue-specific deletion of the core clock gene .

Results: Here we show the existence of the circadian rhythm in mouse IVD tissue and human disc cells. This rhythm is dampened with ageing in mice and can be abolished by treatment with interleukin-1β but not tumour necrosis factor α. Time-series RNAseq revealed 607 genes with 24-hour patterns of expression representing several essential pathways in IVD physiology. Mice with conditional knockout of in their disc cells demonstrated age-related degeneration of IVDs.

Conclusions: We have established autonomous circadian clocks in mouse and human IVD cells which respond to age and cytokines, and control key pathways involved in the homeostasis of IVDs. Genetic disruption to the mouse IVD molecular clock predisposes to IVD degeneration. These results support the concept that disruptions to circadian rhythms may be a risk factor for degenerative IVD disease and low back pain.
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http://dx.doi.org/10.1136/annrheumdis-2016-209428DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5446006PMC
March 2017

Detection of the Previously Unobserved Stereoisomers of Thujone in the Essential Oil and Consumable Products of Sage (Salvia officinalis L.) Using Headspace Solid-Phase Microextraction-Gas Chromatography-Mass Spectrometry.

J Agric Food Chem 2016 Jun 23;64(21):4319-26. Epub 2016 May 23.

Department of Chemistry and Physics, Florida Gulf Coast University , Fort Myers, Florida 33965, United States.

The discovery of the (+)-α-thujone and (-)-β-thujone stereoisomers in the essential oil of sage (Salvia officinalis L.) and dietary supplements is documented for the first time. The detection was accomplished using a chiral resolution protocol of racemic α-/β-thujone on headspace solid-phase microextraction-gas chromatography-mass spectrometry. Because the previously unreported stereoisomers, (+)-α-thujone and (-)-β-thujone, are not commercially available, a three-step synthesis of racemic thujone from commercially available starting materials was developed. Thermolysis studies demonstrated that no racemization at the cyclopropane stereocenters occurs, corroborating that the detection is not an artifact from the hydrodistillation process. The developed chiral resolution of thujone was also used to provide evidence for the absence of the (+)-α-thujone and (-)-β-thujone enantiomers in other common thujone-containing essential oils.
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http://dx.doi.org/10.1021/acs.jafc.6b01065DOI Listing
June 2016

In vitro cytotoxicity of Artemisia vulgaris L. essential oil is mediated by a mitochondria-dependent apoptosis in HL-60 leukemic cell line.

BMC Complement Altern Med 2014 Jul 7;14:226. Epub 2014 Jul 7.

College of Medicine, King Saud Bin Abdulaziz University for Health Sciences (KSAU-HS), Riyadh, Saudi Arabia.

Background: The essential oil (EO) of Artemisia vulgaris L. has been traditionally used worldwide for treating a large number of diseases. Although major components in A. vulgaris EO have been shown to inhibit growth of different cancer cells, as pure compounds or part of other plants extracted oil, no information is known about its anti-proliferative activities. Therefore, the current investigation has evaluated the toxicity of the plant extracted oil from buds (AVO-b) and leaves (AVO-l) and characterized their growth inhibitory effects on cancer cells.

Methods: AVO-b and AVO-l from A. vulgaris L. were extracted by hydrodistillation, and their effect on the viability of human HL-60 promyelocytic leukemia and various other cancer cell lines was tested using MTT assay. Flow cytometric analysis of apoptosis, DNA fragmentation assay, caspases enzymatic activities and Western blotting were used to determine the apoptotic pathway triggered by their action on HL-60 cells.

Results: Low concentrations of AVO-b and AVO-l inhibited the growth of HL-60 cells in a dose- and time-dependent manner. Employing flow cytometric, DNA fragmentation and caspase activation analyses, demonstrated that the cytotoxic effect of the oils is mediated by a caspase-dependent apoptosis. Kinetic studies in the presence and absence specific caspase inhibitors showed that activation of caspase-8 was dependent and subsequent to the activation of caspases-9 and -3. In addition, the essential oil caused a disruption of the mitochondrial transmembrane potential (ΔΨm), increased the release of cytochrome c to the cytosol, and altered the expression of certain members of Bcl-2 family (Bcl-2, Bax and Bid), Apaf-1 and XIAP. Interestingly, low doses of AVO-b and AVO-1 also induced apoptosis in various cancer cell lines, but not in noncancerous cells.

Conclusions: The results demonstrate that the EO-induced apoptosis in HL-60 cells is mediated by caspase-dependent pathways, involving caspases-3, -9, and -8, which are initiated by Bcl-2/Bax/Bid-dependent loss of ΔΨm leading to release of cytochrome c to the cytoplasm to activate the caspase cascade. The finding that AVO-b and AVO-l are more efficient to induce apoptosis in different cancer cell lines than noncancerous cells, suggests that A. vulgaris might be a promising source for new anticancer agents.
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http://dx.doi.org/10.1186/1472-6882-14-226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4227289PMC
July 2014

Reply to Dr. Bland: despite error in formula, EPIC scale still precise.

J Contin Educ Health Prof 2013 ;33(4):283

Associate Professor, Department of Physical Therapy, University of Toronto.

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http://dx.doi.org/10.1002/chp.21195DOI Listing
April 2015
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