A prospective study of MRI biomarkers in the brain and lower limb muscles for prediction of lower limb motor recovery following stroke.

The aim of this prospective observational longitudinal study was to explore and decipher the predictive value of prospective MRI biomarkers in the brain and lower limb muscles for 3-month lower limb motor recovery following stroke. In the brain, we measured the integrity of the corticospinal tract (fractional anisotropy/"FA"). In the muscles, we measured volume, fatty replacement (fat fraction analysis and proton spectroscopy) and oedema. Measurements were taken at two time points: (1) within 4 weeks of stroke (baseline measurement, clinical and imaging) and (2) 3 months following stroke (follow up measurement, clinical only). Clinical measurements consisted of assessments of functional ability and strength (Fugl-Meyer score, motor NIHSS, Functional Ambulation Category/"FAC", and muscle dynamometry). Twenty-three patients completed imaging and clinical assessments at baseline and follow-up; five patients had partial imaging assessment. The results provided some evidence that damage to the corticospinal tract would result in less motor recovery: recovery of the Fugl-Meyer score and dynamometric ankle plantarflexion, ankle dorsiflexion, and knee extension correlated positively and significantly with fractional anisotropy (0.406-0.457; p = 0.034-p = 0.016). However, fractional anisotropy demonstrated a negative correlation with recovery of the Functional Ambulation Category (-0.359, p = 0.046). For the muscle imaging, significant inverse correlation was observed between vastus lateralis fat fraction vs. NIHSS recovery (-0.401, p = 0.04), and a strong positive correlation was observed between ratio of intra- to extra-myocellular lipid concentrations and the recovery of knee flexion (0.709, p = 0.007). This study supports previous literature indicating a positive correlation between the integrity of the corticospinal tract and motor recovery post-stroke, expanding the limited available literature describing this relationship specifically for the lower limb. However, recovery of functional ambulation behaved differently to other clinical recovery markers by demonstrating an inverse relationship with corticospinal tract integrity. The study also introduces some muscle imaging biomarkers as potentially valuable in the prediction of 3-month lower limb motor recovery following stroke.

Expression of PCOS candidate genes in bovine fetal and adult ovarian somatic cells.

Polycystic ovary syndrome (PCOS) is an endocrine metabolic disorder that appears to have a genetic predisposition and a fetal origin. The fetal ovary has two major somatic cell types shown previously to be of different cellular origins, different morphologies and to differentially express 15 genes. We isolated the somatic gonadal ridge epithelial-like (GREL) cells (n = 7) and ovarian fetal fibroblasts (n = 6) by clonal expansion. Using qRT-PCR, we compared the gene expression levels of PCOS candidate genes with previous data on the expression levels in whole fetal ovaries across gestation. We also compared these levels with those in bovine adult ovarian cells including fibroblasts (n = 4), granulosa cells (n = 5) and surface epithelial cells (n = 5). Adult cell types exhibited clear differences in the expression of most genes. In fetal ovarian cells, DENND1A and ERBB3 had significantly higher expression in GREL cells. HMGA2 and TGFB1I1 tended to have higher expression in fetal fibroblasts than GREL cells. Another 19 genes did not exhibit differences between GREL cells and fetal fibroblasts and FBN3, FSHB, LHCGR, FSHR and ZBTB16 were very lowly expressed in GREL cells and fibroblasts. The culture of fetal fibroblasts in EGF-containing medium resulted in lower expression of NEIL2, but higher expression of MAPRE1 compared to culture in the absence of EGF. Thus, the two fetal ovarian somatic cell types mostly lacked differential expression of PCOS candidate genes.

Auditory rhythmical cueing to improve gait in community-dwelling stroke survivors (ACTIVATE): a pilot randomised controlled trial.

BACKGROUND: Gait impairment limiting mobility and restricting activities is common after stroke. Auditory rhythmical cueing (ARC) uses a metronome beat delivered during exercise to train stepping and early work reports gait improvements. This study aimed to establish the feasibility of a full scale multicentre randomised controlled trial to evaluate an ARC gait and balance training programme for use by stroke survivors in the home and outdoors. METHODS: A parallel-group observer-blind pilot randomised controlled trial was conducted. Adults within 2 years of stroke with a gait-related mobility impairment were recruited from four NHS stroke services and randomised to an ARC gait and balance training programme (intervention) or the training programme without ARC (control). Both programmes consisted of 3x30 min sessions per week for 6 weeks undertaken at home/nearby outdoor community. One session per week was supervised and the remainder self-managed. Gait and balance performance assessments were undertaken at baseline, 6 and 10 weeks. Key trial outcomes included recruitment and retention rates, programme adherence, assessment data completeness and safety. RESULTS: Between November 2018 and February 2020, 59 participants were randomised (intervention n=30, control n=29), mean recruitment rate 4/month. At baseline, 6 weeks and 10 weeks, research assessments were conducted for 59/59 (100%), 47/59 (80%) and 42/59 (71%) participants, respectively. Missing assessments were largely due to discontinuation of data collection from mid-March 2020 because of the UK COVID-19 pandemic lockdown. The proportion of participants with complete data for each individual performance assessment ranged from 100% at baseline to 68% at 10 weeks. In the intervention group, 433/540 (80%) total programme exercise sessions were undertaken, in the control group, 390/522 (75%). Falls were reported by five participants in the intervention group, six in the control group. Three serious adverse events occurred, all unrelated to the study. CONCLUSION: We believe that a definitive multicentre RCT to evaluate the ARC gait and balance training programme is feasible. Recruitment, programme adherence and safety were all acceptable. Although we consider that the retention rate and assessment data completeness were not sufficient for a future trial, this was largely due to the UK COVID-19 pandemic lockdown. TRIAL REGISTRATION: ISRCTN, ISRCTN10874601 , Registered on 05/03/2018.

Isolation, culture, and characterisation of bovine ovarian fetal fibroblasts and gonadal ridge epithelial-like cells and comparison to their adult counterparts.

During ovarian development, gonadal ridge epithelial-like (GREL) cells arise from the epithelial cells of the ventral surface of the mesonephros. They ultimately develop into follicular granulosa cells or into ovarian surface epithelial cells. Stromal fibroblasts arise from the mesonephros and penetrate the ovary. We developed methods for isolating and culturing fetal ovarian GREL cells and ovarian fibroblasts by expansion of colonies without passage. In culture, these two cell types were morphologically different. We examined the expression profile of 34 genes by qRT-PCR, of which 24 genes had previously been studied in whole fetal ovaries. Expression of nine of the 10 newly-examined genes in fetal ovaries correlated with gestational age (MUC1, PKP2, CCNE1 and CCNE2 negatively; STAR, COL4A1, GJA1, LAMB2 and HSD17B1 positively). Comparison between GREL cells and fetal fibroblasts revealed higher expression of KRT19, PKP2, OCLN, MUC1, ESR1 and LGR5 and lower expression of GJA1, FOXL2, NR2F2, FBN1, COL1A1, NR5A1, CCND2, CCNE1 and ALDH1A1. Expression of CCND2, CCNE1, CCNE2, ESR2 and TGFBR1 was higher in the fetal fibroblasts than in adult fibroblasts; FBN1 was lower. Expression of OCLN, MUC1, LAMB2, NR5A1, ESR1, ESR2, and TGFBR3 was lower in GREL cells than ovarian surface epithelial cells. Expression of KRT19, DSG2, PKP2, OCLN, MUC1, FBN1, COL1A1, COL3A1, STAR and TGFBR2 was higher and GJA1, CTNNB1, LAMB2, NR5A1, CYP11A1, HSD3B1, CYP19A1, HSD17B1, FOXL2, ESR1, ESR2, TGFBR3 and CCND2 was lower in GREL cells compared to granulosa cells. TGFß1 altered the expression of COL1A1, COL3A1 and FBN1 in fetal fibroblasts and epidermal growth factor altered the expression of FBN1 and COL1A1. In summary, the two major somatic cell types of the developing ovary have distinct gene expression profiles. They, especially GREL cells, also differ from the cells they ultimately differentiate in to. The regulation of cell fate determination, particularly of the bi-potential GREL cells, remains to be elucidated.

Candidate genes for polycystic ovary syndrome are regulated by TGFß in the bovine foetal ovary.

STUDY QUESTION: Could changes in transforming growth factor ß (TGFß) signalling during foetal ovary development alter the expression of polycystic ovary syndrome (PCOS) candidate genes leading to a predisposition to PCOS? SUMMARY ANSWER: TGFß signalling molecules are dynamically expressed during foetal ovary development and TGFß1 inhibits expression of the androgen receptor (AR) and 7 (INSR, C8H9orf3, RAD50, ERBB3, NEIL2, IRF1 and ZBTB16) of the 25 PCOS candidate genes in foetal ovarian fibroblasts in vitro, whilst increasing expression of the AR cofactor TGFß-induced transcript 1 (TGFB1I1 or Hic5). WHAT IS KNOWN ALREADY: The ovarian stroma arises from the mesonephros during foetal ovary development. Changes in the morphology of the ovarian stroma are cardinal features of PCOS. The ovary is more fibrous and has more tunica and cortical and subcortical stroma. It is not known why this is and when this arises. PCOS has a foetal origin and perhaps ovarian stroma development is altered during foetal life to determine the formation of a polycystic ovary later in life. PCOS also has a genetic origin with 19 loci containing 25 PCOS candidate genes. In many adult tissues, TGFß is known to stimulate fibroblast replication and collagen deposition in stroma, though it has the opposite effect in the non-scaring foetal tissues. Our previous studies showed that TGFß signalling molecules [TGFßs and their receptors, latent TGFß binding proteins (LTBPs) and fibrillins, which are extracellular matrix proteins that bind LTBPs] are expressed in foetal ovaries. Also, we previously showed that TGFß1 inhibited expression of AR and 3 PCOS candidate genes (INSR, C8H9orf3 and RAD50) and stimulated expression of TGFB1I1 in cultured foetal ovarian fibroblasts. STUDY DESIGN, SIZE, DURATION: We used Bos taurus for this study as we can ethically collect foetal ovaries from across the full 9-month gestational period. Foetal ovaries (62-276 days, n = 19) from across gestation were collected from pregnant B. taurus cows for RNA-sequencing (RNA-seq) analyses. Foetal ovaries from B. taurus cows were collected (160-198 days, n = 6) for culture of ovarian fibroblasts. PARTICIPANTS/MATERIALS, SETTING, METHODS: RNA-seq transcriptome profiling was performed on foetal ovaries and the data on genes involved in TGFß signalling were extracted. Cells were dispersed from foetal ovaries and fibroblasts cultured and treated with TGFß1. The effects of TGFß regulation on the remaining eight PCOS candidate genes not previously studied (ERBB3, MAPRE1, FDFT1, NEIL2, ARL14EP, PLGRKT, IRF1 and ZBTB16) were examined. MAIN RESULTS AND THE ROLE OF CHANCE: Many TGFß signalling molecules are expressed in the foetal ovary, and for most, their expression levels increased accross gestation (LTBP1/2/3/4, FBN1, TGFB2/3, TGFBR2/3 and TGFB1I1), while a few decreased (FBN3, TGFBR3L, TGFBI and TGFB1) and others remained relatively constant (TGFBRAP1, TGFBR1 and FBN2). TGFß1 significantly decreased expression of PCOS candidate genes ERBB3, NEIL2, IRF1 and ZBTB16 in cultured foetal ovarian fibroblasts. LARGE SCALE DATA: The FASTQ files, normalized data and experimental information have been deposited in the Gene Expression Omnibus (GEO) accessible by accession number GSE178450. LIMITATIONS, REASONS FOR CAUTION: Regulation of PCOS candidate genes by TGFß was carried out in vitro and further studies in vivo are required. This study was carried out in bovine where foetal ovaries from across all of the 9-month gestational period were available, unlike in the human where it is not ethically possible to obtain ovaries from the second half of gestation. WIDER IMPLICATIONS OF THE FINDINGS: From our current and previous results we speculate that inhibition of TGFß signalling in the foetal ovary is likely to (i) increase androgen sensitivity by enhancing expression of AR, (ii) increase stromal activity by stimulating expression of COL1A1 and COL3A1 and (iii) increase the expression of 7 of the 25 PCOS candidate genes. Thus inhibition of TGFß signalling could be part of the aetiology of PCOS or at least the aetiology of polycystic ovaries. STUDY FUNDING/COMPETING INTEREST(S): Funding was received from Adelaide University China Fee Scholarship (M.L.), Australian Research Training Program (R.A.) and the Faculty of Health and Medical Science Divisional Scholarship (R.A.), Adelaide Graduate Research Scholarships (R.A. and N.A.B.), Australia Awards Scholarship (M.D.H.), Robinson Research Institute Career Development Fellowship (K.H.) and Building On Ideas Grant (K.H.), National Health and Medical Research Council of Australia Centre for Research Excellence in the Evaluation, Management and Health Care Needs of Polycystic Ovary Syndrome (N.A.B., M.D.H. and R.J.R.; GTN1078444) and the Centre for Research Excellence on Women's Health in Reproductive life (R.A., R.J.R. and K.H.; GTN1171592) and the UK Medical Research Council (R.A.A.; grant no. G1100357). The funders did not play any role in the study design, data collection and analysis, decision to publish or preparation of the manuscript. The authors of this manuscript have nothing to declare and no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.

Acceptability and deliverability of an auditory rhythmical cueing (ARC) training programme for use at home and outdoors to improve gait and physical activity post-stroke.

BACKGROUND: Although laboratory studies demonstrate that training programmes using auditory rhythmical cueing (ARC) may improve gait post-stroke, few studies have evaluated this intervention in the home and outdoors where deployment may be more appropriate. This manuscript reports stakeholder refinement of an ARC gait and balance training programme for use at home and outdoors, and a study which assessed acceptability and deliverability of this programme. METHODS: Programme design and content were refined during stakeholder workshops involving physiotherapists and stroke survivors. A two-group acceptability and deliverability study was then undertaken. Twelve patients post-stroke with a gait related mobility impairment received either the ARC gait and balance training programme or the gait and balance training programme without ARC. Programme provider written notes, participant exercise and fall diaries, adverse event monitoring and feedback questionnaires captured data about deliverability, safety and acceptability of the programmes. RESULTS: The training programme consisted of 18 sessions (six supervised, 12 self-managed) of exercises and ARC delivered by a low-cost commercially available metronome. All 12 participants completed the six supervised sessions and 10/12 completed the 12 self-managed sessions. Provider and participant session written records and feedback questionnaires confirmed programme deliverability and acceptability. CONCLUSION: An ARC gait and balance training programme refined by key stakeholders was feasible to deliver and acceptable to participants and providers. TRIAL REGISTRATION: ISCTRN 12/03/2018.

Morphine Resistance in Spinal Cord Injury-Related Neuropathic Pain in Rats is Associated With Alterations in Dopamine and Dopamine-Related Metabolomics.

Opioids are not universally effective for treating neuropathic pain following spinal cord injury (SCI), a finding that we previously demonstrated in a rat model of SCI. The aim of this study was to determine analgesic response of morphine-responsive and nonresponsive SCI rats to adjunct treatment with dopamine modulators and to establish if the animal groups expressed distinct metabolomic profiles. Thermal thresholds were tested in female Long Evans rats (N = 45) prior to contusion SCI, after SCI and following injection of morphine, morphine combined with dopamine modulators, or dopamine modulators alone. Spinal cord and striatum samples were processed for metabolomics and targeted mass spectrometry. Morphine provided analgesia in 1 of 3 of SCI animals. All animals showed improved analgesia with morphine + pramipexole (D3 receptor agonist). Only morphine nonresponsive animals showed improved analgesia with the addition of SCH 39166 (D1 receptor antagonist). Metabolomic analysis identified 3 distinct clusters related to the tyrosine pathway that corresponded to uninjured, SCI morphine-responsive and SCI morphine-nonresponsive groups. Mass spectrometry showed matching differences in dopamine levels in striatum and spinal cord between these groups. The data suggest an overall benefit of the D3 receptor system in improving analgesia, and an association between morphine responsiveness and metabolomic changes in the tyrosine/dopamine pathways in striatum and spinal cord. PERSPECTIVE: Spinal cord injury (SCI) leads to opioid-resistant neuropathic pain that is associated with changes in dopamine metabolomics in the spinal cord and striatum of rats. We present evidence that adjuvant targeting of the dopamine system may be a novel pain treatment approach to overcome opioid desensitization and tolerance after SCI.

Economic evaluation of robot-assisted training versus an enhanced upper limb therapy programme or usual care for patients with moderate or severe upper limb functional limitation due to stroke: results from the RATULS randomised controlled trial.

OBJECTIVE: To determine whether robot-assisted training is cost-effective compared with an enhanced upper limb therapy (EULT) programme or usual care. DESIGN: Economic evaluation within a randomised controlled trial. SETTING: Four National Health Service (NHS) centres in the UK: Queen's Hospital, Barking, Havering and Redbridge University Hospitals NHS Trust; Northwick Park Hospital, London Northwest Healthcare NHS Trust; Queen Elizabeth University Hospital, NHS Greater Glasgow and Clyde; and North Tyneside General Hospital, Northumbria Healthcare NHS Foundation Trust. PARTICIPANTS: 770 participants aged 18 years or older with moderate or severe upper limb functional limitation from first-ever stroke. INTERVENTIONS: Participants randomised to one of three programmes provided over a 12-week period: robot-assisted training plus usual care; the EULT programme plus usual care or usual care. MAIN ECONOMIC OUTCOME MEASURES: Mean healthcare resource use; costs to the NHS and personal social services in 2018 pounds; utility scores based on EQ-5D-5L responses and quality-adjusted life years (QALYs). Cost-effectiveness reported as incremental cost per QALY and cost-effectiveness acceptability curves. RESULTS: At 6 months, on average usual care was the least costly option (£3785) followed by EULT (£4451) with robot-assisted training being the most costly (£5387). The mean difference in total costs between the usual care and robot-assisted training groups (£1601) was statistically significant (p<0.001). Mean QALYs were highest for the EULT group (0.23) but no evidence of a difference (p=0.995) was observed between the robot-assisted training (0.21) and usual care groups (0.21). The incremental cost per QALY at 6 months for participants randomised to EULT compared with usual care was £74 100. Cost-effectiveness acceptability curves showed that robot-assisted training was unlikely to be cost-effective and that EULT had a 19% chance of being cost-effective at the £20 000 willingness to pay (WTP) threshold. Usual care was most likely to be cost-effective at all the WTP values considered in the analysis. CONCLUSIONS: The cost-effectiveness analysis suggested that neither robot-assisted training nor EULT, as delivered in this trial, were likely to be cost-effective at any of the cost per QALY thresholds considered. TRIAL REGISTRATION NUMBER: ISRCTN69371850.

Analysing the Action Research Arm Test (ARAT): a cautionary tale from the RATULS trial.

Many studies of stroke rehabilitation use the Action Research Arm Test (ARAT) as an outcome, which measures upper limb function by scoring the ability to complete functional tasks. This report describes an issue encountered when analysing the ARAT subscales in a trial of upper limb therapies after stroke. The subscales of the ARAT at three months followed a 'U-shaped' distribution, and therefore, comparing means or medians was not appropriate. A simple alternative approach was chosen that dichotomised the subscales. When analysing the ARAT, the shape of the distributions must be checked in order to choose the most appropriate descriptive and inferential statistical techniques. In particular, if the data follows a 'U-shaped' distribution, a simple dichotomising or a more sophisticated approach is needed. These should also be considered for heavily skewed distributions, often arising from substantial floor or ceiling effects. Inappropriate analyses can lead to misleading conclusions.

Evaluation of the enhanced upper limb therapy programme within the Robot-Assisted Training for the Upper Limb after Stroke trial: descriptive analysis of intervention fidelity, goal selection and goal achievement.

OBJECTIVE: To report the fidelity of the enhanced upper limb therapy programme within the Robot-Assisted Training for the Upper Limb after stroke (RATULS) randomized controlled trial, the types of goals selected and the proportion of goals achieved. DESIGN: Descriptive analysis of data on fidelity, goal selection and achievement from an intervention group within a randomized controlled trial. SETTING: Out-patient stroke rehabilitation within four UK NHS centres. SUBJECTS: 259 participants with moderate-severe upper limb activity limitation (Action Research Arm Test 0-39) between one week and five years post first stroke. INTERVENTION: The enhanced upper limb therapy programme aimed to provide 36 one-hour sessions, including 45 minutes of face-to-face therapy focusing on personal goals, over 12 weeks. RESULTS: 7877/9324 (84%) sessions were attended; a median of 34 [IQR 29-36] per participant. A median of 127 [IQR 70-190] repetitions were achieved per participant per session attended. Based upon the Canadian Occupational Performance Measure, goal categories were: self-care 1449/2664 (54%); productivity 374/2664 (14%); leisure 180/2664 (7%) and 'other' 661/2664 (25%). For the 2051/2664 goals for which data were available, 1287 (51%) were achieved, ranging between 27% by participants more than 12 months post stroke with baseline Action Research Arm Test scores 0-7, and 88% by those less than three months after stroke with scores 8-19. CONCLUSIONS: Intervention fidelity was high. Goals relating to self-care were most commonly selected. The proportion of goals achieved varied, depending on time post stroke and baseline arm activity limitation.

Robot-assisted training compared with an enhanced upper limb therapy programme and with usual care for upper limb functional limitation after stroke: the RATULS three-group RCT.

BACKGROUND: Loss of arm function is common after stroke. Robot-assisted training may improve arm outcomes. OBJECTIVE: The objectives were to determine the clinical effectiveness and cost-effectiveness of robot-assisted training, compared with an enhanced upper limb therapy programme and with usual care. DESIGN: This was a pragmatic, observer-blind, multicentre randomised controlled trial with embedded health economic and process evaluations. SETTING: The trial was set in four NHS trial centres. PARTICIPANTS: Patients with moderate or severe upper limb functional limitation, between 1 week and 5 years following first stroke, were recruited. INTERVENTIONS: Robot-assisted training using the Massachusetts Institute of Technology-Manus robotic gym system (InMotion commercial version, Interactive Motion Technologies, Inc., Watertown, MA, USA), an enhanced upper limb therapy programme comprising repetitive functional task practice, and usual care. MAIN OUTCOME MEASURES: The primary outcome was upper limb functional recovery 'success' (assessed using the Action Research Arm Test) at 3 months. Secondary outcomes at 3 and 6 months were the Action Research Arm Test results, upper limb impairment (measured using the Fugl-Meyer Assessment), activities of daily living (measured using the Barthel Activities of Daily Living Index), quality of life (measured using the Stroke Impact Scale), resource use costs and quality-adjusted life-years. RESULTS: A total of 770 participants were randomised (robot-assisted training, n = 257; enhanced upper limb therapy, n = 259; usual care, n = 254). Upper limb functional recovery 'success' was achieved in the robot-assisted training [103/232 (44%)], enhanced upper limb therapy [118/234 (50%)] and usual care groups [85/203 (42%)]. These differences were not statistically significant; the adjusted odds ratios were as follows: robot-assisted training versus usual care, 1.2 (98.33% confidence interval 0.7 to 2.0); enhanced upper limb therapy versus usual care, 1.5 (98.33% confidence interval 0.9 to 2.5); and robot-assisted training versus enhanced upper limb therapy, 0.8 (98.33% confidence interval 0.5 to 1.3). The robot-assisted training group had less upper limb impairment (as measured by the Fugl-Meyer Assessment motor subscale) than the usual care group at 3 and 6 months. The enhanced upper limb therapy group had less upper limb impairment (as measured by the Fugl-Meyer Assessment motor subscale), better mobility (as measured by the Stroke Impact Scale mobility domain) and better performance in activities of daily living (as measured by the Stroke Impact Scale activities of daily living domain) than the usual care group, at 3 months. The robot-assisted training group performed less well in activities of daily living (as measured by the Stroke Impact Scale activities of daily living domain) than the enhanced upper limb therapy group at 3 months. No other differences were clinically important and statistically significant. Participants found the robot-assisted training and the enhanced upper limb therapy group programmes acceptable. Neither intervention, as provided in this trial, was cost-effective at current National Institute for Health and Care Excellence willingness-to-pay thresholds for a quality-adjusted life-year. CONCLUSIONS: Robot-assisted training did not improve upper limb function compared with usual care. Although robot-assisted training improved upper limb impairment, this did not translate into improvements in other outcomes. Enhanced upper limb therapy resulted in potentially important improvements on upper limb impairment, in performance of activities of daily living, and in mobility. Neither intervention was cost-effective. FUTURE WORK: Further research is needed to find ways to translate the improvements in upper limb impairment seen with robot-assisted training into improvements in upper limb function and activities of daily living. Innovations to make rehabilitation programmes more cost-effective are required. LIMITATIONS: Pragmatic inclusion criteria led to the recruitment of some participants with little prospect of recovery. The attrition rate was higher in the usual care group than in the robot-assisted training or enhanced upper limb therapy groups, and differential attrition is a potential source of bias. Obtaining accurate information about the usual care that participants were receiving was a challenge. TRIAL REGISTRATION: Current Controlled Trials ISRCTN69371850. FUNDING: This project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 24, No. 54. See the NIHR Journals Library website for further project information.

Many people who have arm weakness following a stroke feel that insufficient attention is paid by rehabilitation services to recovery of their arm. Unfortunately, it is currently unclear how best to provide rehabilitation to optimise recovery, but robot-assisted training and therapy programmes that focus on practising functional tasks are promising and require further evaluation. The Robot-Assisted Training for the Upper Limb after Stroke (RATULS) trial evaluated three approaches to rehabilitation for people with moderate or severe difficulty using their arm. These approaches were robot-assisted training using the Massachusetts Institute of Technology-Manus robotic gym system (InMotion commercial version, Interactive Motion Technologies, Inc., Watertown, MA, USA), an enhanced upper limb therapy programme based on repetitive practice of functional tasks and usual care. Robot-assisted training and the enhanced upper limb therapy programme were provided in an outpatient setting for 45 minutes per session, three times per week, for 12 weeks, in addition to usual care. The Massachusetts Institute of Technology-Manus robotic gym system was selected as it was felt to be the best available technology. The participant sits at a table, places their affected arm onto the Massachusetts Institute of Technology-Manus arm support and attempts to move their arm to play a game on the computer screen. Movements are assisted by the Massachusetts Institute of Technology-Manus if the patient cannot perform the movements themselves. The results of the RATULS trial show that robot-assisted training did not result in additional improvement in stroke survivors' arm use when compared with the enhanced upper limb therapy programme or usual care. Stroke survivors who received enhanced upper limb therapy experienced meaningful improvements in undertaking activities of daily living, when compared with those participants who received either robot-assisted training or usual care. Participants who received enhanced upper limb therapy also experienced benefits in their mobility, compared with usual care participants. Participants and therapists found both therapies acceptable, and described various benefits. A health economic analysis found that neither robot-assisted training nor the enhanced upper limb therapy programme was a cost-effective treatment for the NHS.

Analysis of expression of candidate genes for polycystic ovary syndrome in adult and fetal human and fetal bovine ovaries†.

Polycystic ovary syndrome (PCOS) appears to have a genetic predisposition and a fetal origin. We compared the expression levels of 25 PCOS candidate genes from adult control and PCOS human ovaries (n = 16) using microarrays. Only one gene was potentially statistically different. Using qRT-PCR, expression of PCOS candidate genes was examined in bovine fetal ovaries from early stages when they first developed stroma through to completion of development (n = 27; 60-270 days of gestation). The levels of ERBB3 mRNA negatively correlated with gestational age but positively with HMGA2, FBN3, TOX3, GATA4, and DENND1A.X1,2,3,4, previously identified as correlated with each other and expressed early. PLGRKT and ZBTB16, and less so IRF1, were also correlated with AMH, FSHR, AR, INSR, and TGFB1I1, previously identified as correlated with each other and expressed late. ARL14EP, FDFT1, NEIL2, and MAPRE1 were expressed across gestation and not correlated with gestational age as shown previously for THADA, ERBB4, RAD50, C8H9orf3, YAP1, RAB5B, SUOX, and KRR1. LHCGR, because of its unusual bimodal expression pattern, had some unusual correlations with other genes. In human ovaries (n = 15; <150 days of gestation), ERBB3.V1 and ERBB3.VS were expressed and correlated negatively with gestational age and positively with FBN3, HMGA2, DENND1A.V1,3,4, DENND1A.V1-7, GATA4, and FSHR, previously identified as correlated with each other and expressed early. Thus, the general lack of differential expression of candidate genes in adult ovaries contrasting with dynamic patterns of gene expression in fetal ovaries is consistent with a vulnerability to disturbance in the fetal ovary that may underpin development of PCOS.

An extended stroke rehabilitation service for people who have had a stroke: the EXTRAS RCT.

BACKGROUND: There is limited evidence about the effectiveness of rehabilitation in meeting the longer-term needs of stroke patients and their carers. OBJECTIVE: To determine the clinical effectiveness and cost-effectiveness of an extended stroke rehabilitation service (EXTRAS). DESIGN: A pragmatic, observer-blind, parallel-group, multicentre randomised controlled trial with embedded health economic and process evaluations. Participants were randomised (1 : 1) to receive EXTRAS or usual care. SETTING: Nineteen NHS study centres. PARTICIPANTS: Patients with a new stroke who received early supported discharge and their informal carers. INTERVENTIONS: Five EXTRAS reviews provided by an early supported discharge team member between 1 and 18 months post early supported discharge, usually over the telephone. Reviewers assessed rehabilitation needs, with goal-setting and action-planning. Control treatment was usual care post early supported discharge. MAIN OUTCOME MEASURES: The primary outcome was performance in extended activities of daily living (Nottingham Extended Activities of Daily Living Scale) at 24 months post randomisation. Secondary outcomes at 12 and 24 months included patient mood (Hospital Anxiety and Depression Scale), health status (Oxford Handicap Scale), experience of services and adverse events. For carers, secondary outcomes included carers' strain (Caregiver Strain Index) and experience of services. Cost-effectiveness was estimated using resource utilisation costs (adaptation of the Client Service Receipt Inventory) and quality-adjusted life-years. RESULTS: A total of 573 patients (EXTRAS, n = 285; usual care, n = 288) with 194 carers (EXTRAS, n = 103; usual care, n = 91) were randomised. Mean 24-month Nottingham Extended Activities of Daily Living Scale scores were 40.0 (standard deviation 18.1) for EXTRAS (n = 219) and 37.2 (standard deviation 18.5) for usual care (n = 231), giving an adjusted mean difference of 1.8 (95% confidence interval -0.7 to 4.2). The mean intervention group Hospital Anxiety and Depression Scale scores were not significantly different at 12 and 24 months. The intervention did not improve patient health status or carer strain. EXTRAS patients and carers reported greater satisfaction with some aspects of care. The mean cost of resource utilisation was lower in the intervention group: -£311 (95% confidence interval -£3292 to £2787), with a 68% chance of EXTRAS being cost-saving. EXTRAS was associated with 0.07 (95% confidence interval 0.01 to 0.12) additional quality-adjusted life-years. At current conventional thresholds of willingness to pay for a quality-adjusted life-year, there is a 90% chance that EXTRAS is cost-effective. CONCLUSIONS: EXTRAS did not improve stroke survivors' performance in extended activities of daily living but did improve their overall satisfaction with services. Given the impact on costs and quality-adjusted life-years, there is a high chance that EXTRAS could be considered cost-effective. FUTURE WORK: Further research is required to identify whether or not community-based interventions can improve performance of extended activities of daily living, and to understand the improvements in health-related quality of life and costs seen by provision of intermittent longer-term specialist review. TRIAL REGISTRATION: Current Controlled Trials ISRCTN45203373. FUNDING: This project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 24, No. 24. See the NIHR Journals Library website for further project information.

Early supported discharge enables stroke patients with mild or moderate disability to be discharged earlier than usual from hospital to continue rehabilitation at home. Randomised controlled trials have demonstrated that early supported discharge leads to increased independence for stroke survivors, and that early supported discharge is cost-effective. Early supported discharge is usually provided for up to 6 weeks and patients with ongoing physical, psychological or social needs are then referred to other services. In the UK, provision of longer-term rehabilitation is often limited. Lack of research evidence has meant that service development in this aspect of stroke care has lagged behind service development for acute care. This clinical trial evaluated an extended stroke rehabilitation service (EXTRAS) that started when early supported discharge ended. Stroke survivors and their carers were randomly assigned to receive EXTRAS or usual NHS care. EXTRAS involved five rehabilitation reviews conducted over 18 months by an early supported discharge team member, usually over the telephone. Each review consisted of an assessment of current needs, goal-setting and action-planning, and sought to improve patients' abilities and confidence to undertake extended activities of daily living (mobility, kitchen and domestic tasks, and leisure activities). There were no specific assessments or actions for carers but it was important to evaluate the impact that the new service had on carers. Patients and carers were followed up for 2 years and information was collected about their activities, mood, quality of life and services received. EXTRAS did not improve stroke survivors' performance in extended activities of daily living. However, patients who received EXTRAS reported less anxiety and less depression than those who received usual care, and patients and carers were more satisfied with some aspects of their care. EXTRAS did not improve carers' quality of life or stress. Health economic analyses suggest that EXTRAS improved patients' quality of life and may be good value for money. Further research is needed to identify other treatments to address the longer-term consequences of stroke.

Auditory rhythmical cueing to improve gait and physical activity in community-dwelling stroke survivors (ACTIVATE): study protocol for a pilot randomised controlled trial.

BACKGROUND: Mobility problems are present in 70-80% of stroke survivors and can result in impaired gait and reduced physical activity limiting independent living. Auditory rhythmic cueing (ARC) has been used to provide auditory feedback and shows promise in improving a variety of walking parameters following stroke. The aim of this pilot study is to assess the feasibility of conducting a multi-centre, observer blind, randomised controlled trial of auditory rhythmical cueing (ARC) intervention in home and community settings in North East England. METHODS: This pilot observer blind randomised controlled feasibility trial aims to recruit 60 participants over 15 months from community stroke services in the North East of England. Participants will be within 24 months of stroke onset causing new problems with mobility. Each participant will be randomised to the study intervention or control group. Intervention treatment participants will undertake 18 auditory rhythmical cueing (ARC) treatment sessions over 6 weeks (3 × 30 min per week, 6 supervised (physiotherapist/research associate)/12 self-managed) in a home/community setting. A metronome will be used to provide ARC during a series of balance and gait exercises, which will be gradually progressed. The control treatment participants will undertake the same duration balance and gait exercise training programme as the intervention group but without the ARC. Feasibility will be determined in terms of recruitment, retention, adverse events, adherence, collection of descriptive clinical and accelerometer motor performance data at baseline, 6 weeks and 10 weeks and description of participant, provider and clinical therapists' experiences. As well as using questionnaires to collate participant views, qualitative interviews will be undertaken to further understand how the intervention is delivered in practice in a community setting and to identify aspects perceived important by participants. DISCUSSION: The ACTIVATE study will address an important gap in the evidence base by reporting whether it is feasible to deliver auditory rhythmical cueing in the home and community to improve gait and balance parameters following stroke. The feasibility of the study protocol will be established and results will inform the design of a future multi-centre randomised controlled trial. TRIAL REGISTRATION: Trial register: ISRCTN, Trial identifier: ISRCTN10874601: Date of registration: 12/03/2018.

Positive predictive value of stroke identification by ambulance clinicians in North East England: a service evaluation.

INTRODUCTION/BACKGROUND: Accurate prehospital identification of patients who had an acute stroke enables rapid conveyance to specialist units for time-dependent treatments such as thrombolysis and thrombectomy. Misidentification leads to patients who had a 'stroke mimic' (SM) being inappropriately triaged to specialist units. We evaluated the positive predictive value (PPV) of prehospital stroke identification by ambulance clinicians in the North East of England. METHODS: This service evaluation linked routinely collected records from a UK regional ambulance service identifying adults with any clinical impression of suspected stroke to diagnostic data from four National Health Service hospital trusts between 1 June 2013 and 31 May 2016. The reference standard for a confirmed stroke diagnosis was inclusion in Sentinel Stroke National Audit Programme data or a hospital diagnosis of stroke or transient ischaemic attack in Hospital Episode Statistics. PPV was calculated as a measure of diagnostic accuracy. RESULTS: Ambulance clinicians in North East England identified 5645 patients who had a suspected stroke (mean age 73.2 years, 48% male). At least one Face Arm Speech Test (FAST) symptom was documented for 93% of patients who had a suspected stroke but a positive FAST was only documented for 51%. Stroke, or transient ischaemic attack, was the final diagnosis for 3483 (62%) patients. SM (false positives) accounted for 38% of suspected strokes identified by ambulance clinicians and included a wide range of non-stroke diagnoses including infections, seizures and migraine. DISCUSSION: In this large multisite data set, identification of patients who had a stroke by ambulance clinicians had a PPV rate of 62% (95% CI 61 to 63). Most patients who had a suspected stroke had at least one FAST symptom, but failure to document a complete test was common. Training for stroke identification and SM rates need to be considered when planning service provision and capacity.

Effect of an Enhanced Paramedic Acute Stroke Treatment Assessment on Thrombolysis Delivery During Emergency Stroke Care: A Cluster Randomized Clinical Trial.

Importance: Rapid thrombolysis treatment for acute ischemic stroke reduces disability among patients who are carefully selected, but service delivery is challenging. Objective: To determine whether an enhanced Paramedic Acute Stroke Treatment Assessment (PASTA) intervention increased hospital thrombolysis rates. Design, Setting, and Participants: This multicenter, cluster randomized clinical trial took place between December 2015 and July 2018 in 3 ambulance services and 15 hospitals. Clusters were paramedics based within ambulance stations prerandomized to PASTA or standard care. Patients attended by study paramedics were enrolled after admission if a hospital specialist confirmed a stroke and paramedic assessment started within 4 hours of onset. Allocation to PASTA or standard care reflected the attending paramedic's randomization status. Interventions: The PASTA intervention included additional prehospital information collection, a structured hospital handover, practical assistance up to 15 minutes after handover, a predeparture care checklist, and clinician feedback. Standard care reflected national guidelines. Main Outcomes and Measures: Primary outcome was the proportion of patients receiving thrombolysis. Secondary outcomes included time intervals and day 90 health (with poor status defined as a modified Rankin Score >2, to represent dependency or death). Results: A total of 11 478 patients were screened following ambulance transportation; 1391 were eligible and approached, but 177 did not consent. Of 1214 patients enrolled (mean [SD] age, 74.7 [13.2] years; 590 women [48.6%]), 500 were assessed by 242 paramedics trained in the PASTA intervention and 714 were assessed by 355 paramedics continuing with standard care. The paramedics trained in the PASTA intervention took a mean of 13.4 (95% CI, 9.4-17.4) minutes longer (P < .001) to complete patient care episodes. There was less thrombolysis among the patients in the PASTA group, but this was not significant (PASTA group, 197 of 500 patients [39.4%] vs the standard care group, 319 of 714 patients [44.7%]; adjusted odds ratio, 0.81 [95% CI, 0.61-1.08]; P = .15). Time from a paramedic on scene to thrombolysis was a mean of 8.5 minutes longer in the PASTA group (98.1 [37.6] minutes) vs the standard care group (89.4 [31.1] minutes; P = .01). Poor health outcomes did not differ significantly but occurred less often among patients in the PASTA group (313 of 489 patients [64.0%]) vs the standard care group (461 of 690 patients [66.8%]; adjusted odds ratio, 0.86 [95% CI, 0.60-1.20]; P = .39). Conclusions and Relevance: An enhanced paramedic assessment did not facilitate thrombolysis delivery. The unexpected combination of thrombolysis and health outcomes suggests possible alternative influences on treatment decisions by the intervention, requiring further evaluation. Trial Registration: ISRCTN Registry Identifier: ISRCTN12418919.

Dopamine D1 or D3 receptor modulators prevent morphine tolerance and reduce opioid withdrawal symptoms.

The long-term treatment of chronic pain by opioids is limited by tolerance and risk of addiction/dependence. Previously, we have shown that combination treatment of morphine with a dopamine D1 or D3 receptor modulator restored morphine analgesia in morphine-resistant neuropathic pain and decreased morphine's reward potential in an acute setting. Here, we investigated whether such adjunct therapy with a dopamine D1 receptor preferring antagonist (SCH 39166) or a dopamine D3 receptor preferring agonist (pramipexole) could prevent morphine tolerance and reduce withdrawal symptoms. Initially, tolerance to the combination of morphine + pramipexole was assessed in mice. Mice receiving intraperitoneal injections of morphine showed reduced thermal thresholds on Day 7 whereas those receiving morphine + pramipexole maintained analgesia at Day 7. Next, tolerance and withdrawal to both combinations were tested over 14 days in rats. Rats were assigned one of four drug conditions, (1) saline, 2) morphine, 3) morphine + SCH 39166, 4) morphine + pramipexole), for chronic administration via osmotic pumps. Chronic administration of morphine over 14 days resulted in a significant reduction of morphine analgesia. However, analgesia was maintained when morphine was administered with either the dopamine D1 receptor preferring antagonist or the D3 receptor preferring agonist. Withdrawal symptoms peaked at 48 h and were decreased in rats receiving either combination compared to morphine alone. The data suggests that adjunct therapy with dopamine D1 or D3 receptor preferring modulators prevents morphine tolerance and reduces the duration of morphine withdrawal symptoms, and thus this combination has potential for long-term pain management therapy.

Could perturbed fetal development of the ovary contribute to the development of polycystic ovary syndrome in later life?

Polycystic ovary syndrome (PCOS) affects around 10% of young women, with adverse consequences on fertility and cardiometabolic outcomes. PCOS appears to result from a genetic predisposition interacting with developmental events during fetal or perinatal life. We hypothesised that PCOS candidate genes might be expressed in the fetal ovary when the stroma develops; mechanistically linking the genetics, fetal origins and adult ovarian phenotype of PCOS. In bovine fetal ovaries (n = 37) of 18 PCOS candidate genes only SUMO1P1 was not expressed. Three patterns of expression were observed: early gestation (FBN3, GATA4, HMGA2, TOX3, DENND1A, LHCGR and FSHB), late gestation (INSR, FSHR, and LHCGR) and throughout gestation (THADA, ERBB4, RAD50, C8H9orf3, YAP1, RAB5B, SUOX and KRR1). A splice variant of FSHB exon 3 was also detected early in the bovine ovaries, but exon 2 was not detected. Three other genes, likely to be related to the PCOS aetiology (AMH, AR and TGFB1I1), were also expressed late in gestation. Significantly within each of the three gene groups, the mRNA levels of many genes were highly correlated with each other, despite, in some instances, being expressed in different cell types. TGFß is a well-known stimulator of stromal cell replication and collagen synthesis and TGFß treatment of cultured fetal ovarian stromal cells inhibited the expression of INSR, AR, C8H9orf3 and RAD50 and stimulated the expression of TGFB1I1. In human ovaries (n = 15, < 150 days gestation) many of the same genes as in bovine (FBN3, GATA4, HMGA2, FSHR, DENND1A and LHCGR but not TOX3 or FSHB) were expressed and correlated with each other. With so many relationships between PCOS candidate genes during development of the fetal ovary, including TGFß and androgen signalling, we suggest that future studies should determine if perturbations of these genes in the fetal ovary can lead to PCOS in later life.

Asymmetrical Bioimpedance in the Anterior Circulation for Urgent Stratification of suspected Stroke (ABACUS Stroke): study protocol for a diagnostic accuracy study.

BACKGROUND: Early identification and treatment of stroke improve outcome. Ischaemic stroke due to large vessel occlusion (LVO) benefits from time-critical thrombectomy but this is only available in highly specialised healthcare services. Cerebral Bioimpedance Asymmetry (CBA) measurement obtained with the portable and rapid Cerebrotech Visor™ System device may be able to identify certain types of stroke including LVO. This test could be deployed pre-hospital and used to immediately direct patients to the most appropriate healthcare service for treatment. This study is evaluating the diagnostic accuracy of CBA measurements obtained from a real-world population of suspected stroke. METHODS: Study design: Prospective observational cohort study.Setting: A hyperacute stroke unit and neuroscience centre in North East England.Participants: Adults with a paramedic assigned diagnosis of suspected stroke arriving at hospital within 6 hours of symptom onset.Index Test: Cerebral Bioimpedance Asymmetry measurement performed using the Cerebrotech Visor™ System. Measurement values produce continuous data (range 0 -100); pre-defined threshold for positive state ≥ 10.Reference Standard Tests: Standard CT brain +/- CT/MR angiography, and expert clinician opinion will establish the following clinical outcomes which constitute the suspected stroke population: ischaemic stroke +/- large vessel occlusion; symptomatic severe anterior vessel stenosis; large (≥60ml) and small (<60mls) vessel intracerebral haemorrhage; transient ischaemic attack; stroke mimic conditions; prior territorial stroke.Analyses: Sensitivity, specificity, negative and positive predictive values, area under the Receiver Operating Characteristic curve for identification of i) "complex stroke" (ischaemic stroke with large vessel occlusion or symptomatic severe anterior vessel stenosis or intracerebral haemorrhage ≥60ml or prior territorial stroke) and ii) ischaemic stroke with large vessel occlusion in isolation.Sample size: 124 participants. DISCUSSION: The results from this study will determine how accurately CBA measurement using the Cerebrotech Visor™ System can identify key stroke types within the suspected stroke population. Acceptable diagnostic performance would be an important step forwards for access to time-critical treatments. TRIAL REGISTRATION: Registered with ISRCTN (identifier: ISRCTN79169844) on 06/08/2018.

Small fibre pathology in chronic whiplash-associated disorder: A cross-sectional study.

BACKGROUND: Mechanisms underpinning ongoing symptoms in chronic whiplash associated-disorder (WAD) are not well understood. People with chronic WAD can exhibit sensory dysfunction consistent with small nerve fibre pathology, including thermal hypoaesthesia and hyperalgesia. This study investigated small fibre structure and function in chronic WAD. METHODS: Twenty-four people with chronic WAD (median [IQR] age 49 [15] years, 16 females) and 24 pain-free controls (50 [17] years, 16 females) were recruited. Intraepidermal nerve fibre density (IENFD) and dermal innervation were assessed by skin biopsy. This was performed at (a) the lateral index finger on the primary side of pain and (b) superior to the lateral malleolus on the contralateral side. Quantitative sensory testing was performed over the hand. RESULTS: The WAD group exhibited lower IENFD at the finger (WAD: median [IQR] 4.5 [4.9] fibres/mm; control 7.3 [3.9]; p = .010), but not the ankle (WAD: mean [SD] 7.3 [3.7] fibres/mm; control 9.3 [3.8]; p = .09). Dermal innervation was lower in the WAD group at the finger (WAD: median [IQR] 3.7 [2.8] nerve bundles/mm2 ; controls: 4.9 [2.1]; p = .017) but not the ankle (WAD: median [IQR] 2.1 [1.9] nerve bundles/mm2 ; controls: 1.8 [1.8]; p = .70). In the WAD group, hand thermal and light touch detection were impaired, and heat pain thresholds were lowered (p ≤ .037). CONCLUSIONS: Findings suggest small fibre structural and functional deficits in chronic WAD, implicating potential involvement of small fibre pathology. SIGNIFICANCE: Our study found decreased intraepidermal nerve fibre density, reduced dermal innervation, thermal hypoaesthesia and hypersensitivity in people with chronic WAD, suggestive of small fibre pathology. This observation of peripheral nervous system pathology in chronic whiplash provides novel insights on mechanisms underpinning symptoms and challenges commonly held beliefs regarding this condition.