RNA- and ATAC-sequencing Reveals a Unique CD83+ Microglial Population Focally Depleted in Parkinson's Disease.

All brain areas affected in Parkinson's disease (PD) show an abundance of microglia with an activated morphology together with increased expression of pro-inflammatory cytokines, suggesting that neuroinflammation may contribute to the neurodegenerative process in this common and incurable disorder. We applied a single nucleus RNA- and ATAC-sequencing approach using the 10x Genomics Chromium platform to postmortem PD samples to investigate microglial heterogeneity in PD. We created a multiomic dataset using substantia nigra (SN) tissues from 19 PD donors and 14 non-PD controls (NPCs), as well as three other brain regions from the PD donors which are differentially affected in this disease: the ventral tegmental area (VTA), substantia inominata (SI), and hypothalamus (HypoTs). We identified thirteen microglial subpopulations within these tissues as well as a perivascular macrophage and a monocyte population, of which we characterized the transcriptional and chromatin repertoires. Using this data, we investigated whether these microglial subpopulations have any association with PD and whether they have regional specificity. We uncovered several changes in microglial subpopulations in PD, which appear to parallel the magnitude of neurodegeneration across these four selected brain regions. Specifically, we identified that inflammatory microglia in PD are more prevalent in the SN and differentially express PD-associated markers. Our analysis revealed the depletion of a CD83 and HIF1A- expressing microglial subpopulation, specifically in the SN in PD, that has a unique chromatin signature compared to other microglial subpopulations. Interestingly, this microglial subpopulation has regional specificity to the brainstem in non-disease tissues. Furthermore, it is highly enriched for transcripts of proteins involved in antigen presentation and heat-shock proteins, and its depletion in the PD SN may have implications for neuronal vulnerability in disease.

Value, transparency, and inclusion: A values-based study of patient involvement in musculoskeletal research.

BACKGROUND: Patient and public involvement work (PPI) is essential to good research practice. Existing research indicates that PPI offers benefits to research design, conduct, communication, and implementation of findings. Understanding how PPI works and its value helps to provide information about best practice and highlight areas for further development. This study used a values-based approach to reporting PPI at a Research Unit focused on musculoskeletal conditions within a UK medical school. METHODS: The study was conducted between October 2019 and January 2020 using Gradinger's value system framework as a theoretical basis. The framework comprises three value systems each containing five clusters. All PPI members and researchers who had attended PPI groups were invited to participate. Participants completed a structured questionnaire based on the value system framework; PPI members also provided further information through telephone interviews. Data were deductively analysed using a framework approach with data mapped onto value systems. RESULTS: Twelve PPI members and 17 researchers took part. Views about PPI activity mapped onto all three value systems. PPI members felt empowered to provide their views, and that their opinions were valued by researchers. It was important to PPI members that they were able to 'give back' and to do something positive with their experiences. Researchers would have liked the groups to be more representative of the wider population, patients highlighted that groups could include more younger members. Researchers recognised the value of PPI, and the study highlighted areas where researchers members might benefit from further awareness. CONCLUSIONS: Three areas for development were identified: (i) facilitating researcher engagement in training about the value and importance of PPI in research; (ii) support for researchers to reflect on the role that PPI plays in transparency of healthcare research; (iii) work to further explore and address aspects of diversity and inclusion in PPI.

Long-term outcome of sacral nerve stimulation for faecal incontinence.

AIM: Sacral nerve stimulation (SNS) is a minimally invasive treatment for faecal incontinence (FI). We report our experience of patients who have undergone SNS for FI with a minimum of 5 years' follow-up. This is a single centre prospective observational study with the aim to assess the long-term function of SNS. METHOD: All patients implanted with SNS were identified from our prospective database. The date of implantation, first and last clinic follow-up, surgical complications and St Mark's incontinence scores were abstracted and analysed. RESULTS: From 1996 to 2014, 381 patients were considered for SNS. Of these, 256 patients met the study inclusion criteria. Median age at implantation was 52 years (range 18-81). The ratio of women to men was 205:51. Indications were urge FI (25%), passive FI (17.9%) and mixed FI (57%). The median of the incontinence score at baseline was 19/24 and this improved to 7/24 at the 6-month follow-up. Of the total cohort, 235 patients received a medium-term follow-up (median 110 months, range 12-270) with a median continence score of 10/24 which was also confirmed at the telephone long-term follow-up on 185 patients (132 months, range 60-276). CONCLUSION: This study demonstrates that SNS is an effective treatment in the long term. SNS results in an improvement of validated scores for approximately 60% of patients; however, there is a significant reduction of efficacy over time due to underlying causes.

Acceptability, effectiveness and safety of a Renew® anal insert in patients who have undergone restorative proctocolectomy with ileal pouch-anal anastomosis.

INTRODUCTION: Restorative proctocolectomy has gained acceptance in the surgical management of medically refractive ulcerative colitis and cancer prevention in familial adenomatous polyposis. Incontinence following restorative proctocolectomy occurs in up to 25% of patients overnight. The Renew® insert is an inert single-use device which acts as an anal plug. The aim of this study was to assess the acceptability, effectiveness and safety of the Renew® insert in patients who have undergone restorative proctocolectomy. The device has yet to be assessed in patients who have undergone restorative proctocolectomy. METHOD: This was a prospective study exploring the acceptability, effectiveness and safety of the Renew® insert in improving incontinence in patients who had undergone restorative proctocolectomy. A total of 15 patients with incontinence were asked to use the Renew® insert for 14 days following their standard care. The Incontinence Questionnaire-Bowels was used pre- and posttreatment to assess response and patients were asked to report the perceived acceptability, effectiveness and safety of the device at the end of the trial. RESULTS: The device was acceptable to 8/15 (53%) of patients and was effective in 6/15 (40%). Only 2/15 (13%) of patients raised any safety concerns, and these were minor. The device was associated with a significant reduction in night seepage (P = 0.034). CONCLUSION: In a small study, the Renew® insert can be both acceptable and effective and is also associated with few safety concerns. It is also associated with significant reductions in night-time seepage.

Early changes in Achilles tendon behaviour in vivo following downhill backwards walking.

Downhill backwards walking causes repeated, cyclical loading of the muscle-tendon unit. The effect this type of repeated loading has on the mechanical behaviour of the Achilles tendon is presently unknown. This study aimed to investigate the biomechanical response of the Achilles tendon aponeurosis complex following a downhill backwards walking protocol. Twenty active males (age: 22.3 ± 3.0 years; mass: 74.7 ± 5.6 kg; height: 1.8 ± 0.7 m) performed 60 min of downhill (8.5°), backwards walking on a treadmill at -0.67 m · s(-1). Data were collected before, immediately post, and 24-, 48- and 168-h post-downhill backwards walking. Achilles tendon aponeurosis elongation, strain and stiffness were measured using ultrasonography. Muscle force decreased immediately post-downhill backward walking (P = 0.019). There were increases in Achilles tendon aponeurosis stiffness at 24-h post-downhill backward walking (307 ± 179.6 N · mm(-1), P = 0.004), and decreases in Achilles tendon aponeurosis strain during maximum voluntary contraction at 24 (3.8 ± 1.7%, P = 0.008) and 48 h (3.9 ± 1.8%, P = 0.002) post. Repeated cyclical loading of downhill backwards walking affects the behaviour of the muscle-tendon unit, most likely by altering muscle compliance, and these changes result in tendon stiffness increases.

A review of sacral nerve stimulation for faecal incontinence following rectal surgery and radiotherapy.

AIM: Faecal incontinence may occur following rectal surgery and/or radiotherapy for rectal cancer. The aim of this paper was to review the evidence to support the use of sacral nerve stimulation (SNS) for patients with incontinence who had undergone rectal surgery or received rectal radiotherapy. METHOD: A search was performed of PubMed, Medline and Embase. All studies which reported the outcome of SNS in patients who had undergone a rectal resection or radiotherapy were reviewed. RESULTS: The first report of SNS following rectal surgery was in 2002. Since then seven further studies have described its effect in patients who have undergone anterior resection or pelvic radiotherapy. The total number of patients was 57. All studies were single group series, which ranged in size from one to 15 patients. The follow-up ranged from 1 to 36 months. The success of peripheral nerve evaluation ranged from 47% to 100%. Permanent SNS improved the symptoms and in some studies this was reflected in improved quality of life. The wide variation of patient factors, operations performed, the dose of radiotherapy given and time from operation makes interpretation of the results difficult. CONCLUSION: Larger studies with better patient selection are needed to investigate the effect of SNS on incontinence following radiotherapy or rectal surgery.

A double-blinded randomized multicentre study to investigate the effect of changes in stimulation parameters on sacral nerve stimulation for constipation.

AIM: Sacral nerve stimulation (SNS) may be offered to patients with constipation who have failed to improve with conservative treatment. The response to SNS is variable, with a significant loss of efficacy in some patients. An increased frequency of stimulation may improve the efficacy of SNS for faecal incontinence. This study aimed to see if alteration of the pulse width or frequency improved the outcome for those with constipation. METHOD: Eleven patients with constipation currently being treated by SNS were recruited from three centres. They were randomized to five different protocols of stimulation each applied for 5 weeks. Group 1 used standard settings (pulse width 210 µs, frequency 14 Hz); in the other four groups (Groups 2-5) the pulse width and/or frequency were halved or doubled. Patients and investigators were blinded to the group allocation. RESULTS: The Cleveland Clinic constipation score varied significantly between the five groups. Group 1 achieved the lowest score mean (± SD) 13.4 (± 4.4) (P = 0.03). The number of digitations per defaecation was the lowest in Group 4, 90 µs and 14 Hz (P < 0.01). No other variable changed significantly. Standard settings were the most preferred by the recruited patients. CONCLUSION: Alteration of pulse width or frequency of stimulation had no significant effect on the outcome of SNS for constipation.

A pilot study to compare daily with twice weekly transcutaneous posterior tibial nerve stimulation for faecal incontinence.

AIM: Posterior tibial nerve stimulation (PTNS) has been shown to improve faecal incontinence in the short term. The optimal treatment regimen is unclear with wide variations in protocol reported in the literature. The study aimed to assess two different regimens of transcutaneous PTNS and to establish whether increasing the frequency of stimulation increases the effectiveness. METHOD: Thirty patients were randomized to receive once daily or twice weekly PTNS for a 6-week period. The treatment was carried out by the patient at home after instruction. The primary investigator was blinded to the patient allocation until the study had ended, at which point the symptoms were assessed. No further stimulation was given after 6 weeks and the patients were followed until their symptoms returned to the pre-stimulation state (baseline). The primary outcome measure was a change in the frequency of incontinent episodes. RESULTS: Three patients in the daily group and none in the twice weekly group achieved complete continence. Only patients from the daily group showed a significant reduction in median (interquartile range) incontinent episodes per week from 5 (11.13) to 3.5 (4.31) (P = 0.025). There was no significant change in the frequency of defaecation nor in the ability to defer defaecation. Patients in the daily group experienced a significant improvement in the domains of lifestyle [2.2 (1.7) to 2.6 (1.65), P = 0.04] and embarrassment [1.7 (0.85) to 2.15 (0.4), P = 0.04] on the Rockwood Fecal Incontinence Quality of Life assessment. No adverse events were reported. CONCLUSION: Transcutaneous PTNS can safely be used by the patient at home. Daily treatment may be more effective than twice weekly treatment. Larger studies are needed to investigate this further.

Immunological biomarkers: catalysts for translational advances in autoimmune diabetes.

In a recent workshop organized by the JDRF focused on the 'Identification and Utilization of Robust Biomarkers in Type1 Diabetes', leaders in the field of type 1 diabetes (T1D)/autoimmunity and assay technology came together from academia, government and industry to assess the current state of the field, evaluate available resources/technologies and identify gaps that need to be filled for moving the field of T1D research forward. The highlights of this workshop are discussed in this paper, as well as the proposal for a larger, planned consortium effort, incorporating a JDRF Biomarker Core, to foster collaboration and accelerate progress in this critically needed area of T1D research.

Randomized double-blind crossover study of alternative stimulator settings in sacral nerve stimulation for faecal incontinence.

BACKGROUND: Sacral nerve stimulation is an established treatment for faecal incontinence. Nearly half of all patients experience loss of efficacy at some point. Standard reprogramming restores efficacy for some, but not all, patients. This study aimed to determine whether alternative stimulator settings would increase treatment efficacy. METHODS: Patients with sustained loss of efficacy were recruited from two international specialist centres. A number of alternative stimulation parameters were tested using a double-blind randomized crossover study design. Stimulation settings tested were pulse frequencies of 6.9 and 31 Hz, and pulse widths of 90 and 330 µs, compared with one standard setting of 14 Hz/210 µs. Treatment efficacy was evaluated using a disease-specific quality-of-life score (Fecal Incontinence Quality of Life Scale, FIQLS) and a bowel habit diary completed before randomization, during the study period and after 3 months of follow-up with one preferred setting. RESULTS: Fifteen patients were analysed. With one preferred setting, three of four subdomains in the FIQLS improved significantly. The mean(s.d.) total number of incontinence episodes dropped from 11.7(10.8) to 4.8(4.5) per 3 weeks (P = 0.011) and improvements were maintained after 3 months of follow-up. Optimal pacemaker settings were individual, but a trend towards highest patient satisfaction and improved treatment outcome was evident for high-frequency stimulation (31 Hz/210 µs), which was preferred by eight of the 15 patients. CONCLUSION: Patients experiencing loss of efficacy can experience improvement if alternative pacemaker settings are tested. High-frequency stimulation (31 Hz/210 µs) was preferred by more than half of the patients, and improved treatment outcome was sustained at 3 months.

Feasibility of implementing pulse oximetry screening for congenital heart disease in a community hospital.

OBJECTIVE: Pulse oximetry has been recognized as a promising screening tool for critical congenital heart disease (CCHD). The aim of this research was to study the feasibility of implementation in a community hospital setting. STUDY DESIGN: Meetings were conducted to determine an implementation plan. Pulse oximetry was performed on the right hand and foot after 24 h of age. Newborns with a saturation ≤ 95% or a ≥ 3% difference were considered to have a positive screen. Screening barriers, screening time and ability to effectively screen all eligible newborns were noted. RESULT: From January 2009 through May 2010, of 6841 eligible newborns, 6745 newborns (98.6%) were screened. Of the nine infants with positive pulse oximetry screens, one had CCHD, four had CHD and four others were determined to have false positive screens. Average screening time was 3.5 min (0 to 35 min). CONCLUSION: Pulse oximetry can be implemented successfully in community hospitals without an excessive number of false positives or additional nursing staff.

Plasma cells in muscle in inclusion body myositis and polymyositis.

BACKGROUND: Previous immunohistochemical studies of muscle from patients with inclusion body myositis and polymyositis found many more T cells than B cells, suggesting a role for intramuscular cell-mediated immune mechanisms rather than humoral mechanisms. METHODS: Microarray studies were performed on muscle biopsy specimens from 40 patients with inclusion body myositis (IBM; n = 23), polymyositis (PM; n = 6), and without neuromuscular disease (n = 11). Reverse transcription PCR of selected immunoglobulin gene transcripts was performed on two patient samples. Qualitative immunohistochemical studies for B-cell lineage cell surface markers were performed on 28 muscle specimens and quantitative studies performed on a subset of 19 untreated patients with IBM or PM. CD138+ cells were isolated from muscle using laser capture microdissection, and immunoglobulin transcripts were PCR amplified to determine the presence or absence of immunoglobulin gene rearrangements unique to the B-cell lineage. RESULTS: Immunoglobulin gene transcripts accounted for 59% in IBM and 33% in PM of the most stringently defined highest differentially expressed muscle transcripts compared with normal. Plasma cells, terminally differentiated B cells expressing CD138 but not CD19 or CD20, are present in IBM and PM muscle in numbers several times higher than B cells. CONCLUSIONS: There are differentiated B cells in the form of CD138+ plasma cells within the muscle of patients with inclusion body myositis and polymyositis. The principle of linked recognition of B-cell activation predicts several strategies for autoantigen discovery that could not otherwise be pursued through the study of the infiltrating T-cell population alone.

Follicular dendritic cell tumour in a 9-year-old child.

Follicular dendritic cell tumour (FDCT) or sarcoma is a rare tumour first described in 1986. Some 80 cases have been reported, the youngest being in teenagers. Our patient first presented at 9 years of age with a cervical mass that was removed and revealed an apparently benign, but florid reactive process. At age 14 the lump recurred and biopsy was diagnostic of FDCT. Radical block dissection showed disease to level III and 6 weeks of radiotherapy was followed by 6 months adjuvant chemotherapy. Three years after completing his final treatment he shows no signs of recurrent disease.

Mapping QTLs for resistance to the cyst nematode Globodera pallida derived from the wild potato species Solanum vernei.

Resistance to the potato cyst nematode (PCN) species Globodera pallida, derived from the wild diploid potato species Solanum vernei, has been investigated. This source of resistance, which is effective against all of the major pathotypes of G. pallida and Globodera rostochiensis, has been assumed to be due to several genetic factors, but it has proved difficult to deploy effectively in breeding strategies for potato cultivars. Diploid and tetraploid potato populations segregating for 'vernei' resistance were analysed. At the tetraploid level, a bulk segregant analysis (BSA) approach was employed and detected AFLP markers linked to a resistance QTL on potato linkage group V. Conventional linkage analysis of a diploid population identified QTL on linkage groups V and IX. A marker linked to a QTL on linkage group V has been converted to a single-locus PCR-based marker, which can be used to detect the presence of the QTL in diploid and tetraploid potato germplasm. Moreover, there is evidence that one of the AFLPs detected by BSA appears to be specific to an introgressed segment of DNA from S. vernei. These results are compared with those obtained from other studies on resistance to the PCN species G. pallida.

Effects of approach velocity and foot-target characteristics on the visual regulation of step length.

Two questions emerge from the literature concerning the perceptual-motor processes underlying the visual regulation of step length. The first concerns the effects of velocity on the onset of visual control (VCO), when visual regulation of step length begins during goal-directed locomotion. The second concerns the effects of different obstacles such as a target or raised surface on step length regulation. In two separate experiments, participants (Experiment 1 & 2: n=12, 6 female, 6 male) walked, jogged, or sprinted towards an obstacle along a 10 m walkway, consisting of two marker-strips with alternating black and white 0.50 m markings. Each experiment consisted of three targeting or obstacle tasks with the requirement to both negotiate and continue moving (run-through) through the target. Five trials were conducted for each task and approach speed, with trials block randomised between the six participants of each gender. One 50 Hz video camera panned and filmed each trial from an elevated position, adjacent to the walkway. Video footage was digitized to deduce the gait characteristics. Results for the targeting tasks indicate a linear relationship between approach velocity and accuracy of final foot placement (r=0.89). When foot placement was highly constrained by the obstacle step length shortened during the entire approach. VCO was found to occur at an earlier tau-margin for lower approach velocities for both experiments, indicating that the optical variable 'tau' is affected by approach velocity. A three-phase kinematic profile was found for all tasks, except for the take-off board condition when sprinting. Further research is needed to determine whether this velocity affect on VCO is due to 'whole-body' approach velocity or whether it is a function of the differences between gait modes.

Sediment evidence of early eutrophication and heavy metal pollution of Lake Mälaren, central Sweden.

Lake Mälaren is the water supply and recreation area for more than 1 million people in central Sweden and subject to considerable environmental concern. To establish background data for assessments of contemporary levels of trophy and heavy metal pollution, sediment cores from the lake were analyzed. Diatom-inferred lake-water phosphorus concentrations suggest that pre-20th century nutrient levels in Södra Björkfjärden, a basin in the eastern part of Mälaren, were higher (c. 10-20 micrograms TP L-1) than previously assumed (c. 6 micrograms TP L-1). Stable lead isotope and lead concentration analyses from 3 basins (S. Björkfjärden, Gisselfjärden and Asköfjärden) show that the lake was polluted in the 19th century and earlier from extensive metal production and processing in the catchment, particularly in the Bergslagen region. The lake has experienced a substantial improvement of the lead pollution situation in the 20th century following closure of the mining and metal industry. The lead pollution from the old mining industry was large compared to late-20th century pollution from car emissions, burning of fossil fuels and modern industries.

Unusual 1H NMR chemical shifts support (His) C(epsilon) 1...O==C H-bond: proposal for reaction-driven ring flip mechanism in serine protease catalysis.

13C-selective NMR, combined with inhibitor perturbation experiments, shows that the C(epsilon)(1)H proton of the catalytic histidine in resting alpha-lytic protease and subtilisin BPN' resonates, when protonated, at 9.22 ppm and 9.18 ppm, respectively, which is outside the normal range for such protons and approximately 0.6 to 0.8 ppm further downfield than previously reported. They also show that the previous alpha-lytic protease assignments [Markley, J. L., Neves, D. E., Westler, W. M., Ibanez, I. B., Porubcan, M. A. & Baillargeon, M. W. (1980) Front. Protein Chem. 10, 31-61] were to signals from inactive or denatured protein. Simulations of linewidth vs. pH demonstrate that the true signal is more difficult to detect than corresponding signals from inactive derivatives, owing to higher imidazole pK(a) values and larger chemical shift differences between protonated and neutral forms. A compilation and analysis of available NMR data indicates that the true C(epsilon)(1)H signals from other serine proteases are similarly displaced downfield, with past assignments to more upfield signals probably in error. The downfield displacement of these proton resonances is shown to be consistent with an H-bond involving the histidine C(epsilon)(1)H as donor, confirming the original hypothesis of Derewenda et al. [Derewenda, Z. S., Derewenda, U. & Kobos, P. M. (1994) J. Mol. Biol. 241, 83-93], which was based on an analysis of literature x-ray crystal structures of serine hydrolases. The invariability of this H-bond among enzymes containing Asp-His-Ser triads indicates functional importance. Here, we propose that it enables a reaction-driven imidazole ring flip mechanism, overcoming a major dilemma inherent in all previous mechanisms, namely how these enzymes catalyze both the formation and productive breakdown of tetrahedral intermediates.

Physiological effects of hunting red deer (Cervus elaphus).

When red deer (Cervus elaphus) were hunted by humans with hounds the average distance travelled was at least 19 km. This study of 64 hunted red deer provides the first empirical evidence on their state at the time of death. Blood and muscle samples obtained from hunted deer after death were compared with samples from 50 non-hunted red deer that had been cleanly shot with rifles. The effects on deer of long hunts were (i) depletion of carbohydrate resources for powering muscles, (ii) disruption of muscle tissue, and (iii) elevated secretion of beta-endorphin. High concentrations of cortisol, typically associated with extreme physiological and psychological stress, were found. Damage to red blood cells occurred early in the hunts; possible mechanisms are discussed. Taken together, the evidence suggests that red deer are not well-adapted by their evolutionary or individual history to cope with the level of activity imposed on them when hunted with hounds.