Pili of pathogenic Neisseria are major virulence factors associated with adhesion, twitching motility, auto-aggregation, and DNA transformation. Pili of N. meningitidis are subject to several ...different post-translational modifications. Among these pilin modifications, the presence of phosphorylcholine (ChoP) and a glycan on the pilin protein are phase-variable (subject to high frequency, reversible on/off switching of expression). In this study we report the location of two ChoP modifications on the C-terminus of N. meningitidis pilin. We show that the surface accessibility of ChoP on pili is affected by phase variable changes to the structure of the pilin-linked glycan. We identify for the first time that the platelet activating factor receptor (PAFr) is a key, early event receptor for meningococcal adherence to human bronchial epithelial cells and tissue, and that synergy between the pilin-linked glycan and ChoP post-translational modifications is required for pili to optimally engage PAFr to mediate adherence to human airway cells.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
IntroductionThe REFORM (REhabilitation FOR Musculoskeletal conditions) trial is a non-inferiority randomised controlled trial (n=210) designed to determine whether a supported home exercise programme ...is as good or better than a course of face-to-face physiotherapy for the management of some musculoskeletal conditions. The trial is currently being conducted across Sydney government hospitals in Australia. This process evaluation will run alongside the REFORM trial. It combines qualitative and quantitative data to help explain the trial results and determine the feasibility of rolling out supported home exercise programmes in settings similar to the REFORM trial.Methods and analysisTwo theoretical frameworks underpin our process evaluation methodology: the Realist framework (context, mechanism, outcomes) considers the causal assumptions as to why a supported home exercise programme may be as good or better than face-to-face physiotherapy in terms of the context, mechanisms and outcomes of the trial. The RE-AIM framework describes the Reach, Effectiveness, Adoption, Implementation and Maintenance of the intervention. These two frameworks will be broadly used to guide this process evaluation using a mixed-methods approach. For example, qualitative data will be derived from interviews with patients, healthcare professionals and stakeholders, and quantitative data will be collected to determine the cost and feasibility of providing supported home exercise programmes. These data will be analysed iteratively before the analysis of the trial results and will be triangulated with the results of the primary and secondary outcomes.Ethics and disseminationThis trial will be conducted in accordance with the National Health and Medical Research Council National Statement on Ethical Conduct in Human Research (2018) and the Note for Good Clinical Practice (CPMP/ICH-135/95). Ethical approval was obtained on 17 March 2017 from the Northern Sydney Local Health District Human Research Ethics Committee (trial number: HREC/16HAWKE/431-RESP/16/287) with an amendment for the process evaluation approved on 4 February 2020. The results of the process evaluation will be disseminated through publications in peer-reviewed journals and presentations at scientific conferences.Trial registration numberACTRN12619000065190.
Electronic sensor devices can provide an objective assessment of soft tissue balancing in total knee arthroplasty (TKA) which may potentially decrease postoperative pain. We aim to quantify the ...learning curve for operative time (OT) for this technology.
Consecutive TKA cases balanced with an electronic sensor balancing device by one senior surgeon from 2013 to 2017 were included in this study. The OT (in minutes) was analyzed using the cumulative sum analysis to evaluate the learning curve for this technology. Further analysis was done by splitting the 287 patients into 7 cohorts, 41 patients each.
Two hundred eighty-seven patients balanced with sensor technology were available for analysis. The cumulative sum OT learning curve estimated that this technology’s learning curve was 41 cases. This curve consisted of 2 phases: phase 1 which includes the first 41 cases and phase 2 which includes the remaining 246 patients. The mean OT for the first and last sensor-assisted cohorts was 120.4 and 108.9 minutes (P = .021). The mean OT for the first sensor-assisted cohort and the control cohort was 120.4 versus 109 minutes (P = .023). The mean OT for the last sensor-assisted cohort and the control cohort was 108.9 versus 109 minutes (P = .94).
Our findings suggest that it takes approximately 41 cases of sensor-assisted TKA cases to achieve OTs identical to manually balanced TKA cases. This is a relatively shallow learning curve for the sensor technology, and allows arthroplasty surgeons to objectively achieve soft tissue balancing without adding OT to the surgery.
Uropathogenic
(UPEC) is the major cause of urinary tract infections. Nearly half of all UPEC strains secrete hemolysin, a cytotoxic pore-forming toxin. Here, we show that the prevalence of the ...hemolysin toxin gene (
) is highly variable among the most common 83
sequence types (STs) represented on the EnteroBase genome database. To explore this diversity in the context of a defined monophyletic lineage, we contextualized sequence variation of the
operon within the genealogy of the globally disseminated multidrug-resistant ST131 clone. We show that sequence changes in
and its newly defined 1.616-kb-long leader sequence correspond to phylogenetic designation, and that ST131 strains with the strongest hemolytic activity belong to the most extensive multidrug-resistant sublineage (clade C2). To define the set of genes involved in hemolysin production, the clade C2 strain S65EC was completely sequenced and subjected to a genome-wide screen by combining saturated transposon mutagenesis and transposon-directed insertion site sequencing with the capacity to lyse red blood cells. Using this approach, and subsequent targeted mutagenesis and complementation, 13 genes were confirmed to be specifically required for production of active hemolysin. New hemolysin-controlling elements included discrete sets of genes involved in lipopolysaccharide (LPS) inner core biosynthesis (
,
,
, and
) and cytoplasmic chaperone activity (
and
), and we show these are required for hemolysin secretion. Overall, this work provides a unique description of hemolysin sequence diversity in a single clonal lineage and describes a complex multilevel system of regulatory control for this important toxin.
Uropathogenic
(UPEC) is the major cause of urinary tract infections and a frequent cause of sepsis. Nearly half of all UPEC strains produce the potent cytotoxin hemolysin, and its expression is associated with enhanced virulence. In this study, we explored hemolysin variation within the globally dominant UPEC ST131 clone, finding that strains from the ST131 sublineage with the greatest multidrug resistance also possess the strongest hemolytic activity. We also employed an innovative forward genetic screen to define the set of genes required for hemolysin production. Using this approach, and subsequent targeted mutagenesis and complementation, we identified new hemolysin-controlling elements involved in LPS inner core biosynthesis and cytoplasmic chaperone activity, and we show that mechanistically they are required for hemolysin secretion. These original discoveries substantially enhance our understanding of hemolysin regulation, secretion and function.
Recessive mutations in
, an endoplasmic reticulum (ER)-resident BiP co-chaperone, have been identified in patients with multisystemic neurodegeneration and diabetes mellitus. To further unravel these ...pathomechanisms, we employed a non-biased proteomic approach and identified dysregulation of several key cellular pathways, suggesting a pathophysiological interplay of perturbed lipid metabolism, mitochondrial bioenergetics, ER-Golgi function, and amyloid-beta processing. Further functional investigations in fibroblasts of patients with
mutations detected cellular accumulation of lipids and an increased sensitivity to cholesterol stress, which led to activation of the unfolded protein response (UPR), alterations of the ER-Golgi machinery, and a defect of amyloid precursor protein. In line with the results of previous studies, we describe here alterations in mitochondrial morphology and function, as a major contributor to the
pathophysiology. Hence, we propose that the loss of
affects lipid/cholesterol homeostasis, leading to UPR activation, β-amyloid accumulation, and impairment of mitochondrial oxidative phosphorylation.
Salmonella enterica serovar Typhimurium possesses a multi-copper-ion oxidase (multicopper oxidase), CueO (also known as CuiD), a periplasmic enzyme known to be required for resistance to copper ions. ...CueO from S. Typhimurium was expressed as a recombinant protein in Escherichia coli, and the purified protein exhibited a high cuprous oxidase activity. We have characterized an S. Typhimurium cueO mutant and confirmed that it is more sensitive to copper ions. Using a murine model of infection, it was observed that the cueO mutant was significantly attenuated, as indicated by reduced recovery of bacteria from liver and spleen, although there was no significant difference in recovery from Peyer's patches and mesenteric lymph nodes. However, the intracellular survival of the cueO mutant in unprimed or gamma-interferon-primed murine macrophages was not statistically different from that of wild-type Salmonella, suggesting that additional host factors are involved in clearance of the cueO mutant. Unlike a cueO mutant from E. coli, the S. Typhimurium cueO mutant did not show greater sensitivity to hydrogen peroxide and its sensitivity to copper ions was not affected by siderophores. Similarly, the S. Typhimurium cueO mutant was not rescued from copper ion toxicity by addition of the branched-chain amino acids and leucine.
Background An arteriovenous fistula (AVF) is the preferred vascular access for hemodialysis, offering lower morbidity, mortality, and cost compared with grafts or catheters. Patients with a difficult ...access extremity have often lost all superficial veins, and even basilic veins may be obliterated. We have used brachial vein transposition AVFs (BVT-AVFs) in these challenging patients and review our experience in this report. Methods The study reviewed consecutive patients in whom BVT-AVFs were created from September 2006 to March 2009. Most BVT-AVFs were created in staged procedures, with the second-stage transposition operations completed 4 to 6 weeks after the first-stage AVF operation. A single-stage BVT-AVF was created when the brachial vein diameter was ≥6 mm. Results We identified 58 BVT-AVF procedures, comprising 41 women (71.0%), 28 diabetic patients (48.3%), and 29 (50.0%) had previous access surgery. The operation was completed in two stages in 45 operations (77.6%) and was a primary transposition in 13 patients. However, five of these were secondary AVFs with previous distal AV grafts or AVFs placed elsewhere; effectively, late staged procedures. Follow-up was a mean of 11 months (range, 2.0-31.7 months). Primary patency, primary-assisted patency, and cumulative (secondary) patency were 52.0%, 84.9%, and 92.4% at 12 months and 46.2%, 75.5%, and 92.4% at 24 months, respectively. Harvesting the brachial vein was tedious and more difficult than harvesting other superficial veins. No prosthetic grafts were used. Conclusion BVT-AVFs provide a suitable option for autogenous access when the basilic vein is absent in patients with difficult access extremities. Most patients required intervention for access maturation or maintenance. Most BVT-AVFs were created with staged procedures. Cumulative (secondary) patency was 92.4% at 24 months.
Charcot-Marie-Tooth type 2 (CMT2) neuropathy is characterised by a vast clinical and genetic heterogeneity complicating its diagnosis and therapeutic intervention. Identification of molecular ...signatures that are common to multiple CMT2 subtypes can aid in developing therapeutic strategies and measuring disease outcomes.
A proteomics-based approach was performed on lymphoblasts from CMT2 patients genetically diagnosed with different gene mutations to identify differentially regulated proteins. The candidate proteins were validated through real-time quantitative PCR and western blotting on lymphoblast samples of patients and controls, motor neurons differentiated from patient-derived induced pluripotent stem cells (iPSCs) and sciatic nerves of CMT2 mouse models.
Proteomic profiling of patient lymphoblasts resulted in the identification of profilin 2 (PFN2) and guanidinoacetate methyltransferase (GAMT) as commonly downregulated proteins in different genotypes compared with healthy controls. This decrease was also observed at the transcriptional level on screening 43 CMT2 patients and 22 controls, respectively. A progressive decrease in PFN2 expression with age was observed in patients, while in healthy controls its expression increased with age. Reduced PFN2 expression was also observed in motor neurons differentiated from CMT2 patient-derived iPSCs and sciatic nerves of CMT2 mice when compared with controls. However, no change in GAMT levels was observed in motor neurons and CMT2 mouse-derived sciatic nerves.
We unveil PFN2 and GAMT as molecular determinants of CMT2 with possible indications of the role of PFN2 in the pathogenesis and disease progression. This is the first study describing biomarkers that can boost the development of therapeutic strategies targeting a wider spectrum of CMT2 patients.
Polymerization‐induced self‐assembly (PISA) enables the scalable synthesis of functional block copolymer nanoparticles with various morphologies. Herein we exploit this versatile technique to produce ...so‐called “high χ–low N” diblock copolymers that undergo nanoscale phase separation in the solid state to produce sub‐10 nm surface features. By varying the degree of polymerization of the stabilizer and core‐forming blocks, PISA provides rapid access to a wide range of diblock copolymers, and enables fundamental thermodynamic parameters to be determined. In addition, the pre‐organization of copolymer chains within sterically‐stabilized nanoparticles that occurs during PISA leads to enhanced phase separation relative to that achieved using solution‐cast molecularly‐dissolved copolymer chains.
Polymerization‐induced self‐assembly enables the scalable synthesis of functional block copolymer nanoparticles with various morphologies. This versatile technique is exploited to prepare so‐called „high χ–low N“ diblock copolymers that undergo nanoscale phase separation in the solid state to produce sub‐10 nm domains.
Background The use of catheters or prosthetic grafts for vascular access has significantly higher mortality and morbidity risks, in addition to higher costs, than arteriovenous fistulas (AVF). Many ...patients have a difficult access extremity due to complex medical illnesses, previous vascular access procedures, intravenous catheters, diabetes, vascular disease, female sex, age, and other complicating factors. Transposition AVFs (AVF-T) have been used for these individuals to avoid catheters and grafts. We report our experience with primary and staged basilic vein AVF-Ts and staged brachial vein AVF-Ts. Methods From our database of consecutive vascular access operations, we reviewed patients from May 2003 to September 2006 for all upper extremity AVF-Ts. A primary AVF-T was used when the basilic vein was continuous with a minimum diameter of 4 mm and of adequate length. When the basilic vein was 2.5 to 4 mm, the procedure was staged. The proximal radial artery was used for inflow, if possible. When the basilic vein was not suitable, a radial vein or brachial vein anastomosis was performed as the first stage of a planned brachial vein AVF-T. The second stage operations of staged AVF-Ts were generally done 4 to 6 weeks after the primary AVF construction. All patients were evaluated with preoperative ultrasound imaging by the operating surgeon. Results From a database of 412 consecutive vascular access patients, 78 upper extremity transposition procedures were identified. Of these, 57 patients (73.1%) were women, 44 (56.4%) were diabetic, and 46 (59.0%) had previous access surgery. Fifty-eight operations were staged procedures. The basilic vein was used in 68 AVF-T, the brachial vein in six, and cephalic vein in four. The anastomosis was based on the proximal radial artery in 60 patients. Mean follow-up was 18 months (range, 3-48 months). Primary patency, primary assisted patency, and cumulative patency were 45.7%, 93.5%, and 96.0% at 12 months and 27.6%, 86.5%, and 88.9% at 24 months, respectively. No prosthetic grafts were used in the study period. Conclusion Both primary and staged AVF-T procedures were successfully used in patients with difficult access extremities. AVF-Ts were durable, although many required an interventional procedure for maturation or maintenance. Cumulative (secondary) patency was 96.0% at 12 months and 88.9% at 24 months. The absence of an adequate basilic vein does not preclude the use of a staged AVF-T because the brachial vein offers a suitable alternative.
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