Spectroscopic, biochemical and computational studies of bioactive DNA minor groove binders targeting 5'-WGWWCW-3' motif.

Spectroscopic, biochemical, and computational modelling studies have been used to assess the binding capability of a set of minor groove binding (MGB) ligands against the self-complementary DNA sequences 5'-d(CGCACTAGTGCG)-3' and 5'-d(CGCAGTACTGCG)-3'. The ligands were carefully designed to target the DNA response element, 5'-WGWWCW-3', the binding site for several nuclear receptors. Basic 1D 1H NMR spectra of the DNA samples prepared with three MGB ligands show subtle variations suggestive of how each ligand associates with the double helical structure of both DNA sequences. The variations among the investigated ligands were reflected in the line shape and intensity of 1D 1H and 31P-{1H} NMR spectra. Rapid visual inspection of these 1D NMR spectra proves to be beneficial in providing valuable insights on MGB binding molecules. The NMR results were consistent with the findings from both UV DNA denaturation and molecular modelling studies. Both the NMR spectroscopic and computational analyses indicate that the investigated ligands bind to the minor grooves as antiparallel side-by-side dimers in a head-to-tail fashion. Moreover, comparisons with results from biochemical studies offered valuable insights into the mechanism of action, and antitumor activity of MGBs in relation to their structures, essential pre-requisites for future optimization of MGBs as therapeutic agents.

Dysregulated cholinergic signaling inhibits oligodendrocyte maturation following demyelination.

Dysregulation of oligodendrocyte progenitor cell (OPC) recruitment and oligodendrocyte differentiation contribute to failure of remyelination in human demyelinating diseases such as multiple sclerosis (MS). Deletion of muscarinic receptor enhances OPC differentiation and remyelination. However, the role of ligand-dependent signaling versus constitutive receptor activation is unknown. We hypothesized that dysregulated acetylcholine (ACh) release upon demyelination contributes to ligand mediated activation hindering myelin repair. Following chronic cuprizone induced demyelination (male and female mice), we observed a 2.5-fold increase in ACh concentration. This increase in ACh concentration could be attributed to increased ACh synthesis or decreased acetylcholinesterase (AChE) / butyrylcholinesterase (BChE) mediated degradation. Using ChAT reporter mice, we identified increased ChAT-GFP expression following both lysolecithin and cuprizone demyelination. ChAT-GFP expression was upregulated in a subset of injured and uninjured axons following intraspinal lysolecithin induced demyelination. In cuprizone demyelinated corpus callosum, ChAT-GFP was observed in Gfap+ astrocytes and axons indicating the potential for neuronal and astrocytic ACh release. BChE expression was significantly decreased in the corpus callosum following cuprizone demyelination. This decrease was due to the loss of myelinating oligodendrocytes which were the primary source of BChE. To determine the role of ligand mediated muscarinic signaling following lysolecithin injection, we administered neostigmine, a cholinesterase inhibitor, to artificially raise ACh. We identified a dose-dependent decrease in mature oligodendrocyte density with no effect on OPC recruitment. Together, these results support a functional role of ligand mediated activation of muscarinic receptors following demyelination and suggest that dysregulation of ACh homeostasis directly contributes to failure of remyelination in MS.Significance Statement Demyelinating diseases like Multiple Sclerosis are characterized by failure of remyelination. Oligodendrocyte progenitor cell (OPC) recruitment and differentiation are crucial aspects for remyelination to occur. Here we show that increased acetylcholine (ACh) contributes to activation of muscarinic receptors that inhibit OPC differentiation. Increased choline acetyltransferase synthesis following demyelination was observed in axons and astrocytes suggestive of a potential for acetylcholine synthesis and release. The increase in ACh levels following demyelination was largely due to reduction of oligodendrocyte derived butyrylcholinesterase that modulates ACh concentration. Development of cell specific esterase stimulator to restore ACh levels may serve as an approach towards inhibiting ongoing demyelination and neurodegeneration.

Complex Elbow Fracture-Dislocations: An Algorithmic Approach to Treatment.

Elbow stability arises from a combination of bony congruity, static ligamentous and capsular restraints, and dynamic muscular activation. Elbow trauma can disrupt these static and dynamic stabilizers leading to predictable patterns of instability; these patterns are dependent on the mechanism of injury and a progressive failure of anatomic structures. An algorithmic approach to the diagnosis and treatment of complex elbow fracture-dislocation injuries can improve the diagnostic assessment and reconstruction of the bony and ligamentous restraints to restore a stable and functional elbow. Achieving optimal outcomes requires a comprehensive understanding of pertinent local and regional anatomy, the altered mechanics associated with elbow injury, versatility in surgical approaches and fixation methods, and a strategic rehabilitation plan.

Added Transverse Screw in Tripod Construct Increases Stiffness in Mason III Radial Head Fractures: A Finite Element Analysis.

BACKGROUND: The tripod screw configuration has been shown to offer similar stiffness characteristics to a laterally placed plate. However, concern has been raised that the construct may be prone to failure in scenarios where the screw intersects at the fracture line. We performed a finite element analysis to assess potentially ideal and unideal screw placements in the tripod construct among Mason III radial head fractures. METHODS: A 3-dimensional proximal radius model was developed using a computed tomography scan of an adult male radius. The fracture site was simulated with a uniform gap in transverse and sagittal planes creating a Mason Type III fracture pattern comprising 3 fragments. Three configurations were modelled with varying screw intersection points in relation to the radial neck fracture line. A fourth configuration comprising an added transverse interfragmentary screw was also modelled. Loading scenarios included axial and shear forces to simulate physiological conditions. Von mises stress and displacement were used as outcomes for analysis. RESULTS: Some variation can be seen among the tripod configurations, with a marginal tendency for reduced implant stress and greater stiffness when screw intersection is further from the neck fracture region. The construct with an added transverse interfragmentary screws demonstrated greater stiffness (2269N/mm) than an equivalent tripod construct comprising three screws (612N/mm). CONCLUSION: The results from this study demonstrate biomechanical similarity between tripod screw constructs including where screws intersect at the radial neck fracture line. An added fourth screw, positioned transversely across fragments increased construct stiffness in our model.

Intravenous iron for heart failure, iron deficiency definitions, and clinical response: the IRONMAN trial.

BACKGROUND AND AIMS: What is the relationship between blood tests for iron deficiency, including anaemia, and the response to intravenous iron in patients with heart failure? METHODS: In the IRONMAN trial, 1137 patients with heart failure, ejection fraction ≤ 45%, and either serum ferritin < 100 µg/L or transferrin saturation (TSAT) < 20% were randomized to intravenous ferric derisomaltose (FDI) or usual care. Relationships were investigated between baseline anaemia severity, ferritin and TSAT, to changes in haemoglobin from baseline to 4 months, Minnesota Living with Heart Failure (MLwHF) score and 6-minute walk distance achieved at 4 months, and clinical events, including heart failure hospitalization (recurrent) or cardiovascular death. RESULTS: The rise in haemoglobin after administering FDI, adjusted for usual care, was greater for lower baseline TSAT (Pinteraction < .0001) and ferritin (Pinteraction = .028) and more severe anaemia (Pinteraction = .014). MLwHF scores at 4 months were somewhat lower (better) with FDI for more anaemic patients (overall Pinteraction = .14; physical Pinteraction = .085; emotional Pinteraction = .043) but were not related to baseline TSAT or ferritin. Blood tests did not predict difference in achieved walking distance for those randomized to FDI compared to control. The absence of anaemia or a TSAT ≥ 20% was associated with lower event rates and little evidence of benefit from FDI. More severe anaemia or TSAT < 20%, especially when ferritin was ≥100 µg/L, was associated with higher event rates and greater absolute reductions in events with FDI, albeit not statistically significant. CONCLUSIONS: This hypothesis-generating analysis suggests that anaemia or TSAT < 20% with ferritin > 100 µg/L might identify patients with heart failure who obtain greater benefit from intravenous iron. This interpretation requires confirmation.

Combining multiple investigative approaches to unravel functional responses to global change in the understorey of temperate forests.

Plant communities are being exposed to changing environmental conditions all around the globe, leading to alterations in plant diversity, community composition, and ecosystem functioning. For herbaceous understorey communities in temperate forests, responses to global change are postulated to be complex, due to the presence of a tree layer that modulates understorey responses to external pressures such as climate change and changes in atmospheric nitrogen deposition rates. Multiple investigative approaches have been put forward as tools to detect, quantify and predict understorey responses to these global-change drivers, including, among others, distributed resurvey studies and manipulative experiments. These investigative approaches are generally designed and reported upon in isolation, while integration across investigative approaches is rarely considered. In this study, we integrate three investigative approaches (two complementary resurvey approaches and one experimental approach) to investigate how climate warming and changes in nitrogen deposition affect the functional composition of the understorey and how functional responses in the understorey are modulated by canopy disturbance, that is, changes in overstorey canopy openness over time. Our resurvey data reveal that most changes in understorey functional characteristics represent responses to changes in canopy openness with shifts in macroclimate temperature and aerial nitrogen deposition playing secondary roles. Contrary to expectations, we found little evidence that these drivers interact. In addition, experimental findings deviated from the observational findings, suggesting that the forces driving understorey change at the regional scale differ from those driving change at the forest floor (i.e., the experimental treatments). Our study demonstrates that different approaches need to be integrated to acquire a full picture of how understorey communities respond to global change.

Impact of vasodilators on diuretic response in patients with congestive heart failure: A mechanistic trial of cimlanod (BMS-986231).

AIM: To investigate the effects of Cimlanod, a nitroxyl donor with vasodilator properties, on water and salt excretion after an administration of an intravenos bolus of furosemide. METHODS AND RESULTS: In this randomized, double-blind, mechanistic, crossover trial, 21 patients with left ventricular ejection fraction <45%, increased plasma concentrations of N-terminal pro-B-type natriuretic peptide (NT-proBNP) and receiving loop diuretics were given, on separate study days, either an 8 h intravenous (IV) infusion of cimlanod (12 µg/kg/min) or placebo. Furosemide was given as a 40 mg IV bolus four hours after the start of infusion. The primary endpoint was urine volume in the 4 h after the bolus of furosemide during infusion of cimlanod compared with placebo. Median NT-proBNP at baseline was 1487 (interquartile range: 847-2665) ng/L. Infusion of cimlanod increased cardiac output and reduced blood pressure without affecting cardiac power index consistent with its vasodilator effects. Urine volume in the 4 h post-furosemide was lower with cimlanod (1032 ± 393 ml) versus placebo (1481 ± 560 ml) (p = 0.002), as were total sodium excretion (p = 0.004), fractional sodium excretion (p = 0.016), systolic blood pressure (p < 0.001), estimated glomerular filtration rate (p = 0.012), and haemoglobin (p = 0.010), an index of plasma volume expansion. CONCLUSIONS: For patients with heart failure and congestion, vasodilatation with agents such as cimlanod reduces the response to diuretic agents, which may offset any benefit from acute reductions in cardiac preload and afterload.

MptpB Inhibitor Improves the Action of Antibiotics against Mycobacterium tuberculosis and Nontuberculous Mycobacterium avium Infections.

Treatment of Mycobacterium tuberculosis and Mycobacterium avium infections requires multiple drugs for long time periods. Mycobacterium protein-tyrosine-phosphatase B (MptpB) is a key M. tuberculosis virulence factor that subverts host antimicrobial activity to promote intracellular survival. Inhibition of MptpB reduces the infection burden in vivo and offers new opportunities to improve current treatments. Here, we demonstrate that M. avium produces an MptpB orthologue and that the MptpB inhibitor C13 reduces the M. avium infection burden in macrophages. Combining C13 with the antibiotics rifampicin or bedaquiline showed an additive effect, reducing intracellular infection of both M. tuberculosis and M. avium by 50%, compared to monotreatment with antibiotics alone. This additive effect was not observed with pretomanid. Combining C13 with the minor groove-binding compounds S-MGB-362 and S-MGB-363 also reduced the M. tuberculosis intracellular burden. Similar additive effects of C13 and antibiotics were confirmed in vivo using Galleria mellonella infections. We demonstrate that the reduced mycobacterial burden in macrophages observed with C13 treatments is due to the increased trafficking to lysosomes.

p38γ MAPK delays myelination and remyelination and is abundant in multiple sclerosis lesions.

Multiple sclerosis is a chronic inflammatory disease in which disability results from the disruption of myelin and axons. During the initial stages of the disease, injured myelin is replaced by mature myelinating oligodendrocytes that differentiate from oligodendrocyte precursor cells. However, myelin repair fails in secondary and chronic progressive stages of the disease and with ageing, as the environment becomes progressively more hostile. This may be attributable to inhibitory molecules in the multiple sclerosis environment including activation of the p38MAPK family of kinases. We explored oligodendrocyte precursor cell differentiation and myelin repair using animals with conditional ablation of p38MAPKγ from oligodendrocyte precursors. We found that p38γMAPK ablation accelerated oligodendrocyte precursor cell differentiation and myelination. This resulted in an increase in both the total number of oligodendrocytes and the migration of progenitors ex vivo and faster remyelination in the cuprizone model of demyelination/remyelination. Consistent with its role as an inhibitor of myelination, p38γMAPK was significantly downregulated as oligodendrocyte precursor cells matured into oligodendrocytes. Notably, p38γMAPK was enriched in multiple sclerosis lesions from patients. Oligodendrocyte progenitors expressed high levels of p38γMAPK in areas of failed remyelination but did not express detectable levels of p38γMAPK in areas where remyelination was apparent. Our data suggest that p38γ could be targeted to improve myelin repair in multiple sclerosis.

Outcomes following perioperative red blood cell transfusion in patients undergoing elective major abdominal surgery: a systematic review and meta-analysis.

BACKGROUND: Perioperative red blood cell transfusion is a double-edged sword for surgical patients. While transfusion of red cells can increase oxygen delivery by increasing haemoglobin levels, its impact on short- and long-term postoperative outcomes, particularly in patients undergoing elective major abdominal surgery, is unclear. METHODS: We conducted a systematic review and meta-analysis on the effect of perioperative blood transfusions on postoperative outcomes in elective major abdominal surgery. PubMed, Cochrane, and Scopus databases were searched for studies with data collected between January 1, 2000 and June 6, 2020. The primary outcome was short-term mortality, including all-cause 30-day or in-hospital mortality. Secondary outcomes included long-term all-cause mortality, any morbidity, infectious complications, overall survival, and recurrence-free survival. No randomised controlled trials were found. Thirty-nine observational studies were identified, of which 37 were included in the meta-analysis. RESULTS: Perioperative blood transfusion was associated with short-term all-cause mortality (odds ratio [OR] 2.72, 95% confidence interval [CI] 1.89-3.91, P<0.001), long-term all-cause mortality (hazard ratio 1.35, 95% CI 1.09-1.67, P=0.007), any morbidity (OR 2.18, 95% CI 1.81-2.64, P<0.001), and infectious complications (OR 1.90, 95% CI 1.60-2.26, P<0.001). Perioperative blood transfusion remained associated with short-term mortality in the sensitivity analysis after excluding studies that did not control for preoperative anaemia (OR 2.27, 95% CI 1.59-3.24, P<0.001). CONCLUSIONS: Perioperative blood transfusion in patients undergoing elective major abdominal surgery is associated with poorer short- and long-term postoperative outcomes. This highlights the need to implement patient blood management strategies to manage and preserve the patient's own blood and reduce the need for red blood cell transfusion. TRIAL REGISTRATION: PROSPERO (CRD42021254360).

Histologic Comparison of the Fascicular Area of Processed Nerve Allograft Versus Cabled Sural Nerve Autograft.

Background: The use of multiple cables of sural nerve autograft is common for peripheral nerve reconstruction when injured nerve caliber exceeds the nerve graft caliber. Although the optimal matching of neural to nonneural elements and its association with functional outcomes are unknown, it is reasonable to consider maximizing the neural tissue structure available for nerve regeneration. No prior studies have compared directly the cross-sectional fascicular area between cabled nerve autografts and size-selected nerve allografts. This study evaluated the cross-sectional fascicular area between native nerve stumps and two reconstructive nerve grafting methods: cabled sural nerve autograft (CSNA) and processed nerve allograft (PNA). Methods: CSNA from matched cadaveric specimens and PNA were used to reconstruct nerve defects in the median and ulnar nerves of six pairs of cadaveric specimens. Nerve reconstructions were done by fellowship-trained hand surgeons. The total nerve area, fascicular area, and nonfascicular area were measured histologically. Results: The CSNA grafts had significantly less fascicular area than PNA and caliber-matched native nerve. The PNA grafts had a significantly higher percent fascicular area compared with the intercalary CNSA graft. Conclusions: Fascicular area was significantly greater in PNA versus CSNA. The PNA consistently demonstrated a match in fascicular area closer to the native nerve stumps than CSNA, where CSNA had significantly smaller fascicular area compared with native nerve stumps.

Influence of serum transferrin concentration on diagnostic criteria for iron deficiency in chronic heart failure.

AIMS: Transferrin saturation (TSAT), a marker of iron deficiency, reflects both serum concentrations of iron (SIC) and transferrin (STC). TSAT is susceptible to changes in each of these biomarkers. Little is known about determinants of STC and its influence on TSAT and mortality in patients with heart failure. Accordingly, we studied the relationship of STC to clinical characteristics, to markers of iron deficiency and inflammation and to mortality in chronic heart failure (CHF). METHODS AND RESULTS: Prospective cohort of patients with CHF attending a clinic serving a large local population. A total of 4422 patients were included (median age 75 (68-82) years; 40% women; 32% with left ventricular ejection fraction ≤40%). STC ≤ 2.3 g/L (lowest quartile) was associated with older age, lower SIC and haemoglobin and higher high-sensitivity C-reactive protein, ferritin and N-terminal pro-brain natriuretic peptide compared with those with STC > 2.3 g/L. In the lowest STC quartile, 624 (52%) patients had SIC ≤13 µmol/L, of whom 38% had TSAT ≥20%. For patients in the highest STC quartile, TSAT was <20% when SIC was >13 µmol/L in 185 (17%) patients. STC correlated inversely with ferritin (r = -0.52) and high-sensitivity C-reactive protein (r = -0.17) and directly with albumin (r = 0.29); all P < 0.001. In models adjusted for age, N-terminal pro-brain natriuretic peptide and haemoglobin, both higher SIC (hazard ratio 0.87 [95% CI: 0.81-0.95]) and STC (hazard ratio 0.82 [95% CI: 0.73-0.91]) were associated with lower mortality. SIC was more strongly associated with both anaemia and mortality than either STC or TSAT. CONCLUSIONS: Many patients with CHF and a low STC have low SIC even when TSAT is >20% and serum ferritin >100 µg/L; such patients have a high prevalence of anaemia and a poor prognosis and might have iron deficiency but are currently excluded from clinical trials of iron repletion.

Defining Heart Failure Based on Imaging the Heart and Beyond.

Water and salt retention, in other words congestion, are fundamental to the pathophysiology of heart failure and are important therapeutic targets. Echocardiography is the key tool with which to assess cardiac structure and function in the initial diagnostic workup of patients with suspected heart failure and is essential for guiding treatment and stratifying risk. Ultrasound can also be used to identify and quantify congestion in the great veins, kidneys and lungs. More advanced imaging methods might further clarify the aetiology of heart failure and its consequences for the heart and periphery, thereby improving the efficiency and quality of care tailored with greater precision to individual patient need.

Capacity Assessment Tool to Promote Capacity Building in Global Orthopaedic Surgical Outreach.

BACKGROUND: A growing number of nongovernmental organizations from high-income countries aim to provide surgical outreach for patients in low- and middle-income countries in a manner that builds capacity. There remains, however, a paucity of measurable steps to benchmark and evaluate capacity-building efforts. Based on a framework for capacity building, the present study aimed to develop a Capacity Assessment Tool for orthopaedic surgery (CAT-os) that could be utilized to evaluate and promote capacity building. METHODS: To develop the CAT-os tool, we utilized methodological triangulation-an approach that incorporates multiple different types of data. We utilized (1) the results of a systematic review of capacity-building best practices in surgical outreach, (2) the HEALTHQUAL National Organizational Assessment Tool, and (3) 20 semistructured interviews to develop a draft of the CAT-os. We subsequently iteratively used a modified nominal group technique with a consortium of 8 globally experienced surgeons to build consensus, which was followed by validation through member-checking. RESULTS: The CAT-os was developed and validated as a formal instrument with actionable steps in each of 7 domains of capacity building. Each domain includes items that are scaled for scoring. For example, in the domain of partnership, items range from no formalized plans for sustainable, bidirectional relationships (no capacity) to local surgeons and other health-care workers independently participating in annual meetings of surgical professional societies and independently creating partnership with third party organizations (optimal capacity). CONCLUSIONS: The CAT-os details steps to assess capacity of a local facility, guide capacity-improvement efforts during surgical outreach, and measure the impact of capacity-building efforts. Capacity building is a frequently cited and commendable approach to surgical outreach, and this tool provides objective measurement to aid in improving the capacity in low and middle-income countries through surgical outreach.

Proximal Hamate Reconstruction of Proximal Pole Scaphoid Nonunion: A Case Series and Analysis of Clinical Outcomes.

BACKGROUND: Small proximal pole scaphoid nonunions present a clinical challenge influenced by fragment size, vascular compromise, deforming forces exerted through the scapholunate interosseous ligament (SLIL), and potential articular fragmentation. Osteochondral autograft options for proximal pole reconstruction include the medial femoral trochlea, costochondral rib, or proximal hamate. This study reports the clinical outcomes of patients treated with proximal hamate osteochondral autograft reconstruction. METHODS: A retrospective review identified patients treated with this surgery from 2 institutions with a minimum 6-month follow-up. Clinical outcomes included the Visual Analog Dcale pain score, 12-item Short-Form survey, abbreviated Disabilities of the Arm, Shoulder, and Hand (QuickDASH) score, wrist and forearm range of motion (ROM), radiographic assessment, and complications. We reviewed and compared these outcomes with those of the current published literature. RESULTS: Four patients (mean age: 24 years, 75% men) with a 12.8-month average follow-up (range: 6-20 months) were included. Radiographic union was identified in all cases by 12 weeks (range, 10-12). The average wrist ROM was 67.5% flexion/extension and 100% pronation/supination compared with the contralateral side at the final follow-up. The mean QuickDASH score was 17.6 (SD, 13). No complications were identified. CONCLUSIONS: Proximal pole scaphoid nonunion reconstruction using autologous proximal hamate osteochondral graft demonstrated encouraging clinical and radiographic outcomes. Proximal hamate harvest involves minimal donor site morbidity without a distant operative site, uses an osteochondral graft with similar morphology to the proximal scaphoid, requires no microsurgical technique, and permits reconstruction of the SLIL using the volar capitohamate ligament.

Assessment of haemoglobin and serum markers of iron deficiency in people with cardiovascular disease.

BACKGROUND: The prevalence of anaemia and iron deficiency and their prognostic association with cardiovascular disease have rarely been explored at population level. METHODS: National Health Service records of the Greater Glasgow region for patients aged ≥50 years with a broad range of cardiovascular diagnoses were obtained. During 2013/14, prevalent disease was identified and results of investigations collated. Anaemia was defined as haemoglobin <13 g/dL for men or <12 g/dL for women. Incident heart failure, cancer and death between 2015 and 2018 were identified. RESULTS: The 2013/14 dataset comprised 197 152 patients, including 14 335 (7%) with heart failure. Most (78%) patients had haemoglobin measured, especially those with heart failure (90%). Of those tested, anaemia was common both in patients without (29%) and with heart failure (prevalent cases in 2013/14: 46%; incident cases during 2013/14: 57%). Ferritin was usually measured only when haemoglobin was markedly depressed; transferrin saturation (TSAT) even less often. Incidence rates for heart failure and cancer during 2015-18 were inversely related to nadir haemoglobin in 2013/14. A haemoglobin of 13-15 g/dL for women and 14-16 g/dL for men was associated with the lowest mortality. Low ferritin was associated with a better prognosis and low TSAT with a worse prognosis. CONCLUSION: In patients with a broad range of cardiovascular disorders, haemoglobin is often measured but, unless anaemia is severe, markers of iron deficiency are usually not. Low haemoglobin and TSAT, but not low ferritin, are associated with a worse prognosis. The nadir of risk occurs at haemoglobin 1-3 g/dL above the WHO definition of anaemia.