On the role of dysferlin in striated muscle: membrane repair, t-tubules and Ca2+ handling.

Dysferlin is a 237 kDa membrane-associated protein characterised by multiple C2 domains with a diverse role in skeletal and cardiac muscle physiology. Mutations in DYSF are known to cause various types of human muscular dystrophies, known collectively as dysferlinopathies, with some patients developing cardiomyopathy. A myriad of in vitro membrane repair studies suggest that dysferlin plays an integral role in the membrane repair complex in skeletal muscle. In comparison, less is known about dysferlin in the heart, but mounting evidence suggests that dysferlin's role is similar in both muscle types. Recent findings have shown that dysferlin regulates Ca2+ handling in striated muscle via multiple mechanisms and that this becomes more important in conditions of stress. Maintenance of the transverse (t)-tubule network and the tight coordination of excitation-contraction coupling are essential for muscle contractility. Dysferlin regulates the maintenance and repair of t-tubules, and it is suspected that dysferlin regulates t-tubules and sarcolemmal repair through a similar mechanism. This review focuses on the emerging complexity of dysferlin's activity in striated muscle. Such insights will progress our understanding of the proteins and pathways that regulate basic heart and skeletal muscle function and help guide research into striated muscle pathology, especially that which arises due to dysferlin dysfunction.

Phase II trial of domatinostat (4SC-202) in combination with avelumab in patients with previously treated advanced mismatch repair proficient oesophagogastric and colorectal adenocarcinoma: EMERGE.

BACKGROUND: Most oesophagogastric adenocarcinomas (OGAs) and colorectal cancers (CRCs) are mismatch repair proficient (MMRp), responding poorly to immune checkpoint inhibition. We evaluated the safety and efficacy of domatinostat (histone deacetylase inhibitor) plus avelumab (anti-PD-L1 antibody) in patients with previously treated inoperable, advanced/metastatic MMRp OGA and CRC. PATIENTS AND METHODS: Eligible patients were evaluated in a multicentre, open-label dose escalation/dose expansion phase II trial. In the escalation phase, patients received escalating doses of domatinostat [100 mg once daily (OD), 200 mg OD, 200 mg twice daily (BD)] orally for 14 days followed by continuous dosing plus avelumab 10 mg/kg administered intravenously 2-weekly (2qw) to determine the recommended phase II dose (RP2D). The trial expansion phase evaluated the best objective response rate (ORR) during 6 months by RECIST version 1.1 using a Simon two-stage optimal design with 2/9 and 1/10 responses required to proceed to stage 2 in the OGA and CRC cohorts, respectively. RESULTS: Patients (n = 40) were registered between February 2019 and October 2021. Patients in the dose escalation phase (n = 12) were evaluated to confirm the RP2D of domatinostat 200 mg BD plus avelumab 10 mg/kg. No dose-limiting toxicities were observed. Twenty-one patients were treated at the RP2D, 19 (9 OGA and 10 CRC) were assessable for the best ORR; 2 patients with CRC did not receive combination treatment and were not assessable for the primary endpoint analysis. Six patients were evaluated in the dose escalation and expansion phases. In the OGA cohort, the best ORR was 22.2% (95% one-sided confidence interval lower bound 4.1) and the median duration of disease control was 11.3 months (range 9.9-12.7 months). No responses were observed in the CRC cohort. No treatment-related grade 3-4 adverse events were reported at the RP2D. CONCLUSIONS: Responses in the OGA cohort met the criteria to expand to stage 2 of recruitment with an acceptable safety profile. There was insufficient signal in the CRC cohort to progress to stage 2. TRIAL REGISTRATION: NCT03812796 (registered 23rd January 2019).

Practitioner review: health anxiety in children and young people in the context of the COVID-19 pandemic.

Health-related fear is a normal and common response in the face of the global pandemic of COVID-19. Children and young people are frequently being exposed to messages about the threat to health, including from the media and authorities. Whilst for most, their anxiety will be proportionate to the threat, for some, existing pre-occupation with physical symptoms and illness will become more problematic. There is a growing body of evidence that health anxiety may occur in childhood, however much of the literature is taken from research using adult samples. This practitioner review aims to give an overview of the assessment and treatment of health-related worries in children and young people in the context of the COVID-19 pandemic. This review is based on the limited existing evidence in this population and the more substantial evidence base for treating health anxiety in adults. We consider the adaptations needed to ensure such interventions are developmentally appropriate.

An economic analysis of ransomware and its welfare consequences.

We present in this work an economic analysis of ransomware, a relatively new form of cyber-enabled extortion. We look at how the illegal gains of the criminals will depend on the strategies they use, examining uniform pricing and price discrimination. We also explore the welfare costs to society of such strategies. In addition, we present the results of a pilot survey which demonstrate proof of concept in evaluating the costs of ransomware attacks. We discuss at each stage whether the different strategies we analyse have been encountered already in existing malware, and the likelihood of them being implemented in the future. We hope this work will provide some useful insights for predicting how ransomware may evolve in the future.

Effectiveness of self-management interventions in young adults with type 1 and 2 diabetes: a systematic review and meta-analysis.

AIM: Diabetes in young adulthood has been associated with poor outcomes. Self-management is fundamental to good diabetes care, and self-management interventions have been found to improve outcomes in older adults. We performed a systematic review and meta-analysis to assess the effectiveness of self-management interventions in young adults (aged 15-39 years) with type 1 or type 2 diabetes. METHODS: We searched five databases and two clinical trial registries from 2003 to February 2019, without language restrictions. We included randomized controlled trials (RCTs) comparing the effectiveness of self-management interventions with usual care or enhanced usual care in young adults. Outcomes of interest included clinical outcomes, psychological health, self-care behaviours, diabetes knowledge and self-efficacy. Pairwise meta-analysis was conducted using a random effects model and quality of evidence was assessed using the Grading of Recommendations, Assessment, Development and Evaluations (GRADE) criteria. We followed Cochrane gold standard systematic review methodology and reported this systematic review according to PRISMA guidelines. The protocol was registered with PROSEPRO (CRD42018110868). RESULTS: In total, 13 studies (1002 participants) were included. Meta-analysis showed no difference between self-management interventions and controls in post-intervention HbA1c levels, BMI, depression, diabetes-related distress, overall self-care, diabetes knowledge and self-efficacy. Quality of evidence ranged from very low to moderate due to study limitations, inconsistency and imprecision. CONCLUSIONS: Current self-management interventions did not improve outcomes in young adults with diabetes. Our findings, which contrast with those from systematic reviews in older adults, highlight the need for the development of more effective interventions for young adults with diabetes.

A Multistate Investigation of Antibiotic-Resistant Salmonella enterica Serotype I 4,[5],12:i:- Infections as Part of an International Outbreak Associated with Frozen Feeder Rodents.

While most human Salmonella infections result from exposure to contaminated foods, an estimated 11% of all Salmonella infections are attributed to animal exposures, including both direct animal handling and indirect exposures such as cleaning cages and handling contaminated pet food. This report describes the epidemiologic, environmental and laboratory investigations conducted in the United States as part of the response to an international outbreak of tetracycline-resistant Salmonella enterica serotype I 4,[5],12:i:- infections with over 500 illnesses occurring from 2008 to 2010. This investigation found that illness due to the outbreak strain was significantly associated with exposure to pet reptiles and frozen feeder rodents used as food for pet reptiles. Salmonella isolates indistinguishable from the outbreak strain were isolated from a frozen feeder mice-fed reptile owned by a case patient, as well as from frozen feeder mice and environmental samples collected from a rodent producing facility (Company A). An international voluntary recall of all Company A produced frozen feeder animals sold between May 2009 and July 2010 occurred. Only 13% of cases in our investigation were aware of the association between Salmonella infection and mice or rats. Consumers, the pet industry, healthcare providers and veterinarians need to be aware of the potential health risk posed by feeder rodents, whether live or frozen. Frozen feeder rodent producers, suppliers and distributors should follow the animal food labelling requirements as described in 21 CFR §501.5, and all packages of frozen feeder rodents should include safe handling instructions. Persons should wash their hands thoroughly with soap and water after handling live or frozen feeder rodents, as well as reptiles or anything in the area where the animals live. Continued opportunities exist for public health officials, the pet industry, veterinarians and consumers to work together to prevent salmonellosis associated with pet food, pets and other animals.

Impact of routine bedside infectious disease consultation on clinical management and outcome of Staphylococcus aureus bacteraemia in adults.

Staphylococcus aureus bacteraemia (SAB) is a common, serious infection that is associated with high rates of morbidity and mortality. Evidence suggests that infectious disease consultation (IDC) improves clinical management in patients with SAB. We examined whether the introduction of a routine bedside IDC service for adults with SAB improved clinical management and outcomes compared to telephone consultation. We conducted an observational cohort study of 571 adults with SAB at a teaching hospital in the United Kingdom between July 2006 and December 2012. A telephone consultation was provided on the day of positive blood culture in all cases, but an additional bedside IDC was provided after November 2009 (routine IDC group). Compared to patients in the pre-IDC group, those in the routine IDC group were more likely to have a removable focus of infection identified, echocardiography performed and follow-up blood cultures performed. They also received longer courses of antimicrobial therapy, were more likely to receive combination antimicrobial therapy and were more likely to have SAB recorded in the hospital discharge summary. There was a trend towards lower mortality at 30 days in the routine IDC group compared to the pre-IDC group (12% vs. 22%, p 0.07). Our findings suggest that routine bedside IDC should become the standard of care for adults with SAB.

Macitentan treatment retards the progression of established pulmonary arterial hypertension in an animal model.

BACKGROUND: Macitentan is a new endothelin receptor antagonist that is used to treat pulmonary arterial hypertension in humans. Treatment of established pulmonary hypertension with macitentan was studied using the monocrotaline model of pulmonary hypertension. METHODS: Three groups of rats were created (n=12): control (CON: macitentan only), monocrotaline (MCT: monocrotaline only) and macitentan (MACI: macitentan and monocrotaline). Monocrotaline (60 mg/kg) was injected in the MCT and MACI groups on day 0; volume matched saline was injected in the CON groups. Macitentan therapy (30 mg/kg/day) was commenced on day 11 in the CON and MACI groups. Serial echocardiography and ECGs were performed. The rats were sacrificed if they showed clinical deterioration. RESULTS: The MCT and MACI rats showed signs of pulmonary hypertension by day 7 (maximum pulmonary velocity, CON 1.15 ± 0.15m/s vs MCT 1.04 ± 0.10 m/s vs MACI 0.99 ± 0.18 m/s; p<0.05). Both the MCT and MACI groups developed pulmonary hypertension, but this was less severe in the MACI group (day 21 pulmonary artery acceleration time, MCT 17.55 ± 1.56 ms vs MACI 22.55 ± 1.00 ms; pulmonary artery deceleration, MCT 34.72 ± 3.72 m/s(2) vs MACI 17.30 ± 1.89 m/s(2); p<0.05). Right ventricular hypertrophy and QT interval increases were more pronounced in MCT than MACI (right ventricle wall thickness, MCT 0.13 ± 0.1cm vs MACI 0.10 ± 0.1cm; QT interval, MCT 85 ± 13 ms vs MACI 71 ± 14 ms; p<0.05). Survival benefit was not seen in the MACI group (p=0.50). CONCLUSIONS: Macitentan treatment improves haemodynamic parameters in established pulmonary hypertension. Further research is required to see if earlier introduction of macitentan has greater effects.

Prevalence and characterization of human mecC methicillin-resistant Staphylococcus aureus isolates in England.

OBJECTIVES: There are limited data available on the epidemiology and prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in the human population that encode the recently described mecA homologue, mecC. To address this knowledge gap we undertook a prospective prevalence study in England to determine the prevalence of mecC among MRSA isolates. PATIENTS AND METHODS: Three hundred and thirty-five sequential MRSA isolates from individual patients were collected from each of six clinical microbiology laboratories in England during 2011-12. These were tested by PCR or genome sequencing to differentiate those encoding mecA and mecC. mecC-positive isolates were further characterized by multilocus sequence typing, spa typing, antimicrobial susceptibility profile and detection of PBP2a using commercially available kits. RESULTS: Nine out of the 2010 MRSA isolates tested were mecC positive, indicating a prevalence among MRSA in England of 0.45% (95% CI 0.24%-0.85%). The remainder were mecA positive. Eight out of these nine mecC MRSA isolates belonged to clonal complex 130, the other being sequence type 425. Resistance to non-ß-lactam antibiotics was rare among these mecC MRSA isolates and all were phenotypically identified as MRSA using oxacillin and cefoxitin according to BSAC disc diffusion methodology. However, all nine mecC isolates gave a negative result using three different commercial PBP2a detection assays. CONCLUSIONS: mecC MRSA are currently rare among MRSA isolated from humans in England and this study provides an important baseline prevalence rate to monitor future changes, which may be important given the increasing prevalence of mecC MRSA reported in Denmark.

Recurrent epidemic cholera with high mortality in Cameroon: persistent challenges 40 years into the seventh pandemic.

Cameroon has experienced recurrent cholera epidemics with high mortality rates. In September 2009, epidemic cholera was detected in the Far North region of Cameroon and the reported case-fatality rate was 12%. We conducted village-, healthcare facility- and community-level surveys to investigate reasons for excess cholera mortality. Results of this investigation suggest that cholera patients who died were less likely to seek care, receive rehydration therapy and antibiotics at a healthcare facility, and tended to live further from healthcare facilities. Furthermore, use of oral rehydration salts at home was very low in both decedents and survivors. Despite the many challenges inherent to delivering care in Cameroon, practical measures could be taken to reduce cholera mortality in this region, including the timely provision of treatment supplies, training of healthcare workers, establishment of rehydration centres, and promotion of household water treatment and enhanced handwashing with soap.

The plasma membrane calcium ATPase modulates calcium homeostasis, intracellular signaling events and function in platelets.

BACKGROUND: The plasma membrane calcium ATPase (PMCA) regulates localized signaling events in a variety of cell types, although its functional role in platelets remains undefined. OBJECTIVES: To investigate the role of PMCA in determining platelet intracellular calcium concentration ([Ca²(+) ](i) ) at rest and following agonist stimulation, and to define the corresponding effects upon different stages of platelet activation. METHODS: [Ca²(+) ](i) was continuously measured in Fura-2-loaded platelets and in vitro and in vivo functional analyses performed in the presence of the PMCA inhibitor carboxyeosin (CE). RESULTS: Concentrations of CE that selectively inhibited Ca²(+) extrusion through PMCA were established in human platelets. [Ca²(+) ](i) was elevated by CE in resting platelets, although collagen-stimulated Ca²(+) release was reduced. Impaired Ca²(+) mobilization upon agonist stimulation was accompanied by reduced dense granule secretion and impaired platelet aggregation. Platelet aggregation responses were also reduced in PMCA4(-/-) mice and in an in vivo mouse model of platelet thromboembolism. Conversely, inhibition of PMCA promoted the early and later stages of platelet activation, observed as enhanced adhesion to fibrinogen, and accelerated clot retraction. Investigations into the signaling mechanisms underlying CE-mediated inhibition of platelet aggregation implicated cGMP-independent vasodilator-stimulated phosphoprotein phosphorylation. CONCLUSIONS: Disruption of PMCA activity perturbs platelet Ca²(+) homeostasis and function in a time-dependent manner, demonstrating that PMCA differentially regulates Ca²(+) -dependent signaling events, and hence function, throughout the platelet activation process.

Stage I twin-twin transfusion syndrome: rates of progression and regression in relation to outcome.

OBJECTIVES: Twin-twin transfusion syndrome (TTTS) results in high rates of perinatal mortality and neurological morbidity. Fetoscopic laser ablation of placental anastomoses is now established as the treatment of choice for advanced disease. However, there remains controversy about its use in early-stage TTTS, in which laser-related fetal losses need to be balanced against relatively favorable outcomes with more conservative approaches. We investigated rates of progression and regression in Stage I TTTS and determined factors influencing the course of the disease. METHODS: We undertook a retrospective observational study of all TTTS cases referred to our tertiary referral fetal medicine service from 2000 to 2006. In patients presenting with Stage I TTTS, the following variables were evaluated for their ability to predict the course and progression of the disease: gestational age (GA) at presentation, amniotic fluid index, recipient and donor deepest vertical pool, presence of artery-artery anastomoses, small-sized bladder compared to normal donor bladder and fetal size discordance. Study end-points were disease regression or progression, and neonatal survival at 28 days. RESULTS: Among 132 consecutive cases of TTTS, 46 women presented with Stage I disease. In the majority (69.6%), disease remained stable (28.3%) or regressed (41.3%). Of cases that progressed, 79% did so within 2 weeks and 93% progressed to at least Stage III. No factor was significantly linked with progression or regression, although there was a trend towards the absence of an artery-artery anastomosis (P = 0.10) and the presence of a small rather than normal donor bladder (P = 0.10) influencing progression, and later GA at presentation (P = 0.07) influencing regression. At least one infant survived in 83% of cases and there was double survival in 59%. Perinatal outcome was significantly better in cases that regressed (the rates of at least one survivor and double survival being 89% and 89%, respectively) or remained Stage I (77% and 61%, respectively), compared with those cases that progressed (79% and 14%, respectively). Treatment with amnioreduction at first presentation did not influence progression or regression. CONCLUSIONS: This study demonstrates that a high percentage of Stage I TTTS cases regress or remain early stage. Identification of factors predicting progression would facilitate the selection of patients for definitive therapy, while avoiding treatment-related morbidities in mild or transient disease.

The regulatory function of plasma-membrane Ca(2+)-ATPase (PMCA) in the heart.

The PMCA (plasma-membrane Ca(2+)-ATPase) is a ubiquitously expressed calcium-extruding enzymatic pump important in the control of intracellular calcium concentration. Unlike in non-excitable cells, where PMCA is the only system for calcium extrusion, in excitable cells, such as cardiomyocytes, PMCA has been shown to play only a minor role in calcium homoeostasis compared with the NCX (sodium/calcium exchanger), another system of calcium extrusion. However, increasing evidence points to an important role for PMCA in signal transduction; of particular interest in cardiac physiology is the modulation of nNOS (neuronal nitric oxide synthase) by isoform 4b of PMCA. In the present paper, we will discuss recent advances that support a key role for PMCA4 in modulating the nitric oxide signalling pathway in the heart.

Gastrin releasing peptide and gastrin releasing peptide receptor expression in gastrointestinal carcinoid tumours.

AIMS: To establish whether gastrin releasing peptide (GRP) and the GRP receptor (GRPR) are expressed together in gastrointestinal carcinoid tumours. METHODS: Twenty six carcinoid tumours from the stomach, small intestine, appendix, and colorectum were investigated by immunohistochemistry for GRP and GRPR. RESULTS: GRP was detected in nine of 19 tumours and GRPR in 22 of 26. Coexpression of both the ligand and receptor was seen in six of 19 cases. GRPR but not GRP was more strongly expressed in appendix and colonic tumours. CONCLUSIONS: GRP and GRPR are produced by a large number of gastrointestinal carcinoid tumours. An autocrine/paracrine pathway may exist for GRP stimulated cell proliferation in some of these neoplasms, analogous to that seen in small cell anaplastic carcinoma of the lung.

Lack of inducible nitric oxide synthase promotes intestinal tumorigenesis in the Apc(Min/+) mouse.

BACKGROUND & AIMS: The role of the inducible isoform of nitric oxide synthase (Nos2 or iNOS) in intestinal tumorigenesis is unclear. Conflicting data also exist regarding the ability of Nos2 to modulate expression and/or activity of cyclooxygenase 2 (Cox-2), which promotes intestinal tumorigenesis. Therefore, we determined the effect of a null Nos2 genotype on intestinal tumorigenesis and Cox-2 expression/activity in the Apc(Min/+) mouse model of familial adenomatous polyposis. METHODS: Apc(Min/+)Nos2(-/-) mice were generated by successive crosses between C57BL/6-Apc(Min/+) and C57BL/6-Nos2(tm1Lau) mice. Adenoma characteristics of age-matched Apc(Min/+)Nos2(+/+) and Apc(Min/+)Nos2(-/-) mice were compared. The level and cellular localization of Nos2 messenger RNA (mRNA) expression in Apc(Min/+)Nos2(+/+) mouse intestine was determined. Cox-2 expression and activity were measured in both intestinal tissue and bone marrow-derived macrophages in vitro. RESULTS: Apc(Min/+)Nos2(-/-) mice developed significantly more intestinal adenomas than Apc(Min/+)Nos2(+/+) littermates. Epithelial cell Nos2 mRNA expression was decreased in adenomas compared with histologically normal Apc(Min/+)Nos2(+/+) intestine. There was no significant difference in Cox-2 expression or activity in either intestine or bone marrow-derived macrophages from Apc(Min/+)Nos2(+/+) and Apc(Min/+)Nos2(-/-) animals. CONCLUSIONS: Nos2 plays an antineoplastic role in the Apc(Min/+) mouse model of familial adenomatous polyposis. Nos2 does not modulate Cox-2 expression or activity in the Apc(Min/+) mouse.

Characterisation and expression of Sox9 in the Leopard gecko, Eublepharis macularius.

Since the discovery of the sex-determining gene, Sry, a number of genes have been identified which are involved in sex determination and gonadogenesis in mammals. Although Sry is known to be the testis-determining factor in mammals, this is not the case in non-mammalian vertebrates. Sox9 is another gene that has been shown to have a male-specific role in sex determination, but, unlike Sry, Sox9 has been shown to be involved in sex determination in mammals, birds, and reptiles. This is the first gene to be described that has a conserved role in sex determination in species with either chromosomal or environmental sex-determining mechanisms. Many reptiles do not have sex chromosomes but exhibit temperature-dependent sex determination (TSD). Sox9 has been shown to be expressed in both turtle and alligator during gonadogenesis. To determine if Sox9 also has a role in a gecko species with TSD, we studied gonadal expression of Sox9 during embryonic development of the Leopard gecko (Eublepharis macularius). Gecko Sox9 was found to be highly conserved at the nucleotide level when compared to other vertebrate species including human, chick, alligator, and turtle. Sox9 was found to be expressed in embryos incubated at the male-producing temperature (32.5 degrees C) as well as in embryos incubated at the female-producing temperatures (26 and 34 degrees C), Northern blot analysis showed that Sox9 was expressed at both temperatures from morphological stages 31 to 37. mRNA in situ hybridisation on isolated urogenital systems showed expression at both female- and male-producing temperatures up to stage 36. After this stage, no expression was seen in the female gonads but expression remained in the male. These data provide further evidence that Sox9 is an essential component of a testis-determining pathway that is conserved in species with differing sex-determining mechanisms.

Localization of cyclooxygenase-2 in human sporadic colorectal adenomas.

A putative target for the anti-colorectal cancer action of nonsteroidal anti-inflammatory drugs is the inducible isoform of cyclooxygenase (COX), COX-2. COX-2 is expressed within intestinal adenomas in murine polyposis models, but expression has been poorly characterized in human colorectal neoplasms. Therefore, we investigated the localization of the COX-2 protein in human sporadic colorectal adenomas. Immunohistochemistry for COX-2 and CD68 (a tissue macrophage marker) was performed on formalin-fixed, paraffin-embedded (n = 52) and frozen, acetone-fixed (n = 6) sections of human sporadic colorectal adenomas. Forty of 52 (77%) formalin-fixed adenomas expressed immunoreactive COX-2. COX-2 was localized to superficial interstitial macrophages in 39 cases (75%) and to deep interstitial macrophages in 9 cases (17%). COX-2 staining of dysplastic epithelial cells was observed in 15 cases (29%). A logistic regression analysis identified the adenoma site (P = 0.012) and histological type (P = 0.001) as independent predictors of superficial macrophage COX-2 expression. There was no relationship between the number of macrophages within an adenoma and macrophage COX-2 expression. These results indicate that COX-2 is expressed predominantly by interstitial macrophages within human sporadic colorectal adenomas. If COX-2 does indeed play a role in the early stages of colorectal carcinogenesis in man, these data suggest COX-2-mediated paracrine signaling between the macrophages and epithelial cells within adenomas.