Validation of suitable internal control genes for expression studies in aging.

Quantitative data from experiments of gene expression are often normalized through levels of housekeeping genes transcription by assuming that expression of these genes is highly uniform. This practice is being questioned as it becomes increasingly clear that the level of housekeeping genes expression may vary considerably in certain biological samples. To date, the validation of reference genes in aging has received little attention and suitable reference genes have not yet been defined. Our aim was to evaluate the expression stability of frequently used reference genes in human peripheral blood mononuclear cells with respect to aging. Using quantitative RT-PCR, we carried out an extensive evaluation of five housekeeping genes, i.e. 18s rRNA, ACTB, GAPDH, HPRT1 and GUSB, for stability of expression in samples from donors in the age range 35-74 years. The consistency in the expression stability was quantified on the basis of the coefficient of variation and two algorithms termed geNorm and NormFinder. Our results indicated GUSB be the most suitable transcript and 18s the least for accurate normalization in PBMCs. We also demonstrated that aging is a confounding factor with respect to stability of 18s, HPRT1 and ACTB expression, which were particularly prone to variability in aged donors

Prevalence and severity of abscesses and cellulitis, and their associations with other health outcomes, in a community-based study of people who inject drugs in London, UK.

BACKGROUND: Skin and soft tissue infections (SSTI) are a common but preventable cause of morbidity and mortality among people who inject drugs (PWID). They can be severe, and hospitalisations of PWID with SSTI are rising. The most common SSTI presentations are abscesses and cellulitis. METHODS: We used data from Care & Prevent, a cross-sectional community survey of PWID in London. We reported the lifetime prevalence of SSTI, severity of infections, key risk factors, and associated sequelae. Pictorial questions were used to assess SSTI severity. RESULTS: We recruited 455 PWID. SSTI lifetime prevalence was high: 64% reported an abscess and/or cellulitis. Over one-third (37%) reported a severe infection, 137 (47%) reported hospitalisation. SSTIrisk factors were: aged 35+ years, injecting once or more times a day, subcutaneous or intra-muscular injections, and making four or more attempts to achieve an injection. Those who reported having other health conditions were at higher odds of having an abscess or cellulitis, with risk tending to increase with number of reported conditions. Half (46%) employed self-care for their worst SSTI, and 43% waited for ten or more days before seeking medical care or not seeking medical care at all. CONCLUSIONS: Abscess and cellulitis are very common among PWID in London. We corroborate findings indicating SSTIs are associated with risks, e.g. venous access problems, as well as other co-morbid conditions: septicaemia, endocarditis, DVT, and kidney disease. These co-morbidities may impact SSTIs severity and outcomes. Delayed healthcare seeking potentially exacerbates infection severity, which in turn increases poorer health outcomes and complications

Poly(ADP-ribosyl)ation Acts in the DNA Demethylation of Mouse Primordial Germ Cells Also with DNA Damage-Independent Roles

Poly(ADP-ribosyl)ation regulates chromatin structure and transcription driving epigenetic events. In particular, Parp1 is able to directly influence DNA methylation patterns controlling transcription and activity of Dnmt1. Here, we show that ADP-ribose polymer levels and Parp1 expression are noticeably high in mouse primordial germ cells (PGCs) when the bulk of DNA demethylation occurs during germline epigenetic reprogramming in the embryo. Notably, Parp1 activity is stimulated in PGCs even before its participation in the DNA damage response associated with active DNA demethylation. We demonstrate that PARP inhibition impairs both genome-wide and locus-specific DNA methylation erasure in PGCs. Moreover, we evidence that impairment of PARP activity causes a significant reduction of expression of the gene coding for Tet1 hydroxylases involved in active DNA demethylation. Taken together these results demonstrate new and adjuvant roles of poly(ADP-ribosyl)ation during germline DNA demethylation and suggest its possible more general involvement in genome reprogramming

Navigating environmental constraints to injection preparation: the use of saliva and other alternatives to sterile water among unstably housed PWID in London.

BACKGROUND: The United Kingdom is experiencing an increase in drug-related deaths and serious bacterial infections among its most vulnerable citizens. Cuts to essential services, coupled with a growing homeless population, create a challenging environment to tackle this public health crisis. In this paper, we highlight an underexplored environmental constraint faced by people living and injecting drugs on the streets. Access to water for injection is restricted in the UK, due to legislative and financial barriers. Austerity measures, such as public toilet closures, further restrict the ability of people made homeless to access clean water and protect themselves from health harms. METHODS: We generated questionnaire (n = 455) and in-depth qualitative interview (n = 32) data with people who inject drugs in London for the Care and Prevent study. Participants provided detail on their life history; drug use, injecting and living environments; health conditions and care seeking practices. FINDINGS: A high proportion of the survey sample reported lifetime history of street homelessness (78%), bacterial infections (65%) and related hospitalisation (30%). Qualitative accounts highlight unsafe, potentially dangerous, injection practices in semi-public spaces. Multiple constraints to sourcing sterile water for injection preparation were reported. Alternatives to sterile water included puddle water, toilet cistern water, whisky, cola soda and saliva. Participants who injected heroin and crack cocaine together unanimously reported adding water at two stages during injection preparation: first, adding water as a vehicle for heroin (which was then heated); second, adding cold water to the heroin mixture prior to adding the crack cocaine. This new finding of a stage addition of solvent may represent an additional risk of infection. CONCLUSION: Currently, harm reduction equipment and resources for safe injecting are not meeting the needs of people who inject drugs who are street homeless or unstably housed. Preparation of injections with non-sterile water sources could precipitate bacterial and fungal infections, particularly when used without the application of heat. It is crucial that water for injection, also skin cleaning, is made available for the unstably housed and that harm reduction messaging is tailored to speak to the everyday realities of people who prepare and inject drugs in public spaces

Risk perception and affective state on work exhaustion in obstetrics during the COVID-19 pandemic

A multicenter cross-sectional survey study involving four Italian University Hospitals was performed to test the hypothesis that negative affect and positive affect (affective dimensions) mediate the association between risk perception (perceived risk of infection and death; cognitive dimensions) and the feeling of work exhaustion (WE) among obstetrics healthcare providers (HCPs) during the Coronavirus Disease 2019 (COVID-19) pandemic. Totally, 570 obstetrics HCPs were invited to complete the 104-item IPSICO survey in May 2020. A theoretical model built on the tested hypothesis was investigated by structural equation modelling. The model explained 32.2% of the WE variance. Only negative affect mediated the association between cognitive dimensions and WE and also the association between WE and psychological well-being before the pandemic, experiences of stressful events, female gender, and dysfunctional coping. Non-mediated associations with WE were observed for work perceived as a duty, experience of stressful events, support received by colleagues, and the shift strategy. Only previous psychological well-being, support by colleagues, and shift strategies were inversely associated with WE. Based on study results, monitoring negative than positive affect appears superior in predicting WE, with practical implications for planning psychological interventions in HCPs at the individual, interpersonal, and organizational levels

Markers of Liver Function as Potential Prognostic Indicators of SARS-CoV-2 infection: A Retrospective Analysis during the First and Second Waves of COVID-19 Pandemic

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic is known to cause a predominant respiratory disease, although extrapulmonary manifestations can also occur. One of the targets of Coronavirus disease 2019 (COVID-19) is the hepatobiliary system. The present study aims to describe the correlation between the increase of liver damage markers (i.e. alanine aminotransferase [ALT], aspartate aminotransferase [AST], total bilirubin [TB]) and COVID-19 outcomes (i.e., in-hospital mortality [IHM] and intensive care unit [ICU] transfer). Methods: All patients with confirmed SARS-CoV-2 infection admitted to the Infectious Diseases Unit of the St. Anna University-Hospital of Ferrara from March 2020 to October 2021 were retrospectively included in this single-centre study. ALT, AST and TB levels were tested in all patients and IHM or ICU transfer were considered as main outcomes. Co-morbidities were assessed using Charlson Comorbidity Index. Results: A total of 106 patients were retrieved. No hepatic marker was able to predict IHM, whereas all of them negatively predicted ICU transfer (ALT: OR 1.005, 95%CI 1.001-1.009, p= 0.011; AST: OR 1.018, 95%CI 1.006-1.030, p= 0.003; TB: OR 1.329, 95%CI 1.025-1.724, p= 0.032). Age was the only parameter significantly related to mortality. Conclusions: The present study, by correlating liver damage markers with COVID-19 outcome, showed that an increase of ALT, AST and TB predicted patients' severity, although not mortality

'Care and Prevent': rationale for investigating skin and soft tissue infections and AA amyloidosis among people who inject drugs in London.

BACKGROUND: Skin and soft tissue infections (SSTIs) are a leading cause of morbidity and mortality among people who inject drugs (PWID). International data indicate up to one third of PWID have experienced an SSTI within the past month. Complications include sepsis, endocarditis and amyloid A (AA) amyloidosis. AA amyloidosis is a serious sequela of chronic SSTI among PWID. Though there is a paucity of literature reporting on AA amyloidosis among PWID, what has been published suggests there is likely a causal relationship between AA amyloidosis and injecting-related SSTI. If left untreated, AA amyloidosis can lead to renal failure; premature mortality among diagnosed PWID is high. Early intervention may reverse disease. Despite the high societal and individual burden of SSTI among PWID, empirical evidence on the barriers and facilitators to injecting-related SSTI prevention and care or the feasibility and acceptability of AA amyloidosis screening and treatment referral are limited. This study aims to fill these gaps and assess the prevalence of AA amyloidosis among PWID. METHODS: Care and Prevent is a UK National Institute for Health Research-funded mixed-methods study. In five phases (P1-P5), we aim to assess the evidence for AA amyloidosis among PWID (P1); assess the feasibility of AA amyloidosis screening, diagnostic and treatment referral among PWID in London (P2); investigate the barriers and facilitators to AA amyloidosis care (P3); explore SSTI protection and risk (P4); and co-create harm reduction resources with the affected community (P5). This paper describes the conceptual framework, methodological design and proposed analysis for the mixed-methods multi-phase study. RESULTS: We are implementing the Care and Prevent protocol in London. The systematic review component of the study has been completed and published. Care and Prevent will generate an estimate of AA amyloidosis prevalence among community recruited PWID in London, with implications for the development of screening recommendations and intervention implementation. We aim to recruit 400 PWID from drug treatment services in London, UK. CONCLUSIONS: Care and Prevent is the first study to assess screening feasibility and the prevalence of positive proteinuria, as a marker for AA amyloidosis, among PWID accessing drug treatment services. AA amyloidosis is a serious, yet under-recognised condition for which early intervention is available but not employed

Injecting-related health harms and overuse of acidifiers among people who inject heroin and crack cocaine in London: a mixed-methods study.

BACKGROUND: Venous access is a priority for people who inject drugs (PWID). Damage and scarring of peripheral veins can exacerbate health harms, such as skin and soft tissue infections (SSTI), and promote transitions to femoral and subcutaneous injecting. Brown heroin available in Europe requires acidification for injection preparation. In this paper, we present mixed-methods data to explore our hypothesis of a link between overly acidic injection solutions, venous damage and SSTI risk. METHODS: We present a structured survey (n = 455) and in-depth qualitative interview (n = 31) data generated with PWID in London for the Care & Prevent study. Participants provided life history data and detail on injecting environments and drug preparation practices, including the use of acidifiers. Bivariate and multivariate analyses were conducted using a logistic regression for binary outcomes to explore associations between outcomes and excessive acidifier use. Grounded theory principles informed inductive qualitative analysis. Mixed-methods triangulation was iterative with results comparison informing the direction and questions asked of further analyses. RESULTS: Of the 455 participants, most (92%) injected heroin and/or crack cocaine, with 84% using citric as their primary acid for drug preparation. Overuse of acidifier was common: of the 418 who provided an estimate, 36% (n = 150) used more than ½ a sachet, with 30% (n = 127) using a whole sachet or more. We found associations between acidifier overuse, femoral injecting and DVT, but not SSTI. Qualitative accounts highlight the role of poor heroin quality, crack cocaine use, information and manufacturing constraints in acidifier overuse. Painful injections and damage to peripheral veins were common and often attributed to the use of citric acid. CONCLUSIONS: To reduce injecting-related injury and associated consequences, it is crucial to understand the interplay of environmental and practice-based risks underpinning venous damage among PWID. Overuse of acidifier is a modifiable risk factor. In the absence of structural supports such as safe injecting facilities or the prescribing of pharmaceutical diamorphine, there is an urgent need to revisit injecting paraphernalia design and distribution in order to alleviate health harms and distress among the most marginalised

Snoring, hypertension and Type 2 diabetes in obesity. Protection by physical activity

Sleep-related breathing disorders are recognized as major health problems in obesity. They are involved in both hypertension and Type 2 diabetes, through mechanisms possibly related to increased sympathetic tone. We studied the association of habitual snoring with diabetes, hypertension, weight cycling and physical activity in a large Italian database of treatment-seeking obese subjects. Clinical and behavioral data were assessed by standardized questionnaires. Consecutive data of 1890 obese patients were analyzed [average body mass index (BMI), 38.2 kg/m(2), median age: 46 yr, 78% females], from 25 obesity Italian centers, with low prevalence of clinical manifestations of cardiovascular disease. Habitual snoring was reported in 56% of the cases, and was associated with day-time sleepiness. The prevalence increased with obesity class and waist circumference, and was positively associated with weight cycling and weight gain since the age of 20, and smoking. Regular physical activity had a protective effect. Snoring was associated with diabetes and hypertension at univariate analysis, but in multivariate analysis an independent effect was only observed for hypertension. After adjustment for age, gender and BMI, physical activity maintained an independent, protective effect on both snoring (odds ratio 0.65, 95% confidence interval 0.49-0.84; p = 0.001), diabetes (0.50, 0.30-0.86; p = 0.011) and hypertension (0.71, 0.53-0.95; p = 0.023). We conclude that in treatment-seeking, obese subjects with low prevalence of cardiovascular disease, snoring independently increases the risk of hypertension, whereas physical activity exerts a protection on both snoring and complications. These data underline the importance of lifestyle interventions to limit the burden of obesity and associated diseases.ABSTRACT. Sleep-related breathing disorders are recognized as major health problems in obesity. They are involved in both hypertension and Type 2 diabetes, through mechanisms possibly related to increased sympathetic tone. We studied the association of habitual snoring with diabetes, hypertension, weight cycling and physical activity in a large Italian database of treatmentseeking obese subjects. Clinical and behavioral data were assessed by standardized questionnaires. Consecutive data of 1890 obese patients were analyzed [average body mass index (BMI), 38.2 kg/m2, median age: 46 yr, 78% females], from 25 obesity Italian centers, with low prevalence of clinical manifestations of cardiovascular disease. Habitual snoring was reported in 56% of the cases, and was associated with day-time sleepiness. The prevalence increased with obesity class and waist circumference, and was positively associated with weight cycling and weight gain since the age of 20, and smoking. Regular physical activity had a protective effect. Snoring was associated with diabetes and hypertension at univariate analysis, but in multivariate analysis an independent effect was only observed for hypertension. After adjustment for age, gender and BMI, physical activity maintained an independent, protective effect on both snoring (odds ratio 0.65, 95% confidence interval 0.49–0.84; p=0.001), diabetes (0.50, 0.30–0.86; p=0.011) and hypertension (0.71, 0.53–0.95; p=0.023). We conclude that in treatment-seeking, obese subjects with low prevalence of cardiovascular disease, snoring independently increases the risk of hypertension, whereas physical activity exerts a protection on both snoring and complications. These data underline the importance of lifestyle interventions to limit the burden of obesity and associated diseases