Gut microbiota produces biofilm-associated amyloids with potential for neurodegeneration.

Age-related neurodegenerative diseases involving amyloid aggregation remain one of the biggest challenges of modern medicine. Alterations in the gastrointestinal microbiome play an active role in the aetiology of neurological disorders. Here, we dissect the amyloidogenic properties of biofilm-associated proteins (BAPs) of the gut microbiota and their implications for synucleinopathies. We demonstrate that BAPs are naturally assembled as amyloid-like fibrils in insoluble fractions isolated from the human gut microbiota. We show that BAP genes are part of the accessory genomes, revealing microbiome variability. Remarkably, the abundance of certain BAP genes in the gut microbiome is correlated with Parkinson's disease (PD) incidence. Using cultured dopaminergic neurons and Caenorhabditis elegans models, we report that BAP-derived amyloids induce α-synuclein aggregation. Our results show that the chaperone-mediated autophagy is compromised by BAP amyloids. Indeed, inoculation of BAP fibrils into the brains of wild-type mice promote key pathological features of PD. Therefore, our findings establish the use of BAP amyloids as potential targets and biomarkers of α-synucleinopathies.

Lessons learned from a multi-data source research collaboration: The mirabegron post-authorization safety study program.

BACKGROUND: Many factors contribute to developing and conducting a successful multi-data source, non-interventional, post-authorization safety study (NI-PASS) for submission to multiple health authorities. Such studies are often large undertakings; evaluating and sharing lessons learned can provide useful insights to others considering similar studies. OBJECTIVES: We discuss challenges and key methodological and organizational factors that led to the delivery of a successful post-marketing requirement (PMR)/PASS program investigating the risk of cardiovascular and cancer events among users of mirabegron, an oral medication for the treatment of overactive bladder. RESULTS: We provide context and share learnings, including sections on research program collaboration, scientific transparency, organizational approach, mitigation of uncertainty around potential delays, validity of study outcomes, selection of data sources and optimizing patient numbers, choice of comparator groups and enhancing precision of estimates of associations, potential confounding and generalizability of study findings, and interpretation of results. CONCLUSIONS: This large PMR/PASS program was a long-term commitment from all parties and benefited from an effective coordinating center and extensive scientific interactions across research partners, scientific advisory board, study sponsor, and health authorities, and delivered useful learnings related to the design and organization of multi-data source NI-PASS.

In vitro modeling of polyclonal infection dynamics within the human airways by Haemophilus influenzae differential fluorescent labeling.

IMPORTANCE: Genomic diversity of nontypeable H. influenzae strains confers phenotypic heterogeneity. Multiple strains of H. influenzae can be simultaneously isolated from clinical specimens, but we lack detailed information about polyclonal infection dynamics by this pathogen. A long-term barrier to our understanding of this host-pathogen interplay is the lack of genetic tools for strain engineering and differential labeling. Here, we present a novel plasmid toolkit named pTBH (toolbox for Haemophilus), with standardized modules for fluorescent or bioluminescent labeling, adapted to H. influenzae requirements but designed to be versatile so it can be utilized in other bacterial species. We present detailed experimental and quantitative image analysis methods, together with proof-of-principle examples, and show the ample possibilities of 3D microscopy, combined with quantitative image analysis, to model H. influenzae polyclonal infection lifestyles and unravel the co-habitation and co-infection dynamics of this respiratory pathogen.

Bacillus thuringiensis Cyt Proteins as Enablers of Activity of Cry and Tpp Toxins against Aedes albopictus.

Aedes albopictus is a species of mosquito, originally from Southeast Asia, that belongs to the Culicidae family and the Dipteran insect order. The distribution of this vector has rapidly changed over the past decade, making most of the temperate territories in the world vulnerable to important human vector-borne diseases such as dengue, yellow fever, zika or chikungunya. Bacillus thuringiensis var. israeliensis (Bti)-based insecticides represent a realistic alternative to the most common synthetic insecticides for the control of mosquito larvae. However, several studies have revealed emerging resistances to the major Bti Crystal proteins such as Cry4Aa, Cry4Ba and Cry11Aa, making the finding of new toxins necessary to diminish the exposure to the same toxicity factors overtime. Here, we characterized the individual activity of Cyt1Aa, Cry4Aa, Cry4Ba and Cry11Aa against A. albopictus and found a new protein, Cyt1A-like, that increases the activity of Cry11Aa more than 20-fold. Additionally, we demonstrated that Cyt1A-like facilitates the activity three new Bti toxins: Cry53-like, Cry56A-like and Tpp36-like. All in all, these results provide alternatives to the currently available Bti products for the control of mosquito populations and position Cyt proteins as enablers of activity for otherwise non-active crystal proteins.

Post-Authorization Safety Studies of Acute Liver Injury and Severe Complications of Urinary Tract Infection in Patients with Type 2 Diabetes Exposed to Dapagliflozin in a Real-World Setting.

INTRODUCTION: At the time of dapagliflozin's approval in Europe (2012) to treat patients with type 2 diabetes mellitus, concerns regarding acute liver injury and severe complications of urinary tract infection (sUTI) led to two post-authorization safety (PAS) studies of these outcomes to monitor the safety of dapagliflozin in real-world use. OBJECTIVE: To investigate the incidence of hospitalization for acute liver injury (hALI) or sUTI (pyelonephritis or urosepsis) among patients initiating dapagliflozin compared with other glucose-lowering drugs (GLDs). METHODS: These two noninterventional cohort studies identified initiators of dapagliflozin and comparator GLDs in November 2012-February 2019 using data from three longitudinal, population-based data sources: Clinical Practice Research Datalink (UK), the HealthCore Integrated Research Database (USA), and the Medicare database (USA). Outcomes (hALI and sUTI) were identified with electronic algorithms. Incidence rates were estimated by exposure group. Incidence rate ratios (IRRs) were calculated comparing dapagliflozin to comparator GLDs, using propensity score trimming and stratification to address confounding. The sUTI analyses were conducted separately by sex. RESULTS: In all data sources, hALI and sUTI incidence rates were generally lower in dapagliflozin initiators than comparator GLD initiators. The adjusted IRR (95% confidence interval) pooled across data sources for hALI was 0.85 (0.59-1.24) and for sUTI was 0.76 (0.60-0.96) in females and 0.74 (0.56-1.00) in males. Findings from sensitivity analyses were largely consistent with the primary analyses. CONCLUSIONS: These real-world studies do not suggest increased risks of hALI or sUTI, and they suggest a potential decreased risk of sUTI with dapagliflozin exposure compared with other GLDs.

Post-Authorization Safety Study of Hospitalization for Acute Kidney Injury in Patients with Type 2 Diabetes Exposed to Dapagliflozin in a Real-World Setting.

INTRODUCTION: Dapagliflozin is a sodium-glucose cotransporter 2 inhibitor approved to treat type 2 diabetes mellitus (T2DM), among other conditions. When dapagliflozin was approved in Europe for treating T2DM (2012), potential safety concerns regarding its effect on kidney function resulted in this post-authorization safety study to assess hospitalization for acute kidney injury (hAKI) among dapagliflozin initiators in a real-world setting. OBJECTIVE: The aim of this study was to evaluate the incidence of hAKI in adults with T2DM initiating dapagliflozin compared with other glucose-lowering drugs (GLDs). METHODS: This noninterventional cohort study identified new users of dapagliflozin and comparator GLDs from November 2012 to February 2019 from three longitudinal, population-based data sources: Clinical Practice Research Datalink (CPRD; United Kingdom), the HealthCore Integrated Research Database (HIRD; United States [US]), and Medicare (US). Electronic algorithms identified occurrences of hAKI, from which a sample underwent validation. Incidence rates for hAKI were calculated, and incidence rate ratios (IRRs) compared hAKI in dapagliflozin with comparator GLDs. Propensity score trimming and stratification were conducted for confounding adjustment. RESULTS: In all data sources, dapagliflozin initiators had a lower hAKI incidence rate than comparator GLD initiators (adjusted IRRs: CPRD, 0.44 [95% confidence interval (CI), 0.22-0.86]; HIRD, 0.76 [95% CI, 0.62-0.93]; Medicare, 0.69 [95% CI, 0.59-0.79]). The adjusted IRR pooled across the data sources was 0.70 (95% CI, 0.62-0.78). Results from sensitivity and stratified analyses were consistent with the primary analysis. CONCLUSIONS: This study, with > 34,000 person-years of real-world dapagliflozin exposure, suggests a decreased risk of hAKI in patients with T2DM exposed to dapagliflozin, aligning with results from dapagliflozin clinical trials. STUDY REGISTRATION: European Union Post-Authorisation Studies Register, EUPAS 11684; ClinicalTrials.gov, NCT02695082.

Bacterial biofilm functionalization through Bap amyloid engineering.

Biofilm engineering has emerged as a controllable way to fabricate living structures with programmable functionalities. The amyloidogenic proteins comprising the biofilms can be engineered to create self-assembling extracellular functionalized surfaces. In this regard, facultative amyloids, which play a dual role in biofilm formation by acting as adhesins in their native conformation and as matrix scaffolds when they polymerize into amyloid-like fibrillar structures, are interesting candidates. Here, we report the use of the facultative amyloid-like Bap protein of Staphylococcus aureus as a tool to decorate the extracellular biofilm matrix or the bacterial cell surface with a battery of functional domains or proteins. We demonstrate that the localization of the functional tags can be change by simply modulating the pH of the medium. Using Bap features, we build a tool for trapping and covalent immobilizing molecules at bacterial cell surface or at the biofilm matrix based on the SpyTag/SpyCatcher system. Finally, we show that the cell wall of several Gram-positive bacteria could be functionalized through the external addition of the recombinant engineered Bap-amyloid domain. Overall, this work shows a simple and modulable system for biofilm functionalization based on the facultative protein Bap.

Staphylococcus aureus ftnA 3'-Untranslated Region Modulates Ferritin Production Facilitating Growth Under Iron Starvation Conditions.

Iron acquisition and modulation of its intracellular concentration are critical for the development of all living organisms. So far, several proteins have been described to be involved in iron homeostasis. Among them, ferritins act as the major iron storage proteins, sequestering internalized iron and modulating its concentration inside bacterial cells. We previously described that the deletion of the 3'-untranslated region (3'UTR) of the ftnA gene, which codes for ferritin in Staphylococcus aureus, increased the ftnA mRNA and ferritin levels. Here, we show that the ferritin levels are affected by RNase III and PNPase, which target the ftnA 3'UTR. Rifampicin mRNA stability experiments revealed that the half-life of the ftnA mRNA is affected by both RNase III and the ftnA 3'UTR. A transcriptional fusion of the ftnA 3'UTR to the gfp reporter gene decreased green fluorescent protein (GFP) expression, indicating that the ftnA 3'UTR could work as an independent module. Additionally, a chromosomal deletion of the ftnA 3'UTR impaired S. aureus growth under conditions of iron starvation. Overall, this work highlights the biological relevance of the ftnA 3'UTR for iron homeostasis in S. aureus.

Regulation of Heterogenous LexA Expression in Staphylococcus aureus by an Antisense RNA Originating from Transcriptional Read-Through upon Natural Mispairings in the sbrB Intrinsic Terminator.

Bacterial genomes are pervasively transcribed, generating a wide variety of antisense RNAs (asRNAs). Many of them originate from transcriptional read-through events (TREs) during the transcription termination process. Previous transcriptome analyses revealed that the lexA gene from Staphylococcus aureus, which encodes the main SOS response regulator, is affected by the presence of an asRNA. Here, we show that the lexA antisense RNA (lexA-asRNA) is generated by a TRE on the intrinsic terminator (TTsbrB) of the sbrB gene, which is located downstream of lexA, in the opposite strand. Transcriptional read-through occurs by a natural mutation that destabilizes the TTsbrB structure and modifies the efficiency of the intrinsic terminator. Restoring the mispairing mutation in the hairpin of TTsbrB prevented lexA-asRNA transcription. The level of lexA-asRNA directly correlated with cellular stress since the expressions of sbrB and lexA-asRNA depend on the stress transcription factor SigB. Comparative analyses revealed strain-specific nucleotide polymorphisms within TTsbrB, suggesting that this TT could be prone to accumulating natural mutations. A genome-wide analysis of TREs suggested that mispairings in TT hairpins might provide wider transcriptional connections with downstream genes and, ultimately, transcriptomic variability among S. aureus strains.

The 3'UTR-derived sRNA RsaG coordinates redox homeostasis and metabolism adaptation in response to glucose-6-phosphate uptake in Staphylococcus aureus.

Staphylococcus aureus RsaG is a 3'-untranslated region (3'UTR) derived sRNA from the conserved uhpT gene encoding a glucose-6-phosphate (G6P) transporter expressed in response to extracellular G6P. The transcript uhpT-RsaG undergoes degradation from 5'- to 3'-end by the action of the exoribonucleases J1/J2, which are blocked by a stable hairpin structure at the 5'-end of RsaG, leading to its accumulation. RsaG together with uhpT is induced when bacteria are internalized into host cells or in the presence of mucus-secreting cells. Using MS2-affinity purification coupled with RNA sequencing, several RNAs were identified as targets including mRNAs encoding the transcriptional factors Rex, CcpA, SarA, and the sRNA RsaI. Our data suggested that RsaG contributes to the control of redox homeostasis and adjusts metabolism to changing environmental conditions. RsaG uses different molecular mechanisms to stabilize, degrade, or repress the translation of its mRNA targets. Although RsaG is conserved only in closely related species, the uhpT 3'UTR of the ape pathogen S. simiae harbors an sRNA, whose sequence is highly different, and which does not respond to G6P levels. Our results hypothesized that the 3'UTRs from UhpT transporter encoding mRNAs could have rapidly evolved to enable adaptation to host niches.

Anti-Biofilm Molecules Targeting Functional Amyloids.

The choice of an effective therapeutic strategy in the treatment of biofilm-related infections is a significant issue. Amyloids, which have been historically related to human diseases, are now considered to be prevailing structural components of the biofilm matrix in a wide range of bacteria. This assumption creates the potential for an exciting research area, in which functional amyloids are considered to be attractive targets for drug development to dissemble biofilm structures. The present review describes the best-characterized bacterial functional amyloids and focuses on anti-biofilm agents that target intrinsic and facultative amyloids. This study provides a better understanding of the different modes of actions of the anti-amyloid molecules to inhibit biofilm formation. This information can be further exploited to improve the therapeutic strategies to combat biofilm-related infections.

Cardiovascular Risk in Users of Mirabegron Compared with Users of Antimuscarinic Treatments for Overactive Bladder: Findings from a Non-Interventional, Multinational, Cohort Study.

INTRODUCTION: During clinical trials, mirabegron, a ß3-adrenoreceptor agonist, was associated with increased vital signs vs placebo in patients with overactive bladder. OBJECTIVE: The purpose of this study was to compare incidence rates of adverse cardiovascular (CV) outcomes following mirabegron or antimuscarinic use. METHODS: We conducted an observational post-marketing safety study utilising real-world data. The study population was identified within five sources: Danish and Swedish National Registers, Clinical Practice Research Datalink (UK), Optum (USA) and Humana (USA). Episodes of time when patients were new users of mirabegron or antimuscarinics (October 2012-December 2018) were sourced from prescriptions and matched on propensity scores. Occurrences of major adverse cardiovascular events (MACE), acute myocardial infarction (AMI), stroke, CV mortality and all-cause mortality were identified. Outcome incidence rates and hazard ratios from Cox models were estimated. RESULTS: Overall, 152,026 mirabegron and 152,026 antimuscarinic episodes were matched. The population consisted of 63.1% women and 72.6% were ≥ 65 years old. There were no appreciable differences in the incidence rates of MACE, AMI or stroke between users of mirabegron and antimuscarinics. Incidence rates of CV mortality (hazard ratio 0.83, 95% confidence interval 0.73-0.95) and all-cause mortality (hazard ratio 0.80, 95% confidence interval 0.76-0.84) were no higher with mirabegron vs antimuscarinics. Results restricted to episodes at high risk for CV events or stratified by age (< 65 years, ≥ 65 years) or prior overactive bladder medication use were consistent with overall findings. CONCLUSIONS: This large, multinational study found no higher risk of MACE, AMI, stroke, CV mortality or all-cause mortality among users of mirabegron relative to users of antimuscarinics.

During clinical trials, mirabegron, which is a treatment for overactive bladder, was associated with small increases in heart rate and blood pressure. This study was conducted to compare the frequency of cardiac events following the use of mirabegron or antimuscarinics, a group of treatments also used to treat overactive bladder. We obtained the data for this study from four countries: Denmark, Sweden, the UK and the USA. We identified people who were new users of mirabegron or antimuscarinics from 2012 to 2018 using prescription or dispensing records. Occurrences of major cardiac events, heart attack, stroke, death due to cardiac events and death from any cause were evaluated. Overall, we identified 152,026 times when mirabegron or antimuscarinics were each used as new treatments. Most of the people in the study were women (63.1%) and at least 65 years old (72.6%). There were no notable differences between the treatment groups with regard to how often major cardiac events, heart attack or stroke occurred. Further, death due to cardiac events and death from any cause were no higher with mirabegron compared with antimuscarinics. We obtained similar results when the data were assessed for patients who were at high risk for cardiac events or split by age (less than 65 years or at least 65 years) or a history of overactive bladder medication use. In conclusion, this large study involving data from several countries found no higher risk of major cardiac events, heart attack, stroke or death among people prescribed mirabegron compared with antimuscarinics.

Phase Variation in HMW1A Controls a Phenotypic Switch in Haemophilus influenzae Associated with Pathoadaptation during Persistent Infection.

Genetic variants arising from within-patient evolution shed light on bacterial adaptation during chronic infection. Contingency loci generate high levels of genetic variation in bacterial genomes, enabling adaptation to the stringent selective pressures exerted by the host. A significant gap in our understanding of phase-variable contingency loci is the extent of their contribution to natural infections. The human-adapted pathogen nontypeable Haemophilus influenzae (NTHi) causes persistent infections, which contribute to underlying disease progression. The phase-variable high-molecular-weight (HMW) adhesins located on the NTHi surface mediate adherence to respiratory epithelial cells and, depending on the allelic variant, can also confer high epithelial invasiveness or hyperinvasion. In this study, we characterize the dynamics of HMW-mediated hyperinvasion in living cells and identify a specific HMW binding domain shared by hyperinvasive NTHi isolates of distinct pathological origins. Moreover, we observed that HMW expression decreased over time by using a longitudinal set of persistent NTHi strains collected from chronic obstructive pulmonary disease (COPD) patients, resulting from increased numbers of simple-sequence repeats (SSRs) downstream of the functional P2hmw1A promoter, which is the one primarily driving HMW expression. Notably, the increased SSR numbers at the hmw1 promoter region also control a phenotypic switch toward lower bacterial intracellular invasion and higher biofilm formation, likely conferring adaptive advantages during chronic airway infection by NTHi. Overall, we reveal novel molecular mechanisms of NTHi pathoadaptation based on within-patient lifestyle switching controlled by phase variation. IMPORTANCE Human-adapted bacterial pathogens have evolved specific mechanisms to colonize their host niche. Phase variation is a contingency strategy to allow adaptation to changing conditions, as phase-variable bacterial loci rapidly and reversibly switch their expression. Several NTHi adhesins are phase variable. These adhesins are required for colonization but also immunogenic, in such a way that bacteria with lower adhesin levels are better equipped to survive an immune response, making their contribution to natural infections unclear. We show here that the major NTHi adhesin HMW1A displays allelic variation, which can drive a phase-variable epithelial hyperinvasion phenotype. Over time, hmw1A phase variation lowers adhesin expression, which controls an NTHi lifestyle switch from high epithelial invasiveness to lower invasion and higher biofilm formation. This reversible loss of function aligns with the previously stated notion that epithelial infection is essential for NTHi infection establishment, but once established, persistence favors gene inactivation, in this case facilitating biofilm growth.

RNA thermoswitches modulate Staphylococcus aureus adaptation to ambient temperatures.

Thermoregulation of virulence genes in bacterial pathogens is essential for environment-to-host transition. However, the mechanisms governing cold adaptation when outside the host remain poorly understood. Here, we found that the production of cold shock proteins CspB and CspC from Staphylococcus aureus is controlled by two paralogous RNA thermoswitches. Through in silico prediction, enzymatic probing and site-directed mutagenesis, we demonstrated that cspB and cspC 5'UTRs adopt alternative RNA structures that shift from one another upon temperature shifts. The open (O) conformation that facilitates mRNA translation is favoured at ambient temperatures (22°C). Conversely, the alternative locked (L) conformation, where the ribosome binding site (RBS) is sequestered in a double-stranded RNA structure, is folded at host-related temperatures (37°C). These structural rearrangements depend on a long RNA hairpin found in the O conformation that sequesters the anti-RBS sequence. Notably, the remaining S. aureus CSP, CspA, may interact with a UUUGUUU motif located in the loop of this long hairpin and favour the folding of the L conformation. This folding represses CspB and CspC production at 37°C. Simultaneous deletion of the cspB/cspC genes or their RNA thermoswitches significantly decreases S. aureus growth rate at ambient temperatures, highlighting the importance of CspB/CspC thermoregulation when S. aureus transitions from the host to the environment.

A study of cancer occurrence in users of mirabegron and antimuscarinic treatments for overactive bladder.

OBJECTIVE: This post-authorization safety study (EU PAS Register Number: EUPAS16088) was designed to compare the incidence of cancer outcomes in patients treated with mirabegron versus antimuscarinic medications. METHODS: Cohorts of mirabegron initiators during 2012-2018 were propensity-score matched to antimuscarinic medication initiators within real-world data sources (Danish National Registers, Swedish National Registers, Clinical Practice Research Datalink [UK], Optum [US], and Humana [US]). Incident cancer cases were identified during follow-up from direct linkage to cancer registers or validated through medical record review or through physician questionnaires. Comparisons of sex-specific composite cancer outcomes (cancer of the lung/bronchus, colon/rectum, melanoma of skin, urinary bladder, non-Hodgkin lymphoma, kidney/renal pelvis, pancreas, prostate in men and breast and uterus in women) were made overall and for person-time in the first year and after the first year following start of treatment, for all ages and for the subgroup ≥65 years. RESULTS: Among the 80,637 mirabegron initiators matched to 169,885 antimuscarinic medication initiators, 68% were at least 65 years of age and 66% were women. Over 5000 incident cancer cases were observed overall. Incidence rates were higher for men than women for composite and individual cancer outcomes. The pooled fixed effects hazard ratios for composite cancer outcomes (all ages) were 1.05 (95% confidence interval [CI]: 0.98-1.14) for women and 1.06 (95% CI: 0.98-1.14) for men. Results were similar in persons ≥65 years. CONCLUSIONS: The results suggest no association between mirabegron use and risk of cancer, compared with antimuscarinic medications, in either men or women. Registration: EU PAS Register Number: EUPAS16088.

Preoperative transarterial chemoembolization for laparoscopic liver resection in Child A cirrhotic patients with hepatocellular carcinoma.

PURPOSE: Laparoscopic liver resection (LLR) for hepatocellular carcinoma (HCC) offers better short-term results than open surgery with similar long-term results although it is technically difficult due to the risk of bleeding. METHODS: This study included patients with HCC in Child A cirrhosis who underwent TACE before LLR between 2009 and 2019. The primary endpoint was to analyze the intraoperative and early results of this technique. We also analyzed the long-term outcomes. Patients with and without clinically significant portal hypertension (CSPH) were compared. RESULTS: A total of 44 cirrhotic patients with HCC were included (24 CSPH and 20 non-CSPH). The Pringle maneuver was used in two cases (4.5%), mean blood losses was 100 ml (range 50-200), and three patients (6.8%) required a blood transfusion. The degree of necrosis achieved was greater than 90% in 27 patients (61.4%). At 1, 3, and 5 years, overall survival was 97.7%, 81.5%, and 63.4%, respectively, and disease-free survival was 85.2%, 52.5%, and 34.5%, respectively. There were no statistically significant differences between non-CSPH and CSPH groups regarding intraoperative, early, and long-term outcomes. CONCLUSION: In our experience, TACE could be beneficial to perform LLR in HCC Child-Pugh A patients with and without CSPH without serious complications and similar oncological outcomes.

Long-Term Risk of Skin Cancer and Lymphoma in Users of Topical Tacrolimus and Pimecrolimus: Final Results from the Extension of the Cohort Study Protopic Joint European Longitudinal Lymphoma and Skin Cancer Evaluation (JOELLE).

PURPOSE: Evidence is insufficient to infer whether topical calcineurin inhibitors (TCIs; tacrolimus and pimecrolimus) cause malignancy. The study objective was to estimate the long-term risk of skin cancer and lymphoma associated with topical TCI use in adults and children, separately. PATIENTS AND METHODS: A cohort study in Denmark, Sweden, UK, and the Netherlands was conducted. Adjusted incidence rate ratios (IRRs) and 95% confidence intervals (CIs) were calculated for nonmelanoma skin cancer (NMSC), melanoma, cutaneous T-cell lymphoma (CTCL), non-Hodgkin lymphoma (NHL) excluding CTCL, and Hodgkin lymphoma (HL) in new users of TCIs versus users of moderate/high-potency topical corticosteroids. RESULTS: The study included 126,908/61,841 adults and 32,605/27,961 children initiating treatment with tacrolimus/pimecrolimus, respectively. Follow-up was ≥10 years for 19% of adults and 32% of children. Incidence rate ratios and (95% confidence intervals) for tacrolimus versus corticosteroid users in adults were <1 for melanoma, non-Hodgkin lymphoma, and Hodgkin lymphoma; and 1.80 (1.25-2.58) for cutaneous T-cell lymphoma. For pimecrolimus, IRRs in adults were <1 for non-Hodgkin lymphoma, cutaneous T-cell lymphoma, and Hodgkin's lymphoma; and 1.21 (1.03-1.41) for melanoma; and 1.28 (1.20-1.35) for nonmelanoma skin cancer. In children, results were inconclusive due to few events. In adults, incidence rate ratios ≥5 years after first topical calcineurin inhibitor exposure were not higher than in overall analyses. CONCLUSION: Overall, we found little evidence associating use of topical calcineurin inhibitors with skin cancer and lymphoma; confounding by indication, surveillance bias, and reverse causation may have influenced these results. Even if causal, the public health impact of these excess risks would be low and confined to the first years of exposure.

Validation of cardiovascular outcomes and risk factors in the Clinical Practice Research Datalink in the United Kingdom.

PURPOSE: Strategies to identify and validate acute myocardial infarction (AMI) and stroke in primary-care electronic records may impact effect measures, but to an unknown extent. Additionally, the validity of cardiovascular risk factors that could act as confounders in studies on those endpoints has not been thoroughly assessed in the United Kingdom Clinical Practice Research Datalink's (CPRD's) GOLD database. We explored the validity of algorithms to identify cardiovascular outcomes and risk factors and evaluated different outcome-identification strategies using these algorithms for estimation of adjusted incidence rate ratios (IRRs). METHODS: First, we identified AMI, stroke, smoking, obesity, and menopausal status in a cohort treated for overactive bladder by applying computerized algorithms to primary care medical records (2004-2012). We validated these cardiovascular outcomes and risk factors with physician questionnaires (gold standard for this analysis). Second, we estimated IRRs for AMI and stroke using algorithm-identified and questionnaire-confirmed cases, comparing these with IRRs from cases identified through linkage with hospitalization/mortality data (best estimate). RESULTS: For AMI, the algorithm's positive predictive value (PPV) was >90%. Initial algorithms for stroke performed less well because of inclusion of codes for prevalent stroke; algorithm refinement increased PPV to 80% but decreased sensitivity by 20%. Algorithms for smoking and obesity were considered valid. IRRs based on questionnaire-confirmed cases only were closer to IRRs estimated from hospitalization/mortality data than IRRs from algorithm-identified cases. CONCLUSIONS: AMI, stroke, smoking, obesity, and postmenopausal status can be accurately identified in CPRD. Physician questionnaire-validated AMI and stroke cases yield IRRs closest to the best estimate.

Assessment of a Sour Water Treatment Unit Using Process Simulation, Parametric Sensitivity, and Exergy Analysis.

In this work, a sour water treatment unit was evaluated combining exergetic analysis and parametric sensitivity analysis. Process simulation was performed using Aspen HYSYS 10.1 following real refinery configurations, and the results were validated with existing data. The parametric sensitivity was evaluated by varying the effect of process variables to identify an alternative case with the best technical performance. The exergy analysis was applied to both base and alternative cases. The outcomes were exergy efficiency by stages, global exergy efficiency, total irreversibilities, and exergy by industrial services. A comparison of both cases was performed to identify opportunities for improvement in real sour water treatment. Results revealed that the overall exergy efficiency for the base case was 44.28%. After improving the technical performance, the overall exergy efficiency decreased to 36.12%; the latter indicated higher irreversibilities due to the increase in the use of industrial services. This finding suggested that those process improvements may affect the performance of this refinery unit from an exergetic point of view.