One pathological hallmark of HIV-1 infection is chronic activation of the immune system, driven, in part, by increased expression of proinflammatory cytokines. The host attempts to counterbalance this prolonged immune activation through compensatory mediators of immune suppression. We recently identified a gene encoding the proinflammatory cytokine IL-32 in microarray studies of HIV-1 infection in lymphatic tissue (LT) and show in this study that increased expression of IL-32 in both gut and LT of HIV-1-infected individuals may have a heretofore unappreciated role as a mediator of immune suppression. We show that: 1) IL-32 expression is increased in CD4(+) T cells, B cells, macrophages, dendritic cells, and epithelial cells in vivo; 2) IL-32 induces the expression of immunosuppressive molecules IDO and Ig-like transcript 4 in immune cells in vitro; and 3) in vivo, IL-32-associated IDO/Ig-like transcript 4 expression in LT macrophages and gut epithelial cells decreases immune activation but also may impair host defenses, supporting productive viral replication, thereby accounting for the correlation between IL-32 levels and HIV-1 replication in LT. Thus, during HIV-1 infection, we propose that IL-32 moderates chronic immune activation to avert associated immunopathology but at the same time dampens the antiviral immune response and thus paradoxically supports HIV-1 replication and viral persistence.

<p><div>(<em>b</em>)Background</<em>b</em>)</div>The pou<em>lt</em>ry industry is in need of effective anti<em>b</em>iotic a<em>lt</em>ernatives to control out<em>b</em>reaks of necrotic enteritis (NE) due to <i>Clostridium perfringens</i>. In the present study, we investigated the effects of dietary supplementation with a <em>b</em>lend of encapsulated essential oils and organic acids (BLJ) on growth performance and gut hea<em>lt</em>h using a coinfection model of NE in <em>b</em>roiler chickens.</p><p><div>(<em>b</em>)Methods</<em>b</em>)</div>Two hundred and eighty-eight one-day-old male Ar<em>b</em>or Acres <em>b</em>roiler chicks were randomly assigned using a 2 × 2 factorial design into two groups fed either 0 or 500 mg/kg dietary BLJ and co-challenged (or not challenged for the control) with <i>Eimeria</i> spp./<i>C. perfringens</i>.</p><p><div>(<em>b</em>)Resu<em>lt</em>s</<em>b</em>)</div>Infected <em>b</em>irds fed the BLJ-supplemented diet exhi<em>b</em>ited an improved feed conversion ratio throughout the trial (<i>P</i> &<em>lt</em>; 0.01), a higher villus height and villus height/crypt depth ratio, and reduced intestinal <i>C. perfringens</i> counts, liver <i>C. perfringens</i> carriage, gut lesion scores and serum fluorescein isothiocyanate dextran (FITC-D) concentrations at 7 d post-infection compared with those of <em>b</em>irds without BLJ supplementation (<i>P</i> &<em>lt</em>; 0.05). NE-infected <em>b</em>irds fed BLJ exhi<em>b</em>ited significantly upregulated claudin-1 and <i>IGF-2</i> mRNA levels (<i>P</i> &<em>lt</em>; 0.05), increased <i>A20</i> mRNA expression and significantly downregulated <i>TRAF-6</i>, <i>TNFSF15</i> and <i>TOLLIP</i> mRNA levels in the jejunum at 7 d post-infection compared with those in <em>b</em>irds without BLJ supplementation (<i>P</i> &<em>lt</em>; 0.05). Compared with the uninfected and untreated <em>b</em>irds, the uninfected <em>b</em>irds fed BLJ displayed increased relative a<em>b</em>undances of <i>Lacto<em>b</em>acillus</i> and <i>Coprococcus</i> <em>b</em>ut reduced Rikenellaceae levels. Compared with the unsupplemented NE-challenged <em>b</em>irds, infected <em>b</em>irds fed BLJ showed an increased relative a<em>b</em>undance of Unclassified_Lachnospiraceae and a significantly decreased relative a<em>b</em>undance of Erysipelotrichaceae.</p><p><div>(<em>b</em>)Conclusion</<em>b</em>)</div>BLJ supplementation improved growth performance and gut hea<em>lt</em>h in NE-infected <em>b</em>roiler chickens <em>b</em>y strengthening the intestinal <em>b</em>arrier function, positively modulating the gut micro<em>b</em>iota community and differentially regulating intestinal immune responses. Our resu<em>lt</em>s also suggested that adding BLJ effectively controlled NE infections after experimental <i>Eimeria</i> and <i>Clostridium perfringens</i> coinfection.</p>

<A<em>b</em>stractText>To identify a synovial fluid (SF) <em>b</em>iomarker profile characteristic of individuals with an inflammatory osteoarthritis (OA) endotype.</A<em>b</em>stractText><A<em>b</em>stractText>A total of 48 knees (of 25 participants) were characterized for an extensive array of SF <em>b</em>iomarkers quantified <em>b</em>y Rules Based Medicine using the high-sensitivity mu<em>lt</em>iplex immunoassay, Myriad Human InflammationMAP® 1.0, which included 47 different cytokines, chemokines, and growth factors related to inflammation. Mu<em>lt</em>ivaria<em>b</em>le regression with generalized estimating equations (GEE) and false discovery rate (FDR) correction was used to assess associations of SF RBM <em>b</em>iomarkers with etarfolatide imaging scores reflecting synovial inflammation; radiographic knee OA severity (<em>b</em>ased on Kellgren-Lawrence (KL) grade, joint space narrowing, and osteophyte scores); knee joint symptoms; and SF <em>b</em>iomarkers associated with activated macrophages and knee OA progression including CD14 and CD163 (shed <em>b</em>y activated macrophages) and elastase (shed <em>b</em>y activated neutrophils).</A<em>b</em>stractText><p><div>(<em>b</em>)RESULTS</<em>b</em>)</div>Significant associations of SF <em>b</em>iomarkers meeting FDR &<em>lt</em>; 0.05 included solu<em>b</em>le (s)VCAM-1 and MMP-3 with synovial inflammation (FDR-adjusted p = 0.025 and 1.06 × 10^-7); sVCAM-1, sICAM-1, TIMP-1, and VEGF with radiographic OA severity (p = 1.85 × 10^-5 to 3.97 × 10^-4); and VEGF, MMP-3, TIMP-1, sICAM-1, sVCAM-1, and MCP-1 with OA symptoms (p = 2.72 × 10^-5 to 0.050). All these SF <em>b</em>iomarkers were highly correlated with macrophage markers CD163 and CD14 in SF (r = 0.43 to 0.90, FDR &<em>lt</em>; 0.05); all <em>b</em>ut MCP-1 were also highly correlated with neutrophil elastase in SF (r = 0.62 to 0.89, FDR &<em>lt</em>; 0.05).</p><A<em>b</em>stractText>A su<em>b</em>set of six SF <em>b</em>iomarkers was related to synovial inflammation in OA, as well as radiographic and symptom severity. These six OA-related SF <em>b</em>iomarkers were specifically linked to indicators of activated macrophages and neutrophils. These resu<em>lt</em>s attest to an inflammatory OA endotype that may serve as the <em>b</em>asis for therapeutic targeting of a su<em>b</em>set of individuals at high risk for knee OA progression.</A<em>b</em>stractText><A<em>b</em>stractText>Written informed consent was received from participants prior to inclusion in the study; the study was registered at ClinicalTrials.gov ( NCT01237405 ) on Novem<em>b</em>er 9, 2010, prior to enrollment of the first participant.</A<em>b</em>stractText>

<A<em>b</em>stractText>Pulmonary reha<em>b</em>ilitation (PR) is an effective, key standard treatment for people with COPD. Nevertheless, low participant uptake, insufficient attendance and high drop-out rates are reported. Investigation is warranted of the <em>b</em>enefits achieved through a<em>lt</em>ernative approaches, such as pulmonary tele-reha<em>b</em>ilitation (PTR).</A<em>b</em>stractText><p><div>(<em>b</em>)OBJECTIVE</<em>b</em>)</div>To investigate whether PTR is superior to conventional PR on 6 min walk distance (6MWD) and secondarily on respiratory symptoms, quality of life, physical activity and lower lim<em>b</em> muscle function in patients with COPD and FEV<su<em>b</em>)1</su<em>b</em>) &<em>lt</em>;50% eligi<em>b</em>le for routine hospital-<em>b</em>ased, outpatient PR.</p><A<em>b</em>stractText>In this single-<em>b</em>linded, mu<em>lt</em>icentre, superiority randomised controlled trial, patients were assigned 1:1 to 10 weeks of groups-<em>b</em>ased PTR (60 min, three times weekly) or conventional PR (90 min, two times weekly). Assessments were performed <em>b</em>y <em>b</em>linded assessors at <em>b</em>aseline, end of intervention and at 22 weeks' follow-up from <em>b</em>aseline. The primary analysis was <em>b</em>ased on the intention-to-treat principle.</A<em>b</em>stractText><p><div>(<em>b</em>)MEASUREMENTS AND MAIN RESULTS</<em>b</em>)</div>The primary outcome was change in 6MWD from <em>b</em>aseline to 10 weeks; 134 participants (74 females, mean±SD age 68±9 years, FEV<su<em>b</em>)1</su<em>b</em>) 33%±9% predicted, 6MWD 327±103 metres) were included and randomised. The analysis showed no <em>b</em>etween-group differences for changes in 6MWD after intervention (9.2 metres (95% CI: -6.6 to 24.9)) or at 22 weeks' follow-up (-5.3 metres (95% CI: -28.9 to 18.3)). More participants completed the PTR intervention (n=57) than conventional PR (n=43) (χ² test p&<em>lt</em>;0.01).</p><p><div>(<em>b</em>)CONCLUSION</<em>b</em>)</div>PTR was not superior to conventional PR on the 6MWD and we found no differences <em>b</em>etween groups. As more participants completed PTR, supervised PTR would <em>b</em>e relevant to compare with conventional PR in a non-inferiority design.(<em>b</em>)Trial registration num<em>b</em>er</<em>b</em>)ClinicalTrials.gov (NCT02667171), 28 January 2016.</p>

<A<em>b</em>stractText>The PRIMA study (ClinicalTrials.gov identifier: NCT00140582) esta<em>b</em>lished that 2 years of rituxima<em>b</em> maintenance after first-line immunochemotherapy significantly improved progression-free survival (PFS) in patients with follicular lymphoma compared with o<em>b</em>servation. Here, we report the final PFS and overall survival (OS) resu<em>lt</em>s from the PRIMA study after 9 years of follow-up and provide a final overview of safety.</A<em>b</em>stractText><p><div>(<em>b</em>)METHODS</<em>b</em>)</div>Patients (> 18 years of age) with previously untreated high-tumor-<em>b</em>urden follicular lymphoma were nonrandomly assigned to receive one of three immunochemotherapy induction regimens. Responding patients were randomly assigned (stratified <em>b</em>y induction regimen, response to induction treatment, treatment center, and geographic region) 1:1 to receive 2 years of rituxima<em>b</em> maintenance (375 mg/m², once every 8 weeks), starting 8 weeks after the last induction treatment, or o<em>b</em>servation (no additional treatment). All patients in the extended follow-up provided their written informed consent (data cutoff: Decem<em>b</em>er 31, 2016).</p><p><div>(<em>b</em>)RESULTS</<em>b</em>)</div>In total, 1,018 patients completed induction treatment and were randomly assigned to rituxima<em>b</em> maintenance (n = 505) or o<em>b</em>servation (n = 513). Consent for the extended follow-up was provided <em>b</em>y 607 patients (59.6%) of 1,018 (rituxima<em>b</em> maintenance, n = 309; o<em>b</em>servation, n = 298). After data cutoff, median PFS was 10.5 years in the rituxima<em>b</em> maintenance arm compared with 4.1 years in the o<em>b</em>servation arm (hazard ratio, 0.61; 95% CI, 0.52 to 0.73; <i>P</i> &<em>lt</em>; .001). No OS difference was seen in patients randomly assigned to rituxima<em>b</em> maintenance or o<em>b</em>servation (hazard ratio, 1.04; 95% CI, 0.77 to 1.40; <i>P</i> = .7948); 10-year OS estimates were approximately 80% in <em>b</em>oth study arms. No new safety signals were o<em>b</em>served.</p><A<em>b</em>stractText>Rituxima<em>b</em> maintenance after induction immunochemotherapy provides a significant long-term PFS, <em>b</em>ut not OS, <em>b</em>enefit over o<em>b</em>servation.</A<em>b</em>stractText>

Chronic Chagas disease cardiomyopathy (CCC), caused by Trypanosoma cruzi, is an inflammatory dilated cardiomyopathy associated with increased circulating levels of TNF-alpha. We investigate whether TNF blockade with Etanercept during the chronic phase of T. cruzi infection could attenuate experimental CCC development. The effect of Etanercept was evaluated after 11 months of T. cruzi infection on survival, parasitism, left ventricular function, intensity of myocarditis, fibrosis, and left ventricular mRNA expression of cytokines and TNF-alpha-induced genes. Left ventricular function was significantly reduced in treated animals as compared to infected untreated animals. Blood and cardiac parasitism as well as survival rate were not altered with Etanercept treatment. Inflammatory infiltrates were located predominantly in the subendocardic region in treated animals, whereas in untreated animals inflammation was scattered throughout the myocardium. Left ventricular mRNA IL-10 expression was significantly higher, and iNOS, significantly lower in treated than in untreated animals. mRNA expression of TNF-alpha, IFN-gamma, TGF-beta, A20 and ANP was similar in both groups. Our results suggest that TNF-alpha/LT-alpha blockade with Etanercept enhances left ventricular dysfunction in T. cruzi-induced chronic cardiomyopathy and the absence of TNF signaling may be deleterious to the failing heart in Chagas disease cardiomyopathy.

Melanocytes produce two chemically distinct types of melanin pigments, eumelanin and pheomelanin. These pigments can be quantitatively analyzed by acidic permanganate oxidation or reductive hydrolysis with hydriodic acid to form pyrrole-2,3,5-tricarboxylic acid or aminohydroxyphenylalanine, respectively. About 30 coat color genes in mice have been cloned, and functions of many of those genes have been elucidated. However, little is known about the interacting functions of these loci. In this study, we used congenic mice to eliminate genetic variability, and analyzed eumelanin and pheomelanin contents of hairs from mice mutant at one or more of the major pigment loci, i.e., the albino (C) locus that encodes tyrosinase, the slaty (Slt) locus that encodes tyrosinase-related protein 2 (TRP2 also known as dopachrome tautomerase, DCT), the brown (B) locus that encodes TRP1, the silver (Si) locus that encodes a melanosomal silver protein, the agouti (A) locus that encodes agouti signaling protein (ASP), the extension (E) locus that encodes melanocortin-1 receptor, and the mahogany (Mg) locus that encodes attractin. We also measured total melanin contents after solubilization of hairs in hot Soluene-350 plus water. Hairs were shaved from 2-3-month-old congenic C57BL/6J mice. The chinchilla (c(ch)) allele is known to encode tyrosinase, whose activity is about one third that of wild type (C). Phenotypes of chinchilla (c(ch)/c(ch)) mice that are wild type or mutant at the brown and/or slaty, loci indicate that functioning TRP2 and TRP1 are necessary, in addition to high levels of tyrosinase, for a full production of eumelanin. The chinchilla allele was found to reduce the amount of pheomelanin in lethal yellow and recessive yellow mice to less than one fifth of that in congenic yellow mice that were wild type at the albino locus. This indicates that reduction in tyrosinase activity affects pheomelanogenesis more profoundly compared with eumelanogenesis. Hairs homozygous for mutation at the slaty locus contain 5,6-dihydroxyindole-2-carboxylic acid (DHICA)-poor melanin, and this chemical phenotype was retained in hairs that were mutant at both the brown locus and the slaty locus. Hair from mice mutant at the brown locus, but not at the slaty locus, do not contain DHICA-poor melanin. This indicates that the proportion of DHICA in eumelanin is determined by TRP2, but not by TRP1. Mutation at the slaty locus (Slt(lt)) was found to have no effect on pheomelanogenesis, supporting a role of TRP2 only in eumelanogenesis. The mutation at silver (si) locus showed an effect similar to brown, a partial suppression of eumelanogenesis. The mutation at mahogany (mg) locus partially suppressed the effect of lethal yellow (Ay) on pheomelanogenesis, supporting a role of mahogany in interfering with agouti signaling. These results show that combination of double mutation study of congenic mice with chemical analysis of melanins is useful in evaluating the interaction of pigment gene functions.

OBJECTIVE

ALR/Lt, a mouse strain with strong resistance to type 1 diabetes, is closely related to autoimmune type 1 diabetes-prone NOD/Lt mice. ALR pancreatic beta cells are resistant to the beta cell toxin alloxan, combinations of cytotoxic cytokines, and diabetogenic NOD T-cell lines. Reciprocal F1 hybrids between either ALR and NOD or ALR and NON/Lt, showed that alloxan resistance was transmitted to F1 progeny only when ALR was the maternal parent. Here we show that the mitochondrial genome (mtDNA) of ALR mice contributes resistance to diabetes.

METHODS

When F1 progeny from reciprocal outcrosses between ALR and NOD were backcrossed to NOD, a four-fold lower frequency of spontaneous type 1 diabetes development occurred when ALR contributed the mtDNA. Because of the apparent interaction between nuclear and mtDNA, the mitochondrial genomes were sequenced.

RESULTS

An ALR-specific sequence variation in the mt-Nd2 gene producing a leucine to methionine substitution at amino acid residue 276 in the NADH dehydrogenase 2 was discovered. An isoleucine to valine mutation in the mt-Co3 gene encoding COX3 distinguished ALR and NOD from NON and ALS. All four strains were distinguished by variation in a mt-encoded arginyl tRNA polyadenine tract. Shared alleles of mt-Co3 and mt-Tr comparing NOD and ALR allowed for exclusion of these two genes as candidates, implicating the mt-Nd2 variation as a potential ALR-derived type 1 diabetes protective gene.

CONCLUSIONS

The unusual resistance of ALR mice to both ROS-mediated and autoimmune type 1 diabete stresses reflects an interaction between the nuclear and mt genomes. The latter contribution is most likely via a single nucleotide polymorphism in mt-Nd2.

Enterotoxigenic Escherichia coli (ETEC) produces both heat-labile (LT) and heat-stable (ST) enterotoxins and is a major cause of diarrhea in infants in developing countries and in travelers to those regions. In addition to inducing fluid secretion, LT is a powerful mucosal adjuvant capable of promoting immune responses to coadministered antigens. In this study, we examined purified A subunit to further understand the toxicity and adjuvanticity of LT. Purified A subunit was enzymatically active but sensitive to proteolytic degradation and unable to bind gangliosides, and even in the presence of admixed B subunit, it displayed low cyclic AMP (cAMP) induction and no enterotoxicity. Thus, the ABLT adjuvanticity is not dependent on the ABB subunit. However, immune responses were maximal when signals were received from both subunits either in an ABB admixed. Furthermore, the quality of the immune response (i.e., IgG1/IgG2 balance and mucosal IgA and IL-17 secretion) was determined by the presence of an A subunit, revealing for the first time induction of Th17 responses with the A subunit alone. These results have important implications for understanding ETEC pathogenesis, unraveling immunologic responses induced by LT-based adjuvants, and developing new mucosal vaccines.

<A<em>b</em>stractText>To compare re-rupture rate, complication rate, and functional outcome after operative versus nonoperative treatment of Achilles tendon ruptures; to compare re-rupture rate after early and late full weight <em>b</em>earing; to evaluate re-rupture rate after functional reha<em>b</em>ilitation with early range of motion; and to compare effect estimates from randomised controlled trials and o<em>b</em>servational studies.</A<em>b</em>stractText><A<em>b</em>stractText>Systematic review and meta-analysis.</A<em>b</em>stractText><A<em>b</em>stractText>Pu<em>b</em>Med/Medline, Em<em>b</em>ase, CENTRAL, and CINAHL data<em>b</em>ases were last searched on 25 April 2018 for studies comparing operative versus nonoperative treatment of Achilles tendon ruptures.</A<em>b</em>stractText><A<em>b</em>stractText>Randomised controlled trials and o<em>b</em>servational studies reporting on comparison of operative versus nonoperative treatment of acute Achilles tendon ruptures.</A<em>b</em>stractText><A<em>b</em>stractText>Data extraction was performed independently in pairs, <em>b</em>y four reviewers, with the use of a predefined data extraction file. Outcomes were pooled using random effects models and presented as risk difference, risk ratio, or mean difference, with 95% confidence interval.</A<em>b</em>stractText><p><div>(<em>b</em>)RESULTS</<em>b</em>)</div>29 studies were included-10 randomised controlled trials and 19 o<em>b</em>servational studies. The 10 trials included 944 (6%) patients, and the 19 o<em>b</em>servational studies included 14 918 (94%) patients. A significant reduction in re-ruptures was seen after operative treatment (2.3%) compared with nonoperative treatment (3.9%) (risk difference 1.6%; risk ratio 0.43, 95% confidence interval 0.31 to 0.60; P&<em>lt</em>;0.001; I²=22%). Operative treatment resu<em>lt</em>ed in a significantly higher complication rate than nonoperative treatment (4.9% <i>v</i> 1.6%; risk difference 3.3%; risk ratio 2.76, 1.84 to 4.13; P&<em>lt</em>;0.001; I²=45%). The main difference in complication rate was attri<em>b</em>uta<em>b</em>le to the incidence of infection (2.8%) in the operative group. A similar reduction in re-rupture rate in favour of operative treatment was seen after <em>b</em>oth early and late full weight <em>b</em>earing. No significant difference in re-rupture rate was seen <em>b</em>etween operative and nonoperative treatment in studies that used accelerated functional reha<em>b</em>ilitation with early range of motion (risk ratio 0.60, 0.26 to 1.37; P=0.23; I²=0%). No difference in effect estimates was seen <em>b</em>etween randomised controlled trials and o<em>b</em>servational studies.</p><A<em>b</em>stractText>This meta-analysis shows that operative treatment of Achilles tendon ruptures reduces the risk of re-rupture compared with nonoperative treatment. However, re-rupture rates are low and differences <em>b</em>etween treatment groups are small (risk difference 1.6%). Operative treatment resu<em>lt</em>s in a higher risk of other complications (risk difference 3.3%). The final decision on the management of acute Achilles tendon ruptures should <em>b</em>e <em>b</em>ased on patient specific factors and shared decision making. This review emphasises the potential <em>b</em>enefits of adding high quality o<em>b</em>servational studies in meta-analyses for the evaluation of o<em>b</em>jective outcome measures after surgical treatment.</A<em>b</em>stractText>

(<em>b</em>)Background:</<em>b</em>) The rapidly evolving coronavirus disease 2019 (COVID-19), was declared a pandemic <em>b</em>y the World Hea<em>lt</em>h Organization on March 11, 2020. It was first detected in the Wuhan city of China and has spread glo<em>b</em>ally resu<em>lt</em>ing in a su<em>b</em>stantial hea<em>lt</em>h and economic crisis in many countries. O<em>b</em>servational studies have partially identified different aspects of this disease. There have <em>b</em>een no pu<em>b</em>lished systematic reviews that com<em>b</em>ine clinical, la<em>b</em>oratory, epidemiologic, and mortality findings. Also, the effect of gender on the outcomes of COVID-19 has not <em>b</em>een well-defined. (<em>b</em>)Methods:</<em>b</em>) We reviewed the scientific literature pu<em>b</em>lished from January 1, 2019 to May 29, 2020. Statistical analyses were performed with STATA (version 14, IC; Stata Corporation, College Station, TX, USA). The pooled frequency with 95% confidence intervals (CI) was assessed using random effect model. <i>P</i> &<em>lt</em>; 0.05 was considered a statistically significant pu<em>b</em>lication <em>b</em>ias. (<em>b</em>)Resu<em>lt</em>s:</<em>b</em>) Out of 1,223 studies, 34 satisfied the inclusion criteria. A total of 5,057 patients with a mean age of 49 years were evaluated. Fever (83.0%, CI 77.5-87.6) and cough (65.2%, CI 58.6-71.2) were the most common symptoms. The most prevalent comor<em>b</em>idities were hypertension (18.5%, CI 12.7-24.4) and Cardiovascular disease (14.9%, CI 6.0-23.8). Among the la<em>b</em>oratory a<em>b</em>normalities, elevated C-Reactive Protein (CRP) (72.0%, CI 54.3-84.6) and lymphopenia (50.1%, CI 38.0-62.4) were the most common. Bilateral ground-glass opacities (66.0%, CI 51.1-78.0) was the most common CT scan presentation. The pooled mortality rate was 6.6%, with males having significantly higher mortality compared to females (OR 3.4; 95% CI 1.2-9.1, <i>P</i> = 0.01). (<em>b</em>)Conclusion:</<em>b</em>) COVID-19 has caused a significant num<em>b</em>er of hospitalization and mortality worldwide. Mortality associated with COVID-19 was higher in our study compared to the previous reports from China. The mortality was significantly higher among the hospitalized male group. Further studies are required to evaluate the effect of different varia<em>b</em>les resu<em>lt</em>ing in sex disparity in COVID-19 mortality.

Keywords: COVID-19; coronavirus; male; mortality; pandemic.

Vaccination of healthcare workers (HCWs) reduces the risk of occupational infections, prevents nosocomial transmission and maintains healthcare delivery during outbreaks. Despite the European directive and national legislation on workers' protection, immunization coverage among HCWs has often been very low. In light of Italian National Vaccination Plan 2012-2014 recommendations, the aim of this study was to assess levels of immunization and factors influencing adherence to vaccinations needed for HCWs in Puglia region, South Italy. The study was conducted using an interview-based standardized anonymous questionnaire administered to hospital employees in the period November 2009-March 2011. A total of 2198 health professionals responded in 51/69 Apulian hospitals (median age: 45 years; 65.2% nurses, 22.6% doctors and 12.2% other hospital personnel). Vaccination coverage was 24.8% for influenza, 70.1% for hepatitis B, 9.7% for MMR, 3.6% for varicella, and 15.5% for Td booster. Receiving counselling from occupational health physicians (OHPs) was associated with influenza (OR = 1.8; 95%CI = 1.5-2.2; P &lt; 0.001), hepatitis B (OR = 4.9; 95%CI = 3.9-6.3; P &lt; 0.001), varicella (OR = 43.7; 95%CI = 18.9-101.7; P &lt; 0.001), MMR (OR = 8.8; 95%CI = 4.1-18.6; P &lt; 0.001) and tetanus (OR = 50.5; 95%CI = 30.1-88.3; P &lt; 0.001) vaccine uptake. OHPs should be trained with standard guidelines specific for healthcare settings and HCWs' risk groups to facilitate their crucial role in improving vaccine coverage among HCWs and increase awareness on the duty to protect both employees and patients.

(<em>b</em>)Purpose:</<em>b</em>) The aim of this retrospective mu<em>lt</em>icentric study was to develop and evaluate a prognostic FDG PET/CT radiomics signature in early-stage non-small cell lung cancer (NSCLC) patients treated with stereotactic radiotherapy (SBRT). (<em>b</em>)Material and Methods:</<em>b</em>) Patients from 3 different centers (<i>n</i> = 27, 29 and 8) were pooled to constitute the training set, whereas the patients from a fourth center (<i>n</i> = 23) were used as the testing set. The primary endpoint was local control (LC). The primary tumour was semi-automatically delineated in the PET images using the Fuzzy locally adaptive Bayesian algorithm, and manually in the low-dose CT images. A total of 184 IBSI-compliant radiomic features were extracted. Seven clinical and treatment parameters were included. We used ComBat to harmonize radiomic features extracted from the four institutions relying on different PET/CT scanners. In the training set, varia<em>b</em>les found significant in the univariate analysis were fed into a mu<em>lt</em>ivariate regression model and models were <em>b</em>ui<em>lt</em> <em>b</em>y com<em>b</em>ining independent prognostic factors. (<em>b</em>)Resu<em>lt</em>s:</<em>b</em>) Median follow-up was 21.1 (1.7 - 63.4) and 25.5 (7.7 - 57.8) months in training and testing sets respectively. In univariate analysis, none of the clinical varia<em>b</em>les, 2 PET and 2 CT features were significantly predictive of LC. The <em>b</em>est predictive models in the training set were o<em>b</em>tained <em>b</em>y com<em>b</em>ining one feature from PET, namely information correlation 2 (IC2) and one from CT (Flatness), reaching a sensitivity of 100% and a specificity of 96%. Another model com<em>b</em>ining 2 PET features (IC2 and Strength), reached sensitivity of 100% and specificity of 88%, <em>b</em>oth with an undefined hazard ratio (HR) (p&<em>lt</em>;0.001). The latter model o<em>b</em>tained an accuracy of 0.91 (sensitivity 100%, specificity 81%), with a HR undefined (<i>P</i> = 0.023) in the testing set, however other models relying on CT radiomics features only or the com<em>b</em>ination of PET and CT features failed to validate in the testing set. (<em>b</em>)Conclusion:</<em>b</em>) We showed that two radiomic features derived from FDG PET were independently associated with LC in patients with NSCLC undergoing SBRT and could <em>b</em>e com<em>b</em>ined in an accurate predictive model. This model could provide local relapse-related information and could <em>b</em>e helpful in clinical decision-making.

The ability of microorganisms to generate resistance outcompetes with the generation of new and efficient antibiotics; therefore, it is critical to develop novel antibiotic agents and treatments to control bacterial infections. An alternative to this worldwide problem is the use of nanomaterials with antimicrobial properties. Silver nanoparticles (AgNPs) have been extensively studied due to their antimicrobial effect in different organisms. In this work, the synergistic antimicrobial effect of AgNPs and conventional antibiotics was assessed in Gram-positive and Gram-negative bacteria. AgNPs minimal inhibitory concentration was 10-12 μg mL-1 in all bacterial strains tested, regardless of their different susceptibility against antibiotics. Interestingly, a synergistic antimicrobial effect was observed when combining AgNPs and kanamycin according to the fractional inhibitory concentration index, FICI: &lt;0.5), an additive effect by combining AgNPs and chloramphenicol (FICI: 0.5 to 1), whereas no effect was found with AgNPs and β-lactam antibiotics combinations. Flow cytometry and TEM analysis showed that sublethal concentrations of AgNPs (6-7 μg mL-1) altered the bacterial membrane potential and caused ultrastructural damage, increasing the cell membrane permeability. No chemical interactions between AgNPs and antibiotics were detected. We propose an experimental supported mechanism of action by which combinatorial effect of antimicrobials drives synergy depending on their specific target, facilitated by membrane alterations generated by AgNPs. Our results provide a deeper understanding about the synergistic mechanism of AgNPs and antibiotics, aiming to combat antimicrobial infections efficiently, especially those by multi-drug resistant microorganisms, in order to mitigate the current crisis due to antibiotic resistance.

OBJECTIVE

Direct-acting antivirals (DAAs) have dramatically improved the outcome of patients with hepatitis C virus (HCV) infection including those with decompensated cirrhosis (DC). We analyzed the evolution of indications and results of liver transplantation (LT) in the past 10 years in Europe, focusing on the changes induced by the advent of DAAs.

METHODS

This is a cohort study based on data from the European Liver Transplant Registry (ELTR). Data of adult LTs performed between January 2007 to June 2017 for HCV, hepatitis B virus (HBV), alcohol (EtOH) and non-alcoholic steatohepatitis (NASH) were analyzed. The period was divided into different eras: interferon (IFN/RBV; 2007-2010), protease inhibitor (PI; 2011-2013) and second generation DAA (DAA; 2014-June 2017).

RESULTS

Out of a total number of 60,527 LTs, 36,382 were performed in patients with HCV, HBV, EtOH and NASH. The percentage of LTs due to HCV-related liver disease varied significantly over time (p <0.0001), decreasing from 22.8% in the IFN/RBV era to 17.4% in the DAA era, while those performed for NASH increased significantly (p <0.0001). In the DAA era, the percentage of LTs for HCV decreased significantly (p <0.0001) from 21.1% (first semester 2014) to 10.6% (first semester 2017). This decline was more evident in patients with DC (HCV-DC, -58.0%) than in those with hepatocellular carcinoma (HCC) associated with HCV (HCV-HCC, -41.2%). Conversely, three-year survival of LT recipients with HCV-related liver disease improved from 65.1% in the IFN/RBV era to 76.9% in the DAA era, and is now comparable to the survival of recipients with HBV infection (p = 0.3807).

CONCLUSIONS

In Europe, the number of LTs due to HCV infection is rapidly declining for both HCV-DC and HCV-HCC indications and post-LT survival has dramatically improved over the last three years. This is the first comprehensive study of the overall impact of DAA treatment for HCV on liver transplantation in Europe.

UNASSIGNED

After the advent of direct-acting antivirals in 2014, a dramatic decline was observed in the number of liver transplants performed both in patients with decompensated cirrhosis due to hepatitis C virus (HCV), minus 60%, and in those with hepatocellular carcinoma associated with HCV, minus 41%. Furthermore, this is the first large-scale study demonstrating that the survival of liver transplant recipients with HCV-related liver disease has dramatically improved over the last three years and is now comparable to the survival of recipients with hepatitis B virus infection. The reduction in HCV-related indications for LT means that there is a greater availability of livers, at least 600 every year, which can be allocated to patients with indications other than HCV.

<A<em>b</em>stractText>Cardiovascular adverse events (CVAEs) can occur during proteasome inhi<em>b</em>itor (PI) therapy. We conducted a prospective, o<em>b</em>servational, mu<em>lt</em>i-institutional study to define risk factors and outcomes in patients with mu<em>lt</em>iple myeloma (MM) receiving PIs.</A<em>b</em>stractText><A<em>b</em>stractText>Patients with relapsed MM initiating carfilzomi<em>b</em>- or <em>b</em>ortezomi<em>b</em>-<em>b</em>ased therapy underwent <em>b</em>aseline assessments and repeated assessments at regular intervals over 6 months, including cardiac <em>b</em>iomarkers (troponin I or T, <em>b</em>rain natriuretic peptide [BNP], and N-terminal proBNP), ECG, and echocardiography. Monitoring occurred over 18 months for development of CVAEs.</A<em>b</em>stractText><p><div>(<em>b</em>)RESULTS</<em>b</em>)</div>Of 95 patients enrolled, 65 received carfilzomi<em>b</em> and 30 received <em>b</em>ortezomi<em>b</em>, with median 25 months of follow-up. Sixty-four CVAEs occurred, with 55% grade 3 or greater in severity. CVAEs occurred in 51% of patients treated with carfilzomi<em>b</em> and 17% of those treated with <em>b</em>ortezomi<em>b</em> (<i>P</i> = .002). Median time to first CVAE from treatment start was 31 days, and 86% occurred within the first 3 months. Patients receiving carfilzomi<em>b</em>-<em>b</em>ased therapy with a <em>b</em>aseline elevated BNP level higher than 100 pg/mL or N-terminal proBNP level higher than 125 pg/mL had increased risk for CVAE (odds ratio, 10.8; <i>P</i> &<em>lt</em>; .001). Elevated natriuretic peptides occurring mid-first cycle of treatment with carfilzomi<em>b</em> were associated with a su<em>b</em>stantially higher risk of CVAEs (odds ratio, 36.0; <i>P</i> &<em>lt</em>; .001). Patients who experienced a CVAE had inferior progression-free survival (log-rank <i>P</i> = .01) and overall survival (log-rank <i>P</i> &<em>lt</em>; .001). PI therapy was safely resumed in 89% of patients, a<em>lt</em>hough 41% required chemotherapy modifications.</p><A<em>b</em>stractText>CVAEs are common during PI therapy for relapsed MM, especially with carfilzomi<em>b</em>, particularly within the first 3 months of therapy. CVAEs were associated with worse overall outcomes, <em>b</em>ut usually, discontinuation of therapy was not required. Natriuretic peptides were highly predictive of CVAEs; however, validation of this finding is necessary <em>b</em>efore uniform incorporation into the routine management of patients receiving carfilzomi<em>b</em>.</A<em>b</em>stractText>

<A<em>b</em>stractText>After acute coronary syndrome, dia<em>b</em>etes conveys an excess risk of ischaemic cardiovascular events. A reduction in mean LDL cholesterol to 1·4-1·8 mmol/L with ezetimi<em>b</em>e or statins reduces cardiovascular events in patients with an acute coronary syndrome and dia<em>b</em>etes. However, the efficacy and safety of further reduction in LDL cholesterol with an inhi<em>b</em>itor of proprotein convertase su<em>b</em>tilisin/kexin type 9 (PCSK9) after acute coronary syndrome is unknown. We aimed to explore this issue in a prespecified analysis of the ODYSSEY OUTCOMES trial of the PCSK9 inhi<em>b</em>itor alirocuma<em>b</em>, assessing its effects on cardiovascular outcomes <em>b</em>y <em>b</em>aseline glycaemic status, while also assessing its effects on glycaemic measures including risk of new-onset dia<em>b</em>etes.</A<em>b</em>stractText><p><div>(<em>b</em>)METHODS</<em>b</em>)</div>ODYSSEY OUTCOMES was a randomised, dou<em>b</em>le-<em>b</em>lind, place<em>b</em>o-controlled trial, done at 1315 sites in 57 countries, that compared alirocuma<em>b</em> with place<em>b</em>o in patients who had <em>b</em>een admitted to hospital with an acute coronary syndrome (myocardial infarction or unsta<em>b</em>le angina) 1-12 months <em>b</em>efore randomisation and who had raised concentrations of atherogenic lipoproteins despite use of high-intensity statins. Patients were randomly assigned (1:1) to receive alirocuma<em>b</em> or place<em>b</em>o every 2 weeks; randomisation was stratified <em>b</em>y country and was done centrally with an interactive voice-response or we<em>b</em>-response system. Alirocuma<em>b</em> was titrated to target LDL cholesterol concentrations of 0·65-1·30 mmol/L. In this prespecified analysis, we investigated the effect of alirocuma<em>b</em> on cardiovascular events <em>b</em>y glycaemic status at <em>b</em>aseline (dia<em>b</em>etes, predia<em>b</em>etes, or normoglycaemia)-defined on the <em>b</em>asis of patient history, review of medical records, or <em>b</em>aseline H<em>b</em>A<su<em>b</em>)1c</su<em>b</em>) or fasting serum glucose-and risk of new-onset dia<em>b</em>etes among those without dia<em>b</em>etes at <em>b</em>aseline. The primary endpoint was a composite of death from coronary heart disease, non-fatal myocardial infarction, fatal or non-fatal ischaemic stroke, or unsta<em>b</em>le angina requiring hospital admission. ODYSSEY OUTCOMES is registered with ClinicalTrials.gov, num<em>b</em>er NCT01663402.</p><p><div>(<em>b</em>)FINDINGS</<em>b</em>)</div>At study <em>b</em>aseline, 5444 patients (28·8%) had dia<em>b</em>etes, 8246 (43·6%) had predia<em>b</em>etes, and 5234 (27·7%) had normoglycaemia. There were no significant differences across glycaemic categories in median LDL cholesterol at <em>b</em>aseline (2·20-2·28 mmol/L), after 4 months' treatment with alirocuma<em>b</em> (0·80 mmol/L), or after 4 months' treatment with place<em>b</em>o (2·25-2·28 mmol/L). In the place<em>b</em>o group, the incidence of the primary endpoint over a median of 2·8 years was greater in patients with dia<em>b</em>etes (16·4%) than in those with predia<em>b</em>etes (9·2%) or normoglycaemia (8·5%); hazard ratio (HR) for dia<em>b</em>etes versus normoglycaemia 2·09 (95% CI 1·78-2·46, p&<em>lt</em>;0·0001) and for dia<em>b</em>etes versus predia<em>b</em>etes 1·90 (1·65-2·17, p&<em>lt</em>;0·0001). Alirocuma<em>b</em> resu<em>lt</em>ed in similar relative reductions in the incidence of the primary endpoint in each glycaemic category, <em>b</em>ut a greater a<em>b</em>solute reduction in the incidence of the primary endpoint in patients with dia<em>b</em>etes (2·3%, 95% CI 0·4 to 4·2) than in those with predia<em>b</em>etes (1·2%, 0·0 to 2·4) or normoglycaemia (1·2%, -0·3 to 2·7; a<em>b</em>solute risk reduction p<su<em>b</em>)interaction</su<em>b</em>)=0·0019). Among patients without dia<em>b</em>etes at <em>b</em>aseline, 676 (10·1%) developed dia<em>b</em>etes in the place<em>b</em>o group, compared with 648 (9·6%) in the alirocuma<em>b</em> group; alirocuma<em>b</em> did not increase the risk of new-onset dia<em>b</em>etes (HR 1·00, 95% CI 0·89-1·11). HRs were 0·97 (95% CI 0·87-1·09) for patients with predia<em>b</em>etes and 1·30 (95% CI 0·93-1·81) for those with normoglycaemia (p<su<em>b</em>)interaction</su<em>b</em>)=0·11).</p><A<em>b</em>stractText>After a recent acute coronary syndrome, alirocuma<em>b</em> treatment targeting an LDL cholesterol concentration of 0·65-1·30 mmol/L produced a<em>b</em>out twice the a<em>b</em>solute reduction in cardiovascular events among patients with dia<em>b</em>etes as in those without dia<em>b</em>etes. Alirocuma<em>b</em> treatment did not increase the risk of new-onset dia<em>b</em>etes.</A<em>b</em>stractText><A<em>b</em>stractText>Sanofi and Regeneron Pharmaceuticals.</A<em>b</em>stractText>

<A<em>b</em>stractText>Immune checkpoint inhi<em>b</em>itor use in oncology is increasing rapidly. We sought to determine the frequency, severity, cause, and predictors of AKI in a real-world population receiving checkpoint inhi<em>b</em>itors.</A<em>b</em>stractText><A<em>b</em>stractText>We included all patients who received checkpoint inhi<em>b</em>itor therapy from May 2011 to Decem<em>b</em>er 2016 at Massachusetts General Hospital. Baseline serum creatinine, averaged 6 months <em>b</em>efore checkpoint inhi<em>b</em>itor start date, was compared with all su<em>b</em>sequent creatinine values within 12 months of starting therapy. AKI was defined <em>b</em>y Kidney Disease: Improving Glo<em>b</em>al Outcomes criteria for fold changes in creatinine from <em>b</em>aseline. Sustained AKI events lasted at least 3 days and was our primary outcome. The cause of sustained AKI was determined <em>b</em>y chart review. Cumulative incidence and su<em>b</em>distri<em>b</em>ution hazard models were used to assess the relationship <em>b</em>etween <em>b</em>aseline demographics, comor<em>b</em>idities, and medications, and sustained AKI and potential checkpoint inhi<em>b</em>itor-related AKI.</A<em>b</em>stractText><p><div>(<em>b</em>)RESULTS</<em>b</em>)</div>We included 1016 patients in the analysis. Average age was 63 (SD 13) years, 61% were men, and 91% were white. Mean <em>b</em>aseline creatinine was 0.9 mg/dl (SD 0.4 mg/dl), and 169 (17%) had CKD (eGFR&<em>lt</em>;60 ml/min per 1.73 m²) at <em>b</em>aseline. A total of 169 patients (17%) experienced AKI, defined <em>b</em>y an increase in creatinine at least 1.5 times the <em>b</em>aseline within 12 months; 82 patients (8%) experienced sustained AKI and 30 patients (3%) had potential checkpoint inhi<em>b</em>itor-related AKI. The first episode of sustained AKI occurred, on average, 106 days (SD 85) after checkpoint inhi<em>b</em>itor initiation. Sixteen (2%) patients experienced stage 3 sustained AKI and four patients required dialysis. Proton pump inhi<em>b</em>itor use at <em>b</em>aseline was associated with sustained AKI.</p><A<em>b</em>stractText>AKI is common in patients receiving checkpoint inhi<em>b</em>itor therapy. The causes of sustained AKI in this population are heterogenous and merit thorough evaluation. The role of PPI and other nephritis-inducing drugs in the development of sustained AKI needs to <em>b</em>e <em>b</em>etter defined.</A<em>b</em>stractText>

(<em>b</em>)OBJECTIVE:</<em>b</em>) To investigate the association <em>b</em>etween chronic o<em>b</em>structive pulmonary disease (COPD) and smoking with outcome in patients with COVID-19.(<em>b</em>)METHODS:</<em>b</em>) A systematic literature search was performed using Pu<em>b</em>Med, EuropePMC, SCOPUS and the Cochrane Central Data<em>b</em>ase. A composite of poor outcome, mortality, severe COVID-19, the need for treatment in an intensive care unit (ICU) and disease progression were the outcomes of interest.(<em>b</em>)RESULTS:</<em>b</em>) Data on 4603 patients were pooled from 21 studies. COPD was associated with an increased risk for composite poor outcome (OR 5.01, 95%CI 3.06-8.22; <i>P</i> &<em>lt</em>; 0.001; <i>I</i>² 0%), mortality (OR 4.36, 95%CI 1.45-13.10; <i>P</i> = 0.009; <i>I</i>² 0%), severe COVID-19 (OR 4.62, 95%CI 2.49-8.56; <i>P</i> &<em>lt</em>; 0.001; <i>I</i>² 0%), ICU care (OR 8.33, 95%CI 1.27-54.56; <i>P</i> = 0.03; <i>I</i>² 0%), and disease progression (OR 8.42, 95%CI 1.60-44.27; <i>P</i> = 0.01; <i>I</i>² 0%). Smoking was found to increase the risk of composite poor outcome (OR 1.52, 95%CI 1.16-2.00; <i>P</i> = 0.005; <i>I</i>² 12%), and su<em>b</em>group analysis showed that smoking was significant for increased risk of severe COVID-19 (OR 1.65, 95%CI 1.17-2.34; <i>P</i> = 0.004; <i>I</i>² 11%). Current smokers were at higher risk of composite poor outcomes (OR 1.58, 95%CI 1.10-2.27; <i>P</i> = 0.01; <i>I</i>² 0%) than former/non-smokers.(<em>b</em>)CONCLUSION:</<em>b</em>) Our systematic review and meta-analysis revealed that COPD and smoking were associated with poor outcomes in patients with COVID-19.

Background: Information on the clinical characteristics and outcomes of hospitalized Covid-19 patients with or without diabetes mellitus (DM) is limited in the Arab region. This study aims to fill this gap.

Methods: In this single-center retrospective study, medical records of hospitalized adults with confirmed Covid-19 [RT-PCR positive for SARS-CoV2] at King Saud University Medical City (KSUMC)-King Khaled University Hospital (KKUH), Riyadh, Saudi Arabia from May to July 2020 were analyzed. Clinical, radiological and serological information, as well as outcomes were recorded and analyzed.

Results: A total of 439 patients were included (median age 55 years; 68.3% men). The most prevalent comorbidities were vitamin D deficiency (74.7%), DM (68.3%), hypertension (42.6%) and obesity (42.2%). During hospitalization, 77 out of the 439 patients (17.5%) died. DM patients have a significantly higher death rate (20.5% versus 12.3%; p = 0.04) and lower survival time (p = 0.016) than non-DM. Multivariate cox proportional hazards regression model revealed that age [Hazards ratio, HR 3.0 (95% confidence interval, CI 1.7-5.3); p &lt; 0.001], congestive heart failure [adjusted HR 3.5 (CI 1.4-8.3); p = 0.006], smoking [adjusted HR 5.8 (CI 2.0-17.2); p &lt; 0.001], β-blocker use [adjusted HR 1.7 (CI 1.0-2.9); p = 0.04], bilateral lung infiltrates [adjusted HR 1.9 (CI 1.1-3.3); p = 0.02], creatinine > 90 µmol/l [adjusted HR 2.1 (CI 1.3-3.5); p = 0.004] and 25(OH)D &lt; 12.5 nmol/l [adjusted HR 7.0 (CI 1.7-28.2); p = 0.007] were significant predictors of mortality among hospitalized Covid-19 patients. Random blood glucose ≥ 11.1 mmol/l was significantly associated with intensive care admission [adjusted HR 1.5 (CI 1.0-2.2); p = 0.04], as well as smoking, β-blocker use, neutrophil > 7.5, creatinine > 90 µmol/l and alanine aminotransferase > 65U/l.

Conclusion: The prevalence of DM is high among hospitalized Covid-19 patients in Riyadh, Saudi Arabia. While DM patients have a higher mortality rate than their non-DM counterparts, other factors such as old age, congestive heart failure, smoking, β-blocker use, presence of bilateral lung infiltrates, elevated creatinine and severe vitamin D deficiency, appear to be more significant predictors of fatal outcome. Patients with acute metabolic dysfunctions, including hyperglycemia on admission are more likely to receive intensive care.

Keywords: Covid-19; Diabetes mellitus; Mortality; Saudi Arabia.

To understand the genetics of type 2 diabetes in people of Japanese ancestry, we conducted A meta-analysis of four genome-wide association studies (GWAS; 36,614 cases and 155,150 controls of Japanese ancestry). We identified 88 type 2 diabetes-associated loci (P &lt; 5.0 × 10^-8) with 115 independent signals (P &lt; 5.0 × 10^-6), of which 28 loci with 30 signals were novel. Twenty-eight missense variants were in linkage disequilibrium (r² > 0.6) with the lead variants. Among the 28 missense variants, three previously unreported variants had distinct minor allele frequency (MAF) spectra between people of Japanese and European ancestry (MAF_JPN > 0.05 versus MAF_EUR &lt; 0.01), including missense variants in genes related to pancreatic acinar cells (GP2) and insulin secretion (GLP1R). Transethnic comparisons of the molecular pathways identified from the GWAS results highlight both ethnically shared and heterogeneous effects of a series of pathways on type 2 diabetes (for example, monogenic diabetes and beta cells).

Microbial-derived short-chain fatty acids (SCFA) acetate, propionate and butyrate may provide a link between gut microbiota and whole-body insulin sensitivity (IS). In this cross-sectional study (160 participants, 64% male, BMI: 19.2-41.0 kg/m², normal or impaired glucose metabolism), associations between SCFA (faecal and fasting circulating) and circulating metabolites, substrate oxidation and IS were investigated. In a subgroup (n = 93), IS was determined using a hyperinsulinemic-euglycemic clamp. Data were analyzed using multiple linear regression analysis adjusted for sex, age and BMI. Fasting circulating acetate, propionate and butyrate concentrations were positively associated with fasting GLP-1 concentrations. Additionally, circulating SCFA were negatively related to whole-body lipolysis (glycerol), triacylglycerols and free fatty acids levels (standardized (std) β adjusted (adj) -0.190, P = 0.023; std β adj -0.202, P = 0.010; std β adj -0.306, P = 0.001, respectively). Circulating acetate and propionate were, respectively, negatively and positively correlated with IS (M-value: std β adj -0.294, P &lt; 0.001; std β adj 0.161, P = 0.033, respectively). We show that circulating rather than faecal SCFA were associated with GLP-1 concentrations, whole-body lipolysis and peripheral IS in humans. Therefore, circulating SCFA are more directly linked to metabolic health, which indicates the need to measure circulating SCFA in human prebiotic/probiotic intervention studies as a biomarker/mediator of effects on host metabolism.

A green fluorescent protein (gfp) gene was ligated to the Lactobacillus reuteri-specific nisin-inducible expression-secretion vector pNIES, generating a pNIES-GFP vector capable of secreting the cloned gene as a GFP-fusion protein with fluorescent activity. To develop this system as a live vehicle carrying the heat-stable enterotoxin (ST) and heat-labile enterotoxin B (LT(B)) of the enterotoxigenic Escherichia coli (ETEC), a recombinant 5'-ST-LT(B)-3' DNA fragment was cloned into pNIES-GFP. The resulting L. reuteri/pNIES-GFP:STLT(B) system was found to possess the capability of adhering to the mice gut, secreting GFP:STLT(B) product at 0.14 and 0.026 pgcell(-1) under induced and noninduced conditions, respectively. Further analysis of the GFP:STLT(B) product confirmed its ganglioside-binding ability, LT(B) antigenicity and relative freedom from the ST-associated toxicity, making it suitable for use as an oral vaccine in mice. Oral inoculation of the L. reuteri/pNIES-GFP:STLT(B) culture in mice elicited significant (P<0.01) serum IgG and mucosal IgA antibodies against the STLT(B) antigen. These immunized mice were subsequently challenged with ETEC and showed full protection against the fluid influx response in the gut. This is the first report of using L. reuteri as a vaccine carrier to induce complete immunologic protection against ETEC.