Gait risk factors for disease progression differ between non-traumatic and post-traumatic knee osteoarthritis.

OBJECTIVE: To examine if relationships between knee osteoarthritis (OA) progression with knee moments and muscle activation during gait vary between patients with non-traumatic and post-traumatic knee OA. DESIGN: This longitudinal study included participants with non-traumatic (n = 17) and post-traumatic (n = 18) knee OA; the latter group had a previous anterior cruciate ligament rupture. Motion capture cameras, force plates, and surface electromyography measured knee moments and lower extremity muscle activation during gait. Cartilage volume change were determined over 2 years using magnetic resonance imaging in four regions: medial and lateral plateau and condyle. Linear regression analysis examined relationships between cartilage change with gait metrics (moments, muscle activation), group, and their interaction. RESULTS: Measures from knee adduction and rotation moments were related to lateral condyle cartilage loss in both groups, and knee adduction moment to lateral plateau cartilage loss in the non-traumatic group only [ß = -1.336, 95% confidence intervals (CI) = -2.653 to -0.019]. Generally, lower levels of stance phase muscle activation were related to greater cartilage loss. The relationship between cartilage loss in some regions with muscle activation characteristics varied between non-traumatic and post-traumatic groups including for: lateral hamstring (lateral condyle ß = 0.128, 95%CI = 0.003 to 0.253; medial plateau ß = 0.199, 95%CI = 0.059 to 0.339), rectus femoris (medial condyle ß = -0.267, 95%CI = -0.460 to -0.073), and medial hamstrings (medial plateau; ß = -0.146, 95%CI = -0.244 to -0.048). CONCLUSION: Findings indicate that gait risk factors for OA progression may vary between patients with non-traumatic and post-traumatic knee OA. These OA subtypes should be considered in studies that investigate gait metrics as risk factors for OA progression.

IL-17C amplifies epithelial inflammation in human psoriasis and atopic eczema.

BACKGROUND: Key pathogenic events of psoriasis and atopic eczema (AE) are misguided immune reactions of the skin. IL-17C is an epithelial-derived cytokine, whose impact on skin inflammation is unclear. OBJECTIVE: We sought to characterize the role of IL-17C in human ISD. METHODS: IL-17C gene and protein expression was assessed by immunohistochemistry and transcriptome analysis. Primary human keratinocytes were stimulated and expression of cytokines chemokines was determined by qRT-PCR and luminex assay. Neutrophil migration towards supernatant of stimulated keratinocytes was assessed. IL-17C was depleted using a new IL-17C-specific antibody (MOR106) in murine models of psoriasis (IL-23 injection model) and AE (MC903 model) as well as in human skin biopsies of psoriasis and AE. Effects on cell influx (mouse models) and gene expression (human explant cultures) were determined. RESULTS: Expression of IL-17C mRNA and protein was elevated in various ISD. We demonstrate that IL-17C potentiates the expression of innate cytokines, antimicrobial peptides (IL-36G, S100A7 and HBD2) and chemokines (CXCL8, CXCL10, CCL5 and VEGF) and the autocrine induction of IL-17C in keratinocytes. Cell-free supernatant of keratinocytes stimulated with IL-17C was strongly chemotactic for neutrophils, thus demonstrating a critical role for IL-17C in immune cell recruitment. IL-17C depletion significantly reduced cell numbers of T cells, neutrophils and eosinophils in murine models of psoriasis and AE and led to a significant downregulation of inflammatory mediators in human skin biopsies of psoriasis and AE ex vivo. CONCLUSION: IL-17C amplifies epithelial inflammation in Th2 and Th17 dominated skin inflammation and represents a promising target for the treatment of ISD.

A comparison of muscle activation and knee mechanics during gait between patients with non-traumatic and post-traumatic knee osteoarthritis.

OBJECTIVE: The objective was to compare muscle activation and knee mechanics during gait between participants with non-traumatic knee osteoarthritis (OA), post-traumatic knee OA, and healthy adults. DESIGN: Participants with non-traumatic knee OA (n = 22), post-traumatic knee OA (n = 19), and healthy adults (n = 22) completed gait trials for this observational, cross-sectional study. Post-traumatic OA group had a history of traumatic anterior cruciate ligament (ACL) rupture. Surface electromyography (EMG) measured activation of seven lower extremity muscles. Motion capture cameras and force plates measured motion and force data. Principal component analysis (PCA) determined waveform characteristics (principal components) from EMG, knee angle, and knee external moment waveforms. Analysis of variance (ANOVA) examined group differences in principal component scores (PC-scores). Regression analyses examined if a variable that coded for OA group could predict PC-scores after accounting for disease severity, alignment, and lateral OA. RESULTS: There was lower gastrocnemius EMG amplitudes (P < 0.01; ANOVA) in the post-traumatic OA group compared to healthy group. Non-traumatic OA group had higher vastus lateralis, vastus medialis, and rectus femoris EMG compared to post-traumatic OA group (P = 0.01 to 0.04) in regression analyses. Also, non-traumatic OA group had higher and prolonged lateral hamstring EMG compared to healthy (P = 0.03; ANOVA) and post-traumatic OA (P = 0.04; regression) groups respectively. The non-traumatic OA group had lower knee extension (P < 0.05) and medial rotation (P < 0.05) moments than post-traumatic and healthy groups. CONCLUSIONS: Muscle activation and knee mechanics differed between participants with non-traumatic and post-traumatic knee OA and healthy adults. These OA subtypes had differences in disease characteristics that may impact disease progression.

The effects of source and concentration of dietary fiber, starch, and fatty acids on the daily patterns of feed intake, rumination, and rumen pH in dairy cows.

The daily patterns of feed intake and rumination influence rumen fermentation, rumen pH, and timing of absorbed nutrients in the dairy cow, but the effects of diet composition on these patterns are not well characterized. Data from 3 previously published experiments were examined to determine the influence of dietary starch, fiber, and fatty acids (FA) on daily patterns of intake, rumination, and rumen pH. Dietary neutral detergent fiber (NDF) and starch were investigated in 2 experiments, each with duplicated 4 × 4 Latin square designs with a 2 × 2 factorial arrangement of treatments in cows fed cows 1×/d at 1200 and 1400 h, respectively. To investigate fiber content and digestibility in the first experiment, brown midrib or isogenic conventional corn silage were fed in low- and high-NDF diets (29 and 38%, respectively). To investigate starch source and concentration in the second experiment, ground high-moisture corn or dry ground corn were fed in low- and high-starch diets (21 and 32%, respectively). Effect of fat concentration and saturation was investigated in the third experiment using a replicated 4 × 4 Latin square design that fed cows 1×/d at 0900 h; treatments included a control diet with no added fat and 2.5% added saturated FA, unsaturated FA, or a mixture of the saturated and unsaturated FA. In the first 2 experiments, intake followed a similar daily pattern regardless of starch and NDF concentration or digestibility. Rumination displayed a treatment by time interaction for both NDF and starch concentration, with high-fiber, low-starch diets causing greater rumination overnight but not midday. High-starch diets decreased total daily rumen pH equally across the day, but did not change the daily pattern. Type of corn silage did not affect the daily patterns of rumination or rumen pH, but pH was reduced throughout the day in brown midrib diets. In the third experiment, no interactions between fatty acid supplement and time of day were observed for intake, rumination, or rumen pH. Within all experiments, rumination fit or tended to fit a 24-h rhythm regardless of diet, with the amplitude of the rumination being reduced in low-starch diets and diets containing saturated FA or a mixture of saturated and unsaturated FA. Overall, intake, rumination, and rumen pH follow a daily pattern that was minimally modified by dietary fiber and starch type and level or fat level and fatty acid profile.

Safety and efficacy of direct-acting antivirals for the treatment of chronic hepatitis C in a real-world population aged 65 years and older.

The availability of direct-acting antiviral agents (DAA) regimens has expanded the pool of patients eligible for treatment. However, data on the virologic response and tolerability of DAAs in elderly patients are lacking. We evaluated the efficacy and safety of DAAs in patients with advanced fibrosis/cirrhosis in real-life practice with the focus on those aged ≥65 years. Between January and December 2015, all consecutive patients with HCV-related advanced fibrosis/cirrhosis treated with DAA at eleven tertiary referral centres in Emilia Romagna (Italy) were enrolled. Regimen choice was based on viral genotype and stage of disease, according to guidelines. The primary end point was sustained virologic response 12 weeks after the end of treatment (SVR12). Overall, 282 of 556 (50.7%) patients evaluated were elderly, most of them with cirrhosis. Antiviral therapy was stopped prematurely in four (1.4%) patients. Two patients, both with cirrhosis, died during treatment due to worsening of liver/renal function. SVR12 was achieved by 94.7% and was comparable to that obtained in patients aged <65 (P=.074). Similar data were also reported in subgroup of patients aged ≥75 years. All patients with advanced fibrosis achieved virologic response. SVR12 was 80.8% in Child-Pugh-Turcotte (CTP)-B cirrhosis and 95.4% in CTP-A (P=.013). According to genotype, the SVR12 was achieved in 172 of 181 (95%) with genotype 1b cirrhosis and in 44 of 48 (91.7%) with genotype 2 cirrhosis. In conclusions, in a real-world setting, DAAs are safe and effective in elderly patients with HCV-related advanced fibrosis/cirrhosis, but SVR12 is lower with worsening CTP class.

Long-term outcomes of direct acting antivirals in post-transplant advanced hepatitis C virus recurrence and fibrosing cholestatic hepatitis.

Long-term functional outcomes of sofosbuvir-based antiviral treatment were evaluated in a cohort study involving 16 Italian centres within the international compassionate use programme for post-transplant hepatitis C virus (HCV) recurrence. Seventy-three patients with cirrhosis (n=52) or fibrosing cholestatic hepatitis (FCH, n=21) received 24-week sofosbuvir with ribavirin±pegylated interferon or interferon-free sofosbuvir-based regimen with daclatasvir/simeprevir+ribavirin. The patients were observed for a median time of 103 (82-112) weeks. Twelve of 73 (16.4%) died (10 non-FCH, 2 FCH) and two underwent re-LT. Sustained virological response was achieved in 46 of 66 (69.7%): 31 of 47 (66%) non-FCH and 15 of 19 (79%) FCH patients. All relapsers were successfully retreated. Comparing the data of baseline with last follow-up, MELD and Child-Turcotte-Pugh scores improved both in non-FCH (15.3±6.5 vs 10.5±3.8, P<.0001 and 8.4±2.1 vs 5.7±1.3, P<.0001, respectively) and FCH (17.3±5.9 vs 10.1±2.8, P=.001 and 8.2±1.6 vs 5.5±1, P=.001, respectively). Short-treatment mortality was higher in patients with baseline MELD≥25 than in those with MELD<25 (42.9% vs 4.8%, P=.011). Long-term mortality was 53.3% among patients with baseline MELD≥20 and 7.5% among those with MELD<20 (P<.0001). Among deceased patients 75% were Child-Turcotte-Pugh class C at baseline, while among survivors 83.9% were class A or B (P<.0001). Direct acting antivirals-based treatments for severe post-transplant hepatitis C recurrence, comprising fibrosing cholestatic hepatitis, significantly improve liver function, even without viral clearance and permit an excellent long-term survival. The setting of severe HCV recurrence may require the identification of "too-sick-to-treat patients" to avoid futile treatments.

Quality control and gap-filling of PM10 daily mean concentrations with the best linear unbiased estimator.

According to the European Directive 2008/50/CE, the air quality assessment consists in the measurement of the concentration fields, and the evaluation of the mean, number of exceedances, etc. of some chemical species dangerous to human health. The measurements provided by an air quality ground-based monitoring network are the main information source but the availability of these data is often limited by several technical and operational problems. In this paper, the best linear unbiased estimator (BLUE) is proposed to validate the pollutant concentration values and to fill the gaps in the measurement of time series collected by a monitoring network. The BLUE algorithm is tested using the daily mean concentrations of particulate matter having aerodynamic diameter less than 10 µ (PM10 concentrations) measured by the air quality monitoring sensors operating in the Lazio Region in Italy. The comparison between the estimated and measured data evidences an error comparable with the measurement uncertainty. Due to its simplicity and reliability, the BLUE will be used in the routine quality test procedures of the Lazio air quality monitoring network measurements.

Characterizing structural neural networks in de novo Parkinson disease patients using diffusion tensor imaging.

Parkinson disease (PD) can be considered as a brain multisystemic disease arising from dysfunction in several neural networks. The principal aim of this study was to assess whether large-scale structural topological network changes are detectable in PD patients who have not been exposed yet to dopaminergic therapy (de novo patients). Twenty-one drug-naïve PD patients and thirty healthy controls underwent a 3T structural MRI. Next, Diffusion Tensor Imaging (DTI) and graph theoretic analyses to compute individual structural white-matter (WM) networks were combined. Centrality (degree, eigenvector centrality), segregation (clustering coefficient), and integration measures (efficiency, path length) were assessed in subject-specific structural networks. Moreover, Network-based statistic (NBS) was used to identify whether and which subnetworks were significantly different between PD and control participants. De novo PD patients showed decreased clustering coefficient and strength in specific brain regions such as putamen, pallidum, amygdala, and olfactory cortex compared with healthy controls. Moreover, NBS analyses demonstrated that two specific subnetworks of reduced connectivity characterized the WM structural organization of PD patients. In particular, several key pathways in the limbic system, basal ganglia, and sensorimotor circuits showed reduced patterns of communications when comparing PD patients to controls. This study shows that PD is characterized by a disruption in the structural connectivity of several motor and non-motor regions. These findings provide support to the presence of disconnectivity mechanisms in motor (basal ganglia) as well as in non-motor (e.g., limbic, olfactory) circuits at an early disease stage of PD. Hum Brain Mapp 37:4500-4510, 2016. © 2016 Wiley Periodicals, Inc.

Validation and in vivo assessment of an innovative satellite-based solar UV dosimeter for a mobile app dedicated to skin health.

We present an innovative satellite-based solar UV (ultraviolet) radiation dosimeter with a mobile app interface that has been validated by exploiting both ground-based measurements and an in vivo assessment of the erythemal effects on some volunteers having controlled exposure to solar radiation. The app with this satellite-based UV dosimeter also includes other related functionalities such as the provision of safe sun exposure time updated in real-time and end exposure visual/sound alert. Both validations showed that the system has a good accuracy and reliability needed for health-related applications. This app will be launched on the market by siHealth Ltd in May 2016 under the name of "HappySun" and is available for both Android and iOS devices (more info on ). Extensive R&D activities are on-going for the further improvement of the satellite-based UV dosimeter's accuracy.

Antifungal prophylaxis in liver transplant recipients: one size does not fit all.

BACKGROUND: Targeted antifungal prophylaxis against Candida species or against Candida species and Aspergillus species, according to individual patient risk factors (RFs), is recommended by experts. However, recent studies have reported fluconazole is as effective as broader spectrum antifungals for preventing invasive fungal infection (IFI) after liver transplantation (LT). METHODS: We performed a retrospective cohort study of all adult patients who underwent LT at our 1420-bed tertiary teaching hospital, from June 2010 to December 2014, to assess the rate and etiology of IFI within 100 days after LT, to investigate the compliance with targeted prophylaxis, and to analyze risk factors for developing IFI. RESULTS: In total, 303 patients underwent LT. Patients were classified as having low (no RFs), intermediate (1 RF for invasive candidiasis [IC]), and high risk (1 RF for invasive aspergillosis [IA] or ≥2 RFs for IC) for IFI in 20%, 30%, and 50% of cases, respectively. A total of 139 patients received antifungal prophylaxis: 98 with a mold-active drug and 41 with fluconazole. Overall adherence to targeted prophylaxis was 53%. Nineteen patients (6.3%) developed IFI: 7 IC and 12 IA. Multivariate Cox regression analysis, adjusted for median model for end-stage liver disease score at LT, stratification risk group, and adherence to targeted prophylaxis, showed that graft dysfunction, renal replacement therapy, and prophylaxis with fluconazole were independent risk factors for IFI. Seven of the 9 patients who received fluconazole prophylaxis and developed IFI were classified as having high risk for IFI, and 6 developed IA. CONCLUSION: Recommended stratification is accurate for predicting patients at very high risk for IFI, who should receive prophylaxis with a mold-active drug.

Risk factors for infection with carbapenem-resistant Klebsiella pneumoniae after liver transplantation: the importance of pre- and posttransplant colonization.

Improved understanding of risk factors associated with carbapenem-resistant-Klebsiella pneumoniae (CR-KP) infection after liver transplantation (LT) can aid development of effective preventive strategies. We performed a prospective cohort study of all adult patients undergoing LT at our hospital during 30-month period to define risk factors associated with CR-KP infection. All patients were screened for CR-KP carriage by rectal swabs before and after LT. No therapy was administered to decolonize or treat asymptomatic CR-KP carriers. All patients were monitored up to 180 days after LT. Of 237 transplant patients screened, 41 were identified as CR-KP carriers (11 at LT, 30 after LT), and 20 developed CR-KP infection (18 bloodstream-infection, 2 pneumonia) a median of 41.5 days after LT. CR-KP infection rates among patients non-colonized, colonized at LT, and colonized after LT were 2%, 18.2% and 46.7% (p < 0.001). Independent risk factors for CR-KP infection identified by multivariate analysis, included: renal-replacement-therapy; mechanical ventilation > 48 h; HCV recurrence, and colonization at any time with CR-KP. Based on these four variables, we developed a risk score that effectively discriminated patients at low versus higher risk for CR-KP infection (AUC 0.93, 95% CI 0.86-1.00, p < 0.001). Our results may help to design preventive strategies for LT recipients in CR-KP endemic areas.

Characterizing the patterns of clonal selection in circulating tumor DNA from patients with colorectal cancer refractory to anti-EGFR treatment.

INTRODUCTION: KRAS and EGFR ectodomain-acquired mutations in patients with metastatic colorectal cancer (mCRC) have been correlated with acquired resistance to anti-EGFR monoclonal antibodies (mAbs). We investigated the frequency, co-occurrence, and distribution of acquired KRAS and EGFR mutations in patients with mCRC refractory to anti-EGFR mAbs using circulating tumor DNA (ctDNA). PATIENTS AND METHODS: Sixty-two post-treatment plasma and 20 matching pretreatment archival tissue samples from KRAS (wt) mCRC patients refractory to anti-EGFR mAbs were evaluated by high-sensitivity emulsion polymerase chain reaction for KRAS codon 12, 13, 61, and 146 and EGFR 492 mutations. RESULTS: Plasma analyses showed newly detectable EGFR and KRAS mutations in 5/62 [8%; 95% confidence interval (CI) 0.02-0.18] and 27/62 (44%; 95% CI 0.3-0.56) samples, respectively. KRAS codon 61 and 146 mutations were predominant (33% and 11%, respectively), and multiple EGFR and/or KRAS mutations were detected in 11/27 (41%) cases. The percentage of mutant allele reads was inversely correlated with time since last treatment with EGFR mAbs (P = 0.038). In the matching archival tissue, these mutations were detectable as low-allele-frequency clones in 35% of patients with plasma mutations after treatment with anti-EGFR mAbs and correlated with shorter progression-free survival (PFS) compared with the cases with no new mutations (3.0 versus 8.0 months, P = 0.0004). CONCLUSION: Newly detected KRAS and/or EGFR mutations in plasma ctDNA from patients refractory to anti-EGFR treatment appear to derive from rare, pre-existing clones in the primary tumors. These rare clones were associated with shorter PFS in patients receiving anti-EGFR treatment. Multiple simultaneous mutations in KRAS and EGFR in the ctDNA and the decline in allele frequency after discontinuation of anti-EGFR therapy in a subset of patients suggest that several resistance mechanisms can co-exist and that relative clonal burdens may change over time. Monitoring treatment-induced genetic alterations by sequencing ctDNA could identify biomarkers for treatment screening in anti-EGFR-refractory patients.

Phenotypic heterogeneity and mutational spectrum in a cohort of 45 Italian males subjects with X-linked ectodermal dysplasia.

Ectodermal dysplasias (EDs) are a group of genetic disorders characterized by the abnormal development of the ectodermal-derived structures. X-linked hypohidrotic ectodermal dysplasia, resulting from mutations in ED1 gene, is the most common form. The main purpose of this study was to characterize the phenotype spectrum in 45 males harboring ED1 mutations. The study showed that in addition to the involvement of the major ectodermal tissues, the majority of patients also have alterations of several minor ectodermal-derived structures. Characterizing the clinical spectrum resulting from ED1 gene mutations improves diagnosis and can direct clinical care.

Discovery and molecular characterization of a new cryptovirus dsRNA genome from Japanese persimmon through conventional cloning and high-throughput sequencing.

Through the application of next generation sequencing, in synergy with conventional cloning of DOP-PCR fragments, two double-stranded RNA (dsRNA) molecules of about 1.5 kbp in size were isolated from leaf tissue of a Japanese persimmon (accession SSPI) from Apulia (southern Italy) showing veinlets necrosis. High-throughput sequencing allowed whole genome sequence assembly, yielding a 1,577 and a 1,491 bp contigs identified as dsRNA-1 and dsRNA-2 of a previously undescribed virus, provisionally named as Persimmon cryptic virus (PeCV). In silico analysis showed that both dsRNA fragments were monocistronic and comprised the RNA-dependent RNA polymerase (RdRp) and the capsid protein (CP) genes, respectively. Phylogenetic reconstruction revealed a close relationship of these dsRNAs with those of cryptoviruses described in woody and herbaceous hosts, recently gathered in genus Deltapartitivirus. Virus-specific primers for RT-PCR, designed in the CP cistron, detected viral RNAs also in symptomless persimmon trees sampled from the same geographical area of SSPI, thus proving that PeCV infection may be fairly common and presumably latent.

Genetic Variability of Grapevine Pinot gris virus and Its Association with Grapevine Leaf Mottling and Deformation.

The role of Grapevine Pinot gris virus (GPGV) in the etiology of grapevine leaf mottling and deformation was investigated by biological and molecular assays. A survey on different cultivars from the Trentino Region in Italy showed a widespread distribution of GPGV, which was associated with symptomatic (79%) but also with symptomless (21%) vines. Symptomatic and GPGV-infected 'Pinot gris' vines induced symptoms on grafted vines of healthy Pinot gris or 'Traminer', whereas GPGV-infected but symptomless vines did not. High-throughput sequencing of small RNA (sRNA) populations of two infected Pinot gris accessions confirmed the existence of nearly overlapping viromes in vines with or without symptoms but phylogenetic analyses of the genomes of seven GPGV isolates from Italy and the Czech and Slovak Republics clearly differentiated those infecting symptomatic vines. The involvement of Grapevine rupestris vein feathering virus (GRVFV) in the disease, which was only infecting the symptomatic vine, was ruled out by reverse-transcription polymerase chain reaction studies. Maximum likelihood and Bayesian phylogenetic analysis of two GPGV genomic regions, encompassing part of the movement protein (MP) and coat protein gene sequences and the RNA-dependent RNA polymerase domain of the replicase gene, showed that isolates from symptomatic vines form a lineage distinct from that of symptomless vines. Moreover, the presence or lack of the MP stop codon identified in viral isolates from symptomatic or symptomless vines, respectively, is likely responsible for an MP six amino acids longer in symptomless isolates.

Clinicopathologic characteristics and gene expression analyses of non-KRAS 12/13, RAS-mutated metastatic colorectal cancer.

BACKGROUND: KRAS mutations in codons 12 and 13 are present in ∼40% of all colorectal cancers (CRC). Activating mutations in codons 61 and 146 of KRAS and in codons 12, 13, and 61 of NRAS also occur but are less frequent. The clinicopathologic features and gene expression profiles of this latter subpopulation of RAS-mutant colorectal tumors have not yet been clearly defined but in general are treated similarly to those with KRAS 12 or 13 mutations. PATIENTS AND METHODS: Records of patients with metastatic CRC (mCRC) treated at MD Anderson Cancer Center between December 2000 and August 2012 were reviewed for RAS (KRAS or NRAS) and BRAF mutation status, clinical characteristics, and survival outcomes. To study further with an independent cohort, data from The Cancer Genome Atlas were analyzed to define a gene expression signature for patients whose tumors feature these atypical RAS mutations and explore differences with KRAS 12/13-mutated colorectal tumors. RESULTS: Among the 484 patients reviewed, KRAS 12/13, KRAS 61/146, NRAS, and BRAF mutations were detected in 47.7%, 3.0%, 4.1%, and 7.4%, respectively, of patients who were tested for each of these aberrations. Lung metastases were more common in both the KRAS 12/13-mutated and atypical RAS-mutated cohorts relative to patients with RAS/BRAF wild-type tumors. Gene expression analyses revealed similar patterns regardless of the site of RAS mutation, and in silico functional algorithms predicted that KRAS and NRAS mutations in codons 12, 13, 61, and 146 alter the protein function and drive tumorgenesis. CONCLUSIONS: Clinicopathologic characteristics, survival outcomes, functional impact, and gene expression profiling were similar between patients with KRAS 12/13 and those with NRAS or KRAS 61/146-mutated mCRC. These clinical and bioinformatic findings support the notion that colorectal tumors driven by these RAS mutations are phenotypically similar.

Pazopanib and sunitinib trigger autophagic and non-autophagic death of bladder tumour cells.

BACKGROUND: Tyrosine kinase inhibitors (TKI) such as sunitinib and pazopanib display their efficacy in a variety of solid tumours. However, their use in therapy is limited by the lack of evidence about the ability to induce cell death in cancer cells. Our aim was to evaluate cytotoxic effects induced by sunitinib and pazopanib in 5637 and J82 bladder cancer cell lines. METHODS: Cell viability was tested by MTT assay. Autophagy was evaluated by western blot using anti-LC3 and anti-p62 antibodies, acridine orange staining and FACS analysis. Oxygen radical generation and necrosis were determined by FACS analysis using DCFDA and PI staining. Cathepsin B activation was evaluated by western blot and fluorogenic Z-Arg-Arg-AMC peptide. Finally, gene expression was performed using RT-PCR Profiler array. RESULTS: We found that sunitinib treatment for 24 h triggers incomplete autophagy, impairs cathepsin B activation and stimulates a lysosomal-dependent necrosis. By contrast, treatment for 48 h with pazopanib induces cathepsin B activation and autophagic cell death, markedly reversed by CA074-Me and 3-MA, cathepsin B and autophagic inhibitors, respectively. Finally, pazopanib upregulates the α-glucosidase and downregulates the TP73 mRNA expression. CONCLUSION: Our results showing distinct cell death mechanisms activated by different TKIs, provide the biological basis for novel molecularly targeted approaches.

Analysis of influence of voltage on potential barrier on BiCuVOX and BiTiVOX ceramics.

The BiMeVOx family of compounds appears to be more attractive for applications at low temperatures when ionic conductivity is the determining parameter. The objective of this study was to analysis the influence of voltage of the behavior of the Schottky barrier in both BiCuVOX and BiTiVOX. The samples were analyzed by atomic force microscopy and electric force microscopy (EFM). EFM experiments were conducted to map the electric field distribution on the surface. The formation of Schottky barriers was observed, and their height and width measured. BiCuVOX samples show a barrier width of 140 nm, and BiTiVOX shows a barrier width of 350 nm. The applied voltage has no effect on the barrier width but increases the peak height as observed in the cantilever frequency as measured with the EFM technique.

An improved quantitative mass spectrometry analysis of tumor specific mutant proteins at high sensitivity.

New disease specific biomarkers, especially for cancer, are urgently needed to improve individual diagnosis, prognosis, and treatment selection, that is, for personalized medicine. Genetic mutations that affect protein function drive cancer. Therefore, the detection of such mutations represents a source of cancer specific biomarkers. Here we confirm the implementation of the mutant protein specific immuno-SRM (where SRM is selective reaction monitoring) mass spectrometry method of RAS proteins reported by Wang et al. [Proc. Natl. Acad. Sci. USA 2011, 108, 2444-2449], which exploits an antibody to simultaneously capture the different forms of the target protein and the resolving power and sensitivity of LC-MS/MS and improve the technique by using a more sensitive mass spectrometer. The mutant form G12D was quantified by SRM on a QTRAP 5500 mass spectrometer and the MIDAS workflow was used to confirm the sequence of the targeted peptides. This assay has been applied to quantify wild type and mutant RAS proteins in patient tumors, xenografted human tissue, and benign human epidermal tumors at high sensitivity. The limit of detection for the target proteins was as low as 12 amol (0.25 pg). It requires low starting amounts of tissue (ca.15 mg) that could be obtained from a needle aspiration biopsy. The described strategy could find application in the clinical arena and be applied to the study of expression of protein variants in disease.

Diagnostic accuracy of second-generation dual-source computed tomography coronary angiography with iterative reconstructions: a real-world experience.

PURPOSE: The authors evaluated the diagnostic accuracy of second-generation dual-source (DSCT) computed tomography coronary angiography (CTCA) with iterative reconstructions for detecting obstructive coronary artery disease (CAD). MATERIALS AND METHODS: Between June 2010 and February 2011, we enrolled 160 patients (85 men; mean age 61.2±11.6 years) with suspected CAD. All patients underwent CTCA and conventional coronary angiography (CCA). For the CTCA scan (Definition Flash, Siemens), we use prospective tube current modulation and 70-100 ml of iodinated contrast material (Iomeprol 400 mgI/ ml, Bracco). Data sets were reconstructed with iterative reconstruction algorithm (IRIS, Siemens). CTCA and CCA reports were used to evaluate accuracy using the threshold for significant stenosis at ≥50% and ≥70%, respectively. RESULTS: No patient was excluded from the analysis. Heart rate was 64.3±11.9 bpm and radiation dose was 7.2±2.1 mSv. Disease prevalence was 30% (48/160). Sensitivity, specificity and positive and negative predictive values of CTCA in detecting significant stenosis were 90.1%, 93.3%, 53.2% and 99.1% (per segment), 97.5%, 91.2%, 61.4% and 99.6% (per vessel) and 100%, 83%, 71.6% and 100% (per patient), respectively. Positive and negative likelihood ratios at the per-patient level were 5.89 and 0.0, respectively. CONCLUSIONS: CTCA with second-generation DSCT in the real clinical world shows a diagnostic performance comparable with previously reported validation studies. The excellent negative predictive value and likelihood ratio make CTCA a first-line noninvasive method for diagnosing obstructive CAD.