A cell sizer network involving Cln3 and Far1 controls entrance into S phase in the mitotic cycle of budding yeast

Saccharomyces cerevisiae must reach a carbon source-modulated critical cell size, protein content per cell at the onset of DNA replication (Ps), in order to enter S phase. Cells grown in glucose are larger than cells grown in ethanol. Here, we show that an increased level of the cyclin-dependent inhibitor Far1 increases cell size, whereas far1Δ cells start bud emergence and DNA replication at a smaller size than wild type. Cln3Δ, far1Δ, and strains overexpressing Far1 do not delay budding during an ethanol glucose shift-up as wild type does. Together, these findings indicate that Cln3 has to overcome Far1 to trigger Cln–Cdc28 activation, which then turns on SBF- and MBF-dependent transcription. We show that a second threshold is required together with the Cln3/Far1 threshold for carbon source modulation of Ps. A new molecular network accounting for the setting of Ps is proposed

Bone demineralization and vertebral fractures in endogenous cortisol excess: role of disease etiology and gonadal status

INTRODUCTION: The effects of endogenous cortisol (F) excess on bone mass and vertebral fractures have still not been thoroughly investigated. The aim of this cross-sectional case-control study was to investigate factors influencing bone demineralization and vertebral fractures in different conditions of F excess, i.e. Cushing's disease and adrenal and ectopic Cushing's syndrome. MATERIALS AND METHODS: Eighty consecutive patients and 80 controls were prospectively enrolled: 37 patients (21 females) with pituitary ACTH-secreting adenoma, 18 (14 females) with adrenocortical adenoma, 15 (11 females) with adrenal carcinoma of mixed secretion, and 10 (three females) with ectopic ACTH secretion. The groups had similar age. At diagnosis, bone mineral density (BMD) was determined by the dual-energy x-ray absorptiometry technique at the lumbar spine (L1-L4) and femoral neck; vertebral fractures were investigated by standard spinal radiographs. RESULTS: When comparing the groups with different etiology of F excess, the patients with ectopic ACTH secretion had higher F and lower BMD values than the other subgroups. Morning F (P = 0.03) and testosterone levels (P = 0.04) correlated with lumbar BMD. Vertebral fractures were found in 61 (76%) of the patients, were multiple in 52 (85%) of the cases, and clinically evident in 32 (52%). Only multiple fractures were more frequent in patients with ectopic ACTH hypersecretion (P < 0.05). Lumbar spine BMD was the best predictor of vertebral fractures (P < 0.01). Surprisingly, amenorrheic and eumenorrheic women had similar BMD values and fracture prevalence. CONCLUSION: A high prevalence (76%) of vertebral fracture was revealed, regardless of the etiology of the patients' hypercortisolism. The harmful effects of F excess at the spine were partly counterbalanced by the increased androgen production but were not affected by gonadal status in women

FOXP1 circular RNA sustains mesenchymal stem cell identity via microRNA inhibition

Stem cell identity and plasticity are controlled by master regulatory genes and complex circuits also involving non-coding RNAs. Circular RNAs (circRNAs) are a class of RNAs generated from protein-coding genes by backsplicing, resulting in stable RNA structures devoid of free 5' and 3' ends. Little is known of the mechanisms of action of circRNAs, let alone in stem cell biology. In this study, for the first time, we determined that a circRNA controls mesenchymal stem cell (MSC) identity and differentiation. High-throughput MSC expression profiling from different tissues revealed a large number of expressed circRNAs. Among those, circFOXP1 was enriched in MSCs compared to differentiated mesodermal derivatives. Silencing of circFOXP1 dramatically impaired MSC differentiation in culture and in vivo. Furthermore, we demonstrated a direct interaction between circFOXP1 and miR-17-3p/miR-127-5p, which results in the modulation of non-canonical Wnt and EGFR pathways. Finally, we addressed the interplay between canonical and non-canonical Wnt pathways. Reprogramming to pluripotency of MSCs reduced circFOXP1 and non-canonical Wnt, whereas canonical Wnt was boosted. The opposing effect was observed during generation of MSCs from human pluripotent stem cells. Our results provide unprecedented evidence for a regulatory role for circFOXP1 as a gatekeeper of pivotal stem cell molecular networks

An Exploratory Study of Field Failures

Field failures, that is, failures caused by faults that escape the testing phase leading to failures in the field, are unavoidable. Improving verification and validation activities before deployment can identify and timely remove many but not all faults, and users may still experience a number of annoying problems while using their software systems. This paper investigates the nature of field failures, to understand to what extent further improving in-house verification and validation activities can reduce the number of failures in the field, and frames the need of new approaches that operate in the field. We report the results of the analysis of the bug reports of five applications belonging to three different ecosystems, propose a taxonomy of field failures, and discuss the reasons why failures belonging to the identified classes cannot be detected at design time but shall be addressed at runtime. We observe that many faults (70%) are intrinsically hard to detect at design-time

Treatment efficacy with electrochemotherapy: A multi-institutional prospective observational study on 376 patients with superficial tumors

BACKGROUND: Cutaneous metastases represent a therapeutic challenge. An increasing body of experience suggests that electrochemotherapy (ECT) provides effective tumor control, although its evidence basis should be strengthened. METHODS: This prospective, multicenter, observational study enrolled patients with superficial metastases, who underwent ECT at 10 centers between 2008 and 2013. Outcomes included adherence to European Standard Operating Procedures of ECT (ESOPE), tumor response, local progression-free survival (LPFS), toxicity and patient-reported outcomes (PROs, EORTC QLQ-C30 plus an 8-item questionnaire). RESULTS: We enrolled 376 eligible patients. Tumor histotype distribution was as follows: melanoma, 56%; squamous cell carcinoma, 11%; Kaposi sarcoma, 11%; breast carcinoma, 8%; basal cell carcinoma, 6%; soft tissue sarcomas, 3%; others, 5%. We registered 1304 target tumors (median size 1 cm). Treatment adhered to ESOPE in 88% of patients as to the route of drug administration, and in 70% as to electrode application. The procedure was mainly performed under sedation (64.6%) and by using intravenous chemotherapy (93.4%). Tumor response rate at 60 days was 88% (complete, 50%). Small tumor size predicted complete response achievement (OR 2.24, p = 0.003), higher LPFS (HR 0.68, p = 0.004) and improved PROs (Global Health Status, p < 0.001; wound bleeding, p < 0.001; healing, p = 0.002; and aesthetics, p < 0.001). Skin toxicity (grade 653, 7.8%) was lower in patients with tumors <2 cm (p 640.001). One-year LPFS was 73.7% (95%CI 68.4-78.3). CONCLUSIONS: ECT represents a valuable skin-directed therapy across a range of malignancies. The most frequently applied treatment modality is intravenous chemotherapy under sedation. Small tumor size predicts durable tumor control, fewer side-effects and better PROs

Ventilatory support and mechanical properties of the fibrotic lung acting as a "squishy ball"

Protective ventilation is the cornerstone of treatment of patients with the acute respiratory distress syndrome (ARDS); however, no studies have yet established the best ventilatory strategy to adopt when patients with acute exacerbation of interstitial lung disease (AE-ILD) are admitted to the intensive care unit. Due to the severe impairment of the respiratory mechanics, the fibrotic lung is at high risk of developing ventilator-induced lung injury, regardless of the lung fibrosis etiology. The purpose of this review is to analyze the effects of mechanical ventilation in AE-ILD and to increase the knowledge on the characteristics of fibrotic lung during artificial ventilation, introducing the concept of "squishy ball lung". The role of positive end-expiratory pressure is discussed, proposing a "lung resting strategy" as opposed to the "open lung approach". The review also discusses the practical management of AE-ILD patients discussing illustrative clinical cases

Substantial Histone Reduction Modulates Genomewide Nucleosomal Occupancy and Global Transcriptional Output

The basic unit of genome packaging is the nucleosome, and nucleosomes have long been proposed to restrict DNA accessibility both to damage and to transcription. Nucleosome number in cells was considered fixed, but recently aging yeast and mammalian cells were shown to contain fewer nucleosomes. We show here that mammalian cells lacking High Mobility Group Box 1 protein (HMGB1) contain a reduced amount of core, linker, and variant histones, and a correspondingly reduced number of nucleosomes, possibly because HMGB1 facilitates nucleosome assembly. Yeast nhp6 mutants lacking Nhp6a and -b proteins, which are related to HMGB1, also have a reduced amount of histones and fewer nucleosomes. Nucleosome limitation in both mammalian and yeast cells increases the sensitivity of DNA to damage, increases transcription globally, and affects the relative expression of about 10% of genes. In yeast nhp6 cells the loss of more than one nucleosome in four does not affect the location of nucleosomes and their spacing, but nucleosomal occupancy. The decrease in nucleosomal occupancy is non-uniform and can be modelled assuming that different nucleosomal sites compete for available histones. Sites with a high propensity to occupation are almost always packaged into nucleosomes both in wild type and nucleosome-depleted cells; nucleosomes on sites with low propensity to occupation are disproportionately lost in nucleosome-depleted cells. We suggest that variation in nucleosome number, by affecting nucleosomal occupancy both genomewide and gene-specifically, constitutes a novel layer of epigenetic regulation

Single tube liquid biopsy for advanced non-small cell lung cancer

The need for a liquid biopsy in non-small cell lung cancer (NSCLC) patients is rapidly increasing. We studied the relation between overall survival (OS) and the presence of four cancer biomarkers from a single blood draw in advanced NSCLC patients: EpCAM(high) circulating tumor cells (CTC), EpCAM(low) CTC, tumor-derived extracellular vesicles (tdEV) and cell-free circulating tumor DNA (ctDNA). EpCAM(high) CTC were detected with CellSearch, tdEV in the CellSearch images and EpCAM(low) CTC with filtration after CellSearch. ctDNA was isolated from plasma and mutations present in the primary tumor were tracked with deep sequencing methods. In 97 patients, 21% had >= 2 EpCAM(high) CTC, 15% had >= 2 EpCAM(low) CTC, 27% had >= 18 tdEV and 19% had ctDNA with >= 10% mutant allele frequency. Either one of these four biomarkers could be detected in 45% of the patients and all biomarkers were present in 2%. In 11 out of 16 patients (69%) mutations were detected in the ctDNA. Two or more unfavorable biomarkers were associated with poor OS. The presence of EpCAM(high) CTC and elevated levels of tdEV and ctDNA was associated with a poor OS; however, the presence of EpCAM(low) CTC was not. This single tube approach enables simultaneous analysis of multiple biomarkers to explore their potential as a liquid biopsy