Which is the best algorithm for evaluating a patient’s candidate to sleeve with suspected reflux or hiatal hernia: is manometry or reflux assessment always necessary

Laparoscopic sleeve gastrectomy (SG) has reached wide popularity during the last 15 years, owing to limited morbidity and mortality rates, very successful weight loss results, and impact on comorbidities. However, the postoperative development or worsening of gastroesophageal reflux disease (GERD) is one of the most important drawbacks of this surgical procedure. To date, there is great heterogeneity concerning the definition of GERD, the indication for SG in patients with GERD, and the standardization of pre and postoperative diagnostic pathways. In patients with severe obesity, a strictly symptom-based diagnosis of GERD is unreliable. In fact, a high rate of silent GERD (s-GERD, asymptomatic patients despite objective evidence of GERD) has been reported. Moreover, patients with preoperative s-GERD have a significantly higher risk of experiencing GERD symptoms after SG. For these reasons, the reflux burden and the competence of the anti-reflux barrier should be carefully assessed during the preoperative work-up of patients undergoing SG. Ambulatory pH monitoring (APM) and high-resolution manometry (HRM) are useful diagnostic tools that could provide valuable evidence in the guidance of surgical strategy. In this review, we evaluate the current literature concerning the use of APM and HRM in the diagnostic pathway before SG, as well as their predictive value for the evolution of GERD in the postoperative course. Moreover, we propose a diagnostic algorithm for preoperative GERD assessment, which includes validated symptom questionnaires, upper gastrointestinal endoscopy, APM, and HRM

Nicotinamide inhibits melanoma in vitro and in vivo

Background: Even though new therapies are available against melanoma, novel approaches are needed to overcome resistance and high-toxicity issues. In the present study the anti-melanoma activity of Nicotinamide (NAM), the amide form of Niacin, was assessed in vitro and in vivo. Methods: Human (A375, SK-MEL-28) and mouse (B16-F10) melanoma cell lines were used for in vitro investigations. Viability, cell-death, cell-cycle distribution, apoptosis, Nicotinamide Adenine Dinucleotide+ (NAD+), Adenosine Triphosphate (ATP), and Reactive Oxygen Species (ROS) levels were measured after NAM treatment. NAM anti-SIRT2 activity was tested in vitro; SIRT2 expression level was investigated by in silico transcriptomic analyses. Melanoma growth in vivo was measured in thirty-five C57BL/6 mice injected subcutaneously with B16-F10 melanoma cells and treated intraperitoneally with NAM. Interferon (IFN)-γ-secreting murine cells were counted with ELISPOT assay. Cytokine/chemokine plasmatic levels were measured by xMAP technology. Niacin receptors expression in human melanoma samples was also investigated by in silico transcriptomic analyses. Results: NAM reduced up to 90% melanoma cell number and induced: I) accumulation in G1-phase (40% increase), ii) reduction in S- A nd G2-phase (about 50% decrease), iii) a 10-fold increase of cell-death and 2.5-fold increase of apoptosis in sub-G1 phase, iv) a significant increase of NAD+, ATP, and ROS levels, v) a strong inhibition of SIRT2 activity in vitro. NAM significantly delayed tumor growth in vivo (p ≤ 0.0005) and improved survival of melanoma-bearing mice (p ≤ 0.0001). About 3-fold increase (p ≤ 0.05) of Interferon-gamma (IFN-γ) producing cells was observed in NAM treated mice. The plasmatic expression levels of 6 cytokines (namely: Interleukin 5 (IL-5), Eotaxin, Interleukin 12 (p40) (IL12(p40)), Interleukin 3 (IL-3), Interleukin 10 (IL-10) and Regulated on Activation Normal T Expressed and Secreted (RANTES) were significantly changed in the blood of NAM treated mice, suggesting a key role of the immune response. The observed inhibitory effect of NAM on SIRT2 enzymatic activity confirmed previous evidence; we show here that SIRT2 expression is significantly increased in melanoma and inversely related to melanoma-patients survival. Finally, we show for the first time that the expression levels of Niacin receptors HCAR2 and HCAR3 is almost abolished in human melanoma samples. Conclusion: NAM shows a relevant anti-melanoma activity in vitro and in vivo and is a suitable candidate for further clinical investigations

Investigating serum and tissue expression identified a cytokine/chemokine signature as a highly effective melanoma marker

The identification of reliable and quantitative melanoma biomarkers may help an early diagnosis and may directly affect melanoma mortality and morbidity. The aim of the present study was to identify effective biomarkers by investigating the expression of 27 cytokines/chemokines in melanoma compared to healthy controls, both in serum and in tissue samples. Serum samples were from 232 patients recruited at the IDI-IRCCS hospital. Expression was quantified by xMAP technology, on 27 cytokines/chemokines, compared to the control sera. RNA expression data of the same 27 molecules were obtained from 511 melanoma-and healthy-tissue samples, from the GENT2 database. Statistical analysis involved a 3-step approach: analysis of the single-molecules by Mann–Whitney analysis; analysis of paired-molecules by Pearson correlation; and profile analysis by the machine learning algorithm Support Vector Machine (SVM). Single-molecule analysis of serum expression identified IL-1b, IL-6, IP-10, PDGF-BB, and RANTES differently expressed in melanoma (p < 0.05). Expression of IL-8, GM-CSF, MCP-1, and TNF-α was found to be significantly correlated with Breslow thickness. Eotaxin and MCP-1 were found differentially expressed in male vs. female patients. Tissue expression analysis identified very effective marker/predictor genes, namely, IL-1Ra, IL-7, MIP-1a, and MIP-1b, with individual AUC values of 0.88, 0.86, 0.93, 0.87, respectively. SVM analysis of the tissue expression data identified the combination of these four molecules as the most effective signature to discriminate melanoma patients (AUC = 0.98). Validation, using the GEPIA2 database on an additional 1019 independent samples, fully confirmed these observations. The present study demonstrates, for the first time, that the IL-1Ra, IL-7, MIP-1a, and MIP-1b gene signature discriminates melanoma from control tissues with extremely high efficacy. We therefore propose this 4-molecule combination as an effective melanoma marker

c-FLIP regulates autophagy by interacting with Beclin-1 and influencing its stability

c-FLIP (cellular FLICE-like inhibitory protein) protein is mostly known as an apoptosis modulator. However, increasing data underline that c-FLIP plays multiple roles in cellular homoeostasis, influencing differently the same pathways depending on its expression level and isoform predominance. Few and controversial data are available regarding c-FLIP function in autophagy. Here we show that autophagic flux is less effective in c-FLIP−/− than in WT MEFs (mouse embryonic fibroblasts). Indeed, we show that the absence of c-FLIP compromises the expression levels of pivotal factors in the generation of autophagosomes. In line with the role of c-FLIP as a scaffold protein, we found that c-FLIPL interacts with Beclin-1 (BECN1: coiled-coil, moesin-like BCL2-interacting protein), which is required for autophagosome nucleation. By a combination of bioinformatics tools and biochemistry assays, we demonstrate that c-FLIPL interaction with Beclin-1 is important to prevent Beclin-1 ubiquitination and degradation through the proteasomal pathway. Taken together, our data describe a novel molecular mechanism through which c-FLIPL positively regulates autophagy, by enhancing Beclin-1 protein stability

Identification of proteins associated with ligand-activated estrogen receptor alpha in human breast cancer cell nuclei by tandem affinity purification and nanoLC-MS/MS.

Estrogen receptor a (ER-a) is a key mediator of estrogen actions in breast cancer (BC) cells. Understanding the effects of ligand-activated ER-a in target cells requires identification of the molecular partners acting in concert with this nuclear receptor to transduce the hormonal signal. We applied tandem affinity purification (TAP), glycerol gradient centrifugation and MS analysis to isolate and identify proteins interacting with ligand-activated ER-a in MCF-7 cell nuclei. This led to the identification of 264 ER-associated proteins, whose functions highlight the hinge role of ER-a in the coordination of multiple hormone-regulated nuclear processes in BC cells

Seminal but not serum levels of holotranscobalamin are altered in morbid obesity and correlate with semen quality: a pilot single centre study

Vitamin B12 (cobalamin) is an essential cofactor in the one-carbon metabolism. One-carbon metabolism is a set of complex biochemical reactions, through which methyl groups are utilised or generated, and thus plays a vital role to many cellular functions in humans. Low levels of cobalamin have been associated to metabolic/reproductive pathologies. However, cobalamin status has never been investigated in morbid obesity in relation with the reduced semen quality. We analysed the cross-sectional data of 47-morbidly-obese and 21 lean men at Careggi University Hospital and evaluated total cobalamin (CBL) and holotranscobalamin (the active form of B12; holoTC) levels in serum and semen. Both seminal and serum concentrations of holoTC and CBL were lower in morbidly obese compared to lean men, although the difference did not reach any statistical significance for serum holoTC. Seminal CBL and holoTC were significantly higher than serum levels in both groups. Significant positive correlations were observed between seminal holoTC and total sperm motility (r = 0.394, p = 0.012), sperm concentration (r = 0.401, p = 0.009), total sperm number (r = 0.343, p = 0.028), and negative correlation with semen pH (r = −0.535, p = 0.0001). ROC analysis supported seminal holoTC as the best predictor of sperm number (AUC = 0.769 ± 0.08, p = 0.006). Our findings suggest that seminal rather than serum levels of holoTC may represent a good marker of semen quality in morbidly obese subjects

Resources and tools for rare disease variant interpretation

Collectively, rare genetic disorders affect a substantial portion of the world’s population. In most cases, those affected face difficulties in receiving a clinical diagnosis and genetic characterization. The understanding of the molecular mechanisms of these diseases and the development of therapeutic treatments for patients are also challenging. However, the application of recent advancements in genome sequencing/analysis technologies and computer-aided tools for predicting phenotype-genotype associations can bring significant benefits to this field. In this review, we highlight the most relevant online resources and computational tools for genome interpretation that can enhance the diagnosis, clinical management, and development of treatments for rare disorders. Our focus is on resources for interpreting single nucleotide variants. Additionally, we present use cases for interpreting genetic variants in clinical settings and review the limitations of these results and prediction tools. Finally, we have compiled a curated set of core resources and tools for analyzing rare disease genomes. Such resources and tools can be utilized to develop standardized protocols that will enhance the accuracy and effectiveness of rare disease diagnosis

Substrate specificity of microbial transglutaminase as revealed by three-dimensional docking simulation and mutagenesis

Transglutaminases (TGases) are used in fields such as food and pharmaceuticals. Unlike other TGases, microbial transglutaminase (MTG) activity is Ca2+-independent, broadening its application. Here, a three-dimensional docking model of MTG binding to a peptide substrate, CBZ-Gln-Gly, was simulated. The data reveal CBZ-Gln-Gly to be stretched along the MTG active site cleft with hydrophobic and/or aromatic residues interacting directly with the substrate. Moreover, an oxyanion binding site for TGase activity may be constructed from the amide groups of Cys64 and/or Val65. Alanine mutagenesis verified the simulated binding region and indicated that large molecules can be widely recognized on the MTG cleft

Acute kidney injury promotes development of papillary renal cell adenoma and carcinoma from renal progenitor cells.

Acute tissue injury causes DNA damage and repair processes involving increased cell mitosis and polyploidization, leading to cell function alterations that may potentially drive cancer development. Here, we show that acute kidney injury (AKI) increased the risk for papillary renal cell carcinoma (pRCC) development and tumor relapse in humans as confirmed by data collected from several single-center and multicentric studies. Lineage tracing of tubular epithelial cells (TECs) after AKI induction and long-term follow-up in mice showed time-dependent onset of clonal papillary tumors in an adenoma-carcinoma sequence. Among AKI-related pathways, NOTCH1 overexpression in human pRCC associated with worse outcome and was specific for type 2 pRCC. Mice overexpressing NOTCH1 in TECs developed papillary adenomas and type 2 pRCCs, and AKI accelerated this process. Lineage tracing in mice identified single renal progenitors as the cell of origin of papillary tumors. Single-cell RNA sequencing showed that human renal progenitor transcriptome showed similarities to PT1, the putative cell of origin of human pRCC. Furthermore, NOTCH1 overexpression in cultured human renal progenitor cells induced tumor-like 3D growth. Thus, AKI can drive tumorigenesis from local tissue progenitor cells. In particular, we find that AKI promotes the development of pRCC from single progenitors through a classical adenoma-carcinoma sequence

ELIXIR‐IT: a growing support to national and international research in life sciences

ELIXIR-IT gathers most of the excellence centres or bioinformatics in Italy and is striving to assume pivotal role for the national and international life science communities. This is reflected by the growing number of bioinformatics services, initiatives and projects supported or participated by ELIXIR-IT, including H2020 grants and a number of training efforts delivering state of the arts courses on basic and advanced topics. In this poster we highlight some of the activities