The histone binding capacity of SPT2 controls chromatin structure and function in Metazoa

Histone chaperones control nucleosome density and chromatin structure. In yeast, the H3-H4 chaperone Spt2 controls histone deposition at active genes but its roles in metazoan chromatin structure and organismal physiology are not known. Here we identify the Caenorhabditis elegans ortholog of SPT2 (CeSPT-2) and show that its ability to bind histones H3-H4 is important for germline development and transgenerational epigenetic gene silencing, and that spt-2 null mutants display signatures of a global stress response. Genome-wide profiling showed that CeSPT-2 binds to a range of highly expressed genes, and we find that spt-2 mutants have increased chromatin accessibility at a subset of these loci. We also show that SPT2 influences chromatin structure and controls the levels of soluble and chromatin-bound H3.3 in human cells. Our work reveals roles for SPT2 in controlling chromatin structure and function in Metazoa.</p

ZFPIP/Zfp462 is involved in P19 cell pluripotency and in their neuronal fate.

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Comparative Analysis of Methods to Induce Myocardial Infarction in a Closed-Chest Rabbit Model

Objective. To develop a rabbit model of closed-chest catheter-induced myocardial infarction. Background. Limitations of rodent and large animal models justify the search for clinically relevant alternatives. Methods. Microcatheterization of the heart was performed in 47 anesthetized 3-4 kg New Zealand rabbits to test five techniques of myocardial ischemia: free coils (n=4), interlocking coils (n=4), thrombogenic gelatin sponge (n=4), balloon occlusion (n=4), and alcohol injection (n=8). In order to limit ventricular fibrillation, an antiarrhythmic protocol was implemented, with beta-blockers/amiodarone before and xylocaine infusion during the procedure. Clinical, angiographic, and echographic data were gathered. End points included demonstration of vessel occlusion (TIMI flow grades 0 and 1 on the angiogram), impairment of left ventricular function at 2 weeks after procedure (by echocardiography), and pathologically confirmed myocardial infarction. Results. The best arterial access was determined to be through the right carotid artery. The internal mammary guiding catheter 4-Fr was selected as the optimal device for selective intracoronary injection. Free coils deployed prematurely and tended to prolapse into the aorta. Interlocking coils did not deploy completely and failed to provide reliable results. Gelatin sponge was difficult to handle, adhered to the catheter, and could not be clearly visualized by fluoroscopy. Balloon occlusion yielded inconsistent results. Alcohol injection was the most efficient and reproducible method for inducing myocardial infarction (4 out of 6 animals), the extent of which could be fine-tuned by using a coaxial balloon catheter as a microcatheter (0.52 mm) to achieve a superselective injection of 0.2 mL of alcohol. This approach resulted in a 20% decrease in LVEF and infarcted myocardium was confirmed histologically. Conclusions. By following a stepwise approach, a minimally invasive, effective, and reproducible rabbit model of catheter-induced myocardial infarction has been developed which addresses the limitations of rodent experiments while avoiding the logistical and cost issues associated with large animal models

Recruitment and activation of hepatic stellate cells by uveal melanoma cells in a xenograft mouse model

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Phrenic nerve stimulation in an ovine model with temporary removable pacing leads

International audienceBackground: The objective of this study was to assess the feasibility and safety of a novel, removable, surgically implanted, temporary neurostimulation approach involving the distal portion of the phrenic nerve.Methods: Temporary phrenic nerve pacing electrodes were implanted surgically using an ovine model (4 animals). The primary endpoint was the ability to successfully match the animal's minute-ventilation upon implantation of both phrenic nerve pacers on day 1. Secondary endpoints were successful phrenic neurostimulation by both electrodes 15 and 30 days after initial implantation. We also assessed safe removal of the electrodes at 15 and 30 days after implementation.Results: In 3 of 4 animals, electrodes were successfully implanted in both right and left phrenic nerves. On day 1, median ventilation-minute induced by neurostimulation was not significantly different from baseline ventilation-minute [4.9 L·min-1 (4.4-5.5) vs. 4.4 L·min-1 (4.3-5.2); P=0.4] after 15 minutes. Neurostimulation was still possible 15 and 30 days after implementation in all left side phrenic nerves. On the right side, stimulation was possible at all times in 1 animal but not in the remaining 3 animals for at least one time point, possibly due to lead displacement. Analysis of pathology after percutaneous electrode removal showed integrity of the distal portion of all phrenic nerves.Conclusions: Efficient temporary neurostimulation through the distal portion of the phrenic nerve was possible at baseline. The main complication was the displacement of electrodes on the right phrenic nerve on two occasions, which was due to the anatomy of the ovine model. It compromised diaphragm pacing on day 15 and day 30. The electrodes could be safely removed percutaneously without damage to the phrenic nerves

Synergic Interactions Between Hepatic Stellate Cells and Uveal Melanoma in Metastatic Growth

Uveal melanoma (UM) is a malignant intraocular tumor that spreads to the liver in half of the cases. Since hepatic cells could play a role in the therapeutic resistance of metastatic UM, the purpose of our study was to investigate the pro-invasive role of hepatic stellate cells (HSteCs) in metastatic UM at the micro- and macro-metastatic stages. We first performed an immunostaining with the alpha-smooth muscle actin (&alpha;SMA) to localize activated HSteCs in UM liver macro-metastases from four patients. Their accumulation of collagen was assessed with Masson&rsquo;s Trichrome stain. Next, we inoculated metastatic UM cells alone or with human HSteCs in triple-immunodeficient mice, in order to determine if HSteCs are recruited as early as the micro-metastatic stage. The growth of metastatic foci was imaged in the liver by ex vivo fluorescence imaging. Histological analyses were performed with Masson&rsquo;s Trichrome and Picrosirius Red stains, and antibodies against Melan-A and &alpha;SMA. The collagen content was measured in xenografts by quantitative polarization microscopy. In patient hepatectomy samples, activated HSteCs and their pathological matrix were localized surrounding the malignant lesions. In the mouse xenograft model, the number of hepatic metastases was increased when human HSteCs were co-inoculated. Histological analyses revealed a significant recruitment of HSteCs near the micro/macrolesions, and an increase in fibrillar collagen production. Our results show that HSteCs can provide a permissive microenvironment and might increase the therapeutic resistance of metastatic UM

Extracellular Vesicles from Ocular Melanoma Have Pro-Fibrotic and Pro-Angiogenic Properties on the Tumor Microenvironment

Uveal melanoma (UM) is the most common primary intraocular tumor and often spreads to the liver. Intercellular communication though extracellular vesicles (EVs) plays an important role in several oncogenic processes, including metastasis, therapeutic resistance, and immune escape. This study examines how EVs released by UM cells modify stellate and endothelial cells in the tumor microenvironment. The surface markers, and the concentration and size of EVs derived from UM cells or choroidal melanocytes were characterized by high-resolution flow cytometry, electron microscopy, and Western blotting. The selective biodistribution of EVs was studied in mice by fluorescence imaging. The activation/contractility of stellate cells and the tubular organization of endothelial cells after exposure to melanomic EVs were determined by traction force microscopy, collagen gel contraction, or endothelial tube formation assays. We showed that large EVs from UM cells and healthy melanocytes are heterogenous in size, as well as their expression of phosphatidylserine, tetraspanins, and Tsg101. Melanomic EVs mainly accumulated in the liver and lungs of mice. Hepatic stellate cells with internalized melanomic EVs had increased contractility, whereas EV-treated endothelial cells developed more capillary-like networks. Our study demonstrates that the transfer of EVs from UM cells leads to a pro-fibrotic and pro-angiogenic phenotype in hepatic stellate and endothelial cells

The impact of raw materials and baking conditions on Maillard reaction products, thiamine, folate, phytic acid and minerals in white bread

The aim of this study was to develop a white bread with improved nutrient contents and reduced levels of potentially harmful Maillard reaction products such as Ne-carboxymethyllysine (CML) and 5-hydroxymethylfurfural (HMF). Assays were carried out through a full factorial experimental design allowing the simultaneous analysis of four factors at two levels: (1) wheat flour extraction rates (ash content: 0.60%-0.72%), (2) leavening agents (bakers' yeast -bakers' yeast and sourdough), (3) prebaking and (4) baking conditions (different sets of time and temperature). The baking conditions affected HMF and CML as well as certain mineral contents. A reduced baking temperature along with a prolonged heat treatment was found to be favourable for reducing both the CML (up to 20%) and HMF concentrations (up to 96%). The presence of sourdough decreased the formation of CML (up to 28%), and increased the apparent amounts of calcium (up to 8%) and manganese (up to 17.5%) probably through acidification of the dough. The extraction rate of flours as well as interactions between multiple factors also affected certain mineral content. However, compounds like folate, thiamine, copper, zinc, iron and phytic acid were not affected by any of the factors studied

Human Organ-Specific 3D Cancer Models Produced by the Stromal Self-Assembly Method of Tissue Engineering for the Study of Solid Tumors

Cancer research has considerably progressed with the improvement of in vitro study models, helping to understand the key role of the tumor microenvironment in cancer development and progression. Over the last few years, complex 3D human cell culture systems have gained much popularity over in vivo models, as they accurately mimic the tumor microenvironment and allow high-throughput drug screening. Of particular interest, in vitrohuman 3D tissue constructs, produced by the self-assembly method of tissue engineering, have been successfully used to model the tumor microenvironment and now represent a very promising approach to further develop diverse cancer models. In this review, we describe the importance of the tumor microenvironment and present the existing in vitro cancer models generated through the self-assembly method of tissue engineering. Lastly, we highlight the relevance of this approach to mimic various and complex tumors, including basal cell carcinoma, cutaneous neurofibroma, skin melanoma, bladder cancer, and uveal melanoma