Selective involvement of serum response factor in pressure-induced myogenic tone in resistance arteries

OBJECTIVE: In resistance arteries, diameter adjustment in response to pressure changes depends on the vascular cytoskeleton integrity. Serum response factor (SRF) is a dispensable transcription factor for cellular growth, but its role remains unknown in resistance arteries. We hypothesized that SRF is required for appropriate microvascular contraction. METHODS AND RESULTS: We used mice in which SRF was specifically deleted in smooth muscle or endothelial cells, and their control. Myogenic tone and pharmacological contraction was determined in resistance arteries. mRNA and protein expression were assessed by quantitative real-time PCR (qRT-PCR) and Western blot. Actin polymerization was determined by confocal microscopy. Stress-activated channel activity was measured by patch clamp. Myogenic tone developing in response to pressure was dramatically decreased by SRF deletion (5.9+/-2.3%) compared with control (16.3+/-3.2%). This defect was accompanied by decreases in actin polymerization, filamin A, myosin light chain kinase and myosin light chain expression level, and stress-activated channel activity and sensitivity in response to pressure. Contractions induced by phenylephrine or U46619 were not modified, despite a higher sensitivity to p38 blockade; this highlights a compensatory pathway, allowing normal receptor-dependent contraction. CONCLUSIONS: This study shows for the first time that SRF has a major part to play in the control of local blood flow via its central role in pressure-induced myogenic tone in resistance arteries

Large oesophageal varice screening by a sequential algorithm using a cirrhosis blood test and optionally capsule endoscopy

BACKGROUND & AIMS: Large oesophageal varice (LEV) screening is recommended in cirrhosis. We performed a prospective study to improve non-invasive LEV screening. DESIGN: 287 patients with cirrhosis had upper gastrointestinal endoscopy (LEV reference), oesophageal capsule endoscopy (ECE), liver elastography and blood marker analyses. CirrhoMeter (cirrhosis blood test), the most accurate non-invasive LEV test, was segmented for cirrhosis (reference comparator) or LEV. VariScreen, a sequential and partially minimally invasive diagnostic algorithm, was developed by multivariate analysis. It uses CirrhoMeter first, then ECE if CirrhoMeter cannot rule LEV out or in, and finally endoscopy if CirrhoMeter+ECE combination remains uninformative. RESULTS: Diagnostic effectiveness rates for LEV were: cirrhosis-segmented CirrhoMeter: 14.6%, LEV-segmented CirrhoMeter: 34.6%, ECE: 60.6% and VariScreen: 66.4% (P ≤ .001 for overall or pair comparison). The respective missed LEV rates were: 2.8%, 5.6%, 8.3% and 5.6% (P = .789). Spared endoscopy rates were, respectively: 15.6%, 36.0%, 70.6% and 69%, (P < .001 for overall or paired comparison except ECE vs VariScreen: P = .743). VariScreen spared 38% of ECE and reduced missed LEV by 87% compared to classical ECE performed in all patients. Excepting cirrhosis-segmented CirrhoMeter, these spared endoscopy rates were significantly higher than that of the Baveno VI recommendation (using platelets and Fibroscan): 18.4% (P < .001). Ascites and Child-Pugh class independently predicted endoscopy sparing by VariScreen: from 86.0% in compensated Child Pugh class A to 24.1% in Child-Pugh class C with ascites. CONCLUSION: VariScreen algorithm significantly reduced the missed LEV rate with ECE by 87%, ECE use by 38% and endoscopy requirement by 69%, and even 86% in compensated cirrhosis

Human bone marrow mesenchymal stem cells : a systematic reappraisal via the genostem experience

Genostem (acronym for “Adult mesenchymal stem cells engineering for connective tissue disorders. From the bench to the bed side”) has been an European consortium of 30 teams working together on human bone marrow Mesenchymal Stem Cell (MSC) biological properties and repair capacity. Part of Genostem activity has been dedicated to the study of basic issues on undifferentiated MSCs properties and on signalling pathways leading to the differentiation into 3 of the connective tissue lineages, osteoblastic, chondrocytic and tenocytic. We have evidenced that native bone marrow MSCs and stromal cells, forming the niche of hematopoietic stem cells, were the same cellular entity located abluminally from marrow sinus endothelial cells. We have also shown that culture-amplified, clonogenic and highly-proliferative MSCs were bona fide stem cells, sharing with other stem cell types the major attributes of self-renewal and of multipotential priming to the lineages to which they can differentiate (osteoblasts, chondrocytes, adipocytes and vascular smooth muscle cells/pericytes). Extensive transcription profiling and in vitro and in vivo assays were applied to identify genes involved in differentiation. Thus we have described novel factors implicated in osteogenesis (FHL2, ITGA5, Fgf18), chondrogenesis (FOXO1A) and tenogenesis (Smad8). Another part of Genostem activity has been devoted to studies of the repair capacity of MSCs in animal models, a prerequisite for future clinical trials. We have developed novel scaffolds (chitosan, pharmacologically active microcarriers) useful for the repair of both bone and cartilage. Finally and most importantly, we have shown that locally implanted MSCs effectively repair bone, cartilage and tendonWork supported by the European Community (Key action 1.2.4-3 Integrated Project Genostem, contract No 503161)

New Algorithm to Determine True Colocalization in Combination with Image Restoration and Time-Lapse Confocal Microscopy to Map Kinases in Mitochondria

The subcellular localization and physiological functions of biomolecules are closely related and thus it is crucial to precisely determine the distribution of different molecules inside the intracellular structures. This is frequently accomplished by fluorescence microscopy with well-characterized markers and posterior evaluation of the signal colocalization. Rigorous study of colocalization requires statistical analysis of the data, albeit yet no single technique has been established as a standard method. Indeed, the few methods currently available are only accurate in images with particular characteristics. Here, we introduce a new algorithm to automatically obtain the true colocalization between images that is suitable for a wide variety of biological situations. To proceed, the algorithm contemplates the individual contribution of each pixel's fluorescence intensity in a pair of images to the overall Pearsońs correlation and Manders' overlap coefficients. The accuracy and reliability of the algorithm was validated on both simulated and real images that reflected the characteristics of a range of biological samples. We used this algorithm in combination with image restoration by deconvolution and time-lapse confocal microscopy to address the localization of MEK1 in the mitochondria of different cell lines. Appraising the previously described behavior of Akt1 corroborated the reliability of the combined use of these techniques. Together, the present work provides a novel statistical approach to accurately and reliably determine the colocalization in a variety of biological images

Prospective Acid Reflux Study of Iran (PARSI): Methodology and study design

<p>Abstract</p> <p>Background</p> <p>Gastroesophageal reflux disease is a common and chronic disorder but long term, prospective studies of the fate of patients seeking medical advice are scarce. This is especially prominent when looking at non-erosive reflux disease (NERD) patients.</p> <p>Methods</p> <p>We designed a prospective cohort to assess the long term outcome of GERD patients referring to gastroenterologists. Consecutive consenting patients, 15 years of age and older, presenting with symptoms suggestive of GERD referring to our outpatient clinics undergo a 30 minute interview. Upper gastrointestinal endoscopy is performed for them with protocol biopsies and blood samples are drawn. Patients are then treated according to a set protocol and followed regularly either in person or by telephone for at least 10 years.</p> <p>Discussion</p> <p>Our data show that such a study is feasible and follow-ups, which are the main concern, can be done in a fairly reliable way to collect data. The results of this study will help to clarify the course of various subgroups of GERD patients after coming to medical attention and their response to treatment considering different variables. In addition, the basic symptoms and biological database will fuel further molecular epidemiologic studies.</p