The Dichotomy of Vascular Smooth Muscle Differentiation/De- Differentiation in Health and Disease

Vascular smooth muscle cells (SMCs) are thought to display cellular plasticity by alternating between a quiescent ‘contractile’ differentiated phenotype and a proliferative ‘synthetic’ de-differentiated phenotype in response to induction of distinct developmental pathways or to local micro-environmental cues. This classic de-differentiation and re-programming process is associated with a significant loss in the expression of key SMC differentiation marker genes for a large number of proliferative vascular diseases in vivo and in sub-cultured cells in vitro. Regarded as essential for vascular regeneration and repair in vivo, phenotypic modulation represents a critical target for therapeutic intervention. However, recent evidence now suggests that this process of vascular regeneration may also involve differentiation of resident vascular stem cells and the accumulation of stem cell-derived myogenic, osteochondrogenic and macrophage-like phenotypes within vascular lesions in vivo and across sub-cultured SMC cell populations in vitro. This review summarises our current knowledge of vascular regeneration, de-differentiation and re-programming of vascular SMCs, and focuses on the accumulating evidence of a putative role for stem cell-derived progeny and the evolving dichotomy of the origin of SMC-like cells during intimal-medial thickening and the progression of arteriosclerotic disease

Differential effects of alcohol and its metabolite acetaldehyde on vascular smooth muscle cell Notch signaling and growth

Alcohol (EtOH) consumption can variously affect cardiovascular disease. Our aim was to compare the effects of EtOH and its primary metabolite acetaldehyde (ACT) on vascular smooth muscle Notch signaling and cell growth, which are important for atherogenesis. Human coronary artery smooth muscle cells (HCASMCs) were treated with EtOH (25 mM) or ACT (10 or 25 μM). As previously reported, EtOH inhibited Notch signaling and growth of HCASMCs. In contrast, ACT treatment stimulated HCASMC proliferation (cell counts) and increased proliferating cell nuclear antigen expression, concomitant with stimulation of Notch signaling, as determined by increased Notch receptor (N1 and N3) and target gene (Hairy-related transcription factor 1–3) mRNA levels. Interaction of the ligand with the Notch receptor initiates proteolytic cleavage by α- and γ-secretase, resulting in the release of the active Notch intracellular domain. Neither EtOH nor ACT had any significant effect on α-secretase activity. A fluorogenic peptide cleavage assay demonstrated almost complete inhibition by EtOH of Delta-like ligand 4-stimulated γ-secretase activity in solubilized HCASMCs (similar to the effect of the control inhibitor DAPT) but no effect of ACT treatment. EtOH, but not ACT, affected the association and distribution of the γ-secretase catalytic subunit presenilin-1 with lipid rafts, as determined by dual fluorescent labeling and confocal microscopic visualization. In conclusion, ACT stimulates vascular smooth muscle cell Notch signaling and growth, effects opposite to those of EtOH. These differential actions on vascular smooth muscle cells of EtOH and its metabolite ACT may be important in mediating the ultimate effects of drinking on cardiovascular disease

Reactive oxygen species (ROS), intimal thickening, and subclinical atherosclerotic disease

Arteriosclerosis causes significant morbidity and mortality worldwide. Central to this process is the development of subclinical non-atherosclerotic intimal lesions before the appearance of pathologic intimal thickening and advanced atherosclerotic plaques. Intimal thickening is associated with several risk factors, including oxidative stress due to reactive oxygen species (ROS), inflammatory cytokines and lipid. The main ROS producing systems in-vivo are reduced nicotinamide dinucleotide phosphate (NADPH) oxidase (NOX). ROS effects are context specific. Exogenous ROS induces apoptosis and senescence, whereas intracellular ROS promotes stem cell differentiation, proliferation, and migration. Lineage tracing studies using murine models of subclinical atherosclerosis have revealed the contributory role of medial smooth muscle cells (SMCs), resident vascular stem cells, circulating bone-marrow progenitors and endothelial cells that undergo endothelial-mesenchymal-transition (EndMT). This review will address the putative physiological and patho-physiological roles of ROS in controlling vascular cell fate and ROS contribution to vascular regeneration and disease progression

Moderate alcohol consumption targets S100β+ vascular stem cells and attenuates injury-induced neointimal hyperplasia

Background Stem cells present in the vessel wall may be triggered in response to injurious stimuli to undergo differentiation and contribute to vascular disease development. Our aim was to determine the effect of moderate alcohol (EtOH) exposure on the expansion and differentiation of S100 calcium-binding protein B positive (S100β+) resident vascular stem cells and their contribution to pathologic vessel remodeling in a mouse model of arteriosclerosis. Methods and Results Lineage tracing analysis of S100β+ cells was performed in male and female S100β-eGFP/Cre/ERT2–dTomato transgenic mice treated daily with or without EtOH by oral gavage (peak BAC: 15 mM or 0.07%) following left common carotid artery ligation for 14 days. Carotid arteries (ligated or sham-operated) were harvested for morphological analysis and confocal assessment of fluorescent-tagged S100 β + cells in FFPE carotid cross sections. Ligation-induced carotid remodeling was more robust in males than in females. EtOH-gavaged mice had less adventitial thickening and markedly reduced neointimal formation compared to controls, with a more pronounced inhibitory effect in males compared to females. There was significant expansion of S100β+-marked cells in vessels postligation, primarily in the neointimal compartment. EtOH treatment reduced the fraction of S100β+ cells in carotid cross sections, concomitant with attenuated remodeling. In vitro, EtOH attenuated Sonic Hedgehog-stimulated myogenic differentiation (as evidenced by reduced calponin and myosin heavy chain expression) of isolated murine S100β+ vascular stem cells. Conclusions These data highlight resident vascular S100β+ stem cells as a novel target population for alcohol and suggest that regulation of these progenitors in adult arteries, particularly in males, may be an important mechanism contributing to the antiatherogenic effects of moderate alcohol consumption

The calcium binding protein S100β marks hedgehog-responsive resident vascular stem cells within vascular lesions

A hallmark of subclinical atherosclerosis is the accumulation of vascular smooth muscle cell (SMC)-like cells leading to intimal thickening. While medial SMCs contribute, the participation of hedgehog-responsive resident vascular stem cells (vSCs) to lesion formation remains unclear. Using transgenic eGFP mice and genetic lineage tracing of S100β vSCs in vivo, we identified S100β/ Sca1 cells derived from a S100β non-SMC parent population within lesions that co-localise with smooth muscle α-actin (SMA) cells following iatrogenic flow restriction, an effect attenuated following hedgehog inhibition with the smoothened inhibitor, cyclopamine. In vitro, S100β/Sca1 cells isolated from atheroprone regions of the mouse aorta expressed hedgehog signalling components, acquired the di-methylation of histone 3 lysine 4 (H3K4me2) stable SMC epigenetic mark at the Myh11 locus and underwent myogenic differentiation in response to recombinant sonic hedgehog (SHh). Both S100β and PTCH1 cells were present in human vessels while S100β cells were enriched in arteriosclerotic lesions. Recombinant SHh promoted myogenic differentiation of human induced pluripotent stem cell-derived S100β neuroectoderm progenitors in vitro. We conclude that hedgehog-responsive S100β vSCs contribute to lesion formation and support targeting hedgehog signalling to treat subclinical arteriosclerosis

Oral abstracts 3: RA Treatment and outcomesO13. Validation of jadas in all subtypes of juvenile idiopathic arthritis in a clinical setting

Background: Juvenile Arthritis Disease Activity Score (JADAS) is a 4 variable composite disease activity (DA) score for JIA (including active 10, 27 or 71 joint count (AJC), physician global (PGA), parent/child global (PGE) and ESR). The validity of JADAS for all ILAR subtypes in the routine clinical setting is unknown. We investigated the construct validity of JADAS in the clinical setting in all subtypes of JIA through application to a prospective inception cohort of UK children presenting with new onset inflammatory arthritis. Methods: JADAS 10, 27 and 71 were determined for all children in the Childhood Arthritis Prospective Study (CAPS) with complete data available at baseline. Correlation of JADAS 10, 27 and 71 with single DA markers was determined for all subtypes. All correlations were calculated using Spearman's rank statistic. Results: 262/1238 visits had sufficient data for calculation of JADAS (1028 (83%) AJC, 744 (60%) PGA, 843 (68%) PGE and 459 (37%) ESR). Median age at disease onset was 6.0 years (IQR 2.6-10.4) and 64% were female. Correlation between JADAS 10, 27 and 71 approached 1 for all subtypes. Median JADAS 71 was 5.3 (IQR 2.2-10.1) with a significant difference between median JADAS scores between subtypes (p < 0.01). Correlation of JADAS 71 with each single marker of DA was moderate to high in the total cohort (see Table 1). Overall, correlation with AJC, PGA and PGE was moderate to high and correlation with ESR, limited JC, parental pain and CHAQ was low to moderate in the individual subtypes. Correlation coefficients in the extended oligoarticular, rheumatoid factor negative and enthesitis related subtypes were interpreted with caution in view of low numbers. Conclusions: This study adds to the body of evidence supporting the construct validity of JADAS. JADAS correlates with other measures of DA in all ILAR subtypes in the routine clinical setting. Given the high frequency of missing ESR data, it would be useful to assess the validity of JADAS without inclusion of the ESR. Disclosure statement: All authors have declared no conflicts of interest. Table 1Spearman's correlation between JADAS 71 and single markers DA by ILAR subtype ILAR Subtype Systemic onset JIA Persistent oligo JIA Extended oligo JIA Rheumatoid factor neg JIA Rheumatoid factor pos JIA Enthesitis related JIA Psoriatic JIA Undifferentiated JIA Unknown subtype Total cohort Number of children 23 111 12 57 7 9 19 7 17 262 AJC 0.54 0.67 0.53 0.75 0.53 0.34 0.59 0.81 0.37 0.59 PGA 0.63 0.69 0.25 0.73 0.14 0.05 0.50 0.83 0.56 0.64 PGE 0.51 0.68 0.83 0.61 0.41 0.69 0.71 0.9 0.48 0.61 ESR 0.28 0.31 0.35 0.4 0.6 0.85 0.43 0.7 0.5 0.53 Limited 71 JC 0.29 0.51 0.23 0.37 0.14 -0.12 0.4 0.81 0.45 0.41 Parental pain 0.23 0.62 0.03 0.57 0.41 0.69 0.7 0.79 0.42 0.53 Childhood health assessment questionnaire 0.25 0.57 -0.07 0.36 -0.47 0.84 0.37 0.8 0.66 0.4

‘Cruel and unusual punishment’: an inter-jurisdictional study of the criminalisation of young people with complex support needs

Although several criminologists and social scientists have drawn attention to the high rates of mental and cognitive disability amongst populations of young people embroiled in youth justice systems, less attention has been paid to the ways in which young people with disability are disproportionately exposed to processes of criminalisation and how the same processes serve to further disable them. In this paper, we aim to make a contribution towards filling this gap by drawing upon qualitative findings from the Comparative Youth Penality Project - an empirical inter-jurisdictional study of youth justice and penality in England and Wales and in four Australian states. We build on, integrate and extend theoretical perspectives from critical disability studies and from critical criminology to examine the presence of, and responses to, socio-economically disadvantaged young people with multiple disabilities (complex support needs) in youth justice systems in our selected jurisdictions. Four key findings emerge from our research pertaining to: (i) the criminalisation of disability and disadvantage; (ii) the management of children and young people with disabilities by youth justice agencies; (iii) the significance of early and holistic responses for children and young people with complex support needs; and (iv) the inadequate nature of community based support

Alcohol and Cardiovascular Disease—Modulation of Vascular Cell Function

Alcohol is a commonly used drug worldwide. Epidemiological studies have identified alcohol consumption as a factor that may either positively or negatively influence many diseases including cardiovascular disease, certain cancers and dementia. Often there seems to be a differential effect of various drinking patterns, with frequent moderate consumption of alcohol being salutary and binge drinking or chronic abuse being deleterious to one’s health. A better understanding of the cellular and molecular mechanisms mediating the many effects of alcohol consumption is beginning to emerge, as well as a clearer picture as to whether these effects are due to the direct actions of alcohol itself, or caused in part by its metabolites, e.g., acetaldehyde, or by incidental components present in the alcoholic beverage (e.g., polyphenols in red wine). This review will discuss evidence to date as to how alcohol (ethanol) might affect atherosclerosis that underlies cardiovascular and cerebrovascular disease, and the putative mechanisms involved, focusing on vascular endothelial and smooth muscle cell effects

Gastro-esophageal reflux disease to Barrett's

Thesis (Ph. D.)--University of Rochester. Department of Biology, 2014.Background: Esophageal Adenocarcinoma (EAC) affects about 18000 of the US population with annual mortality around 15000. Barrett’s esophagus (BE) is the single most important risk factor for the development of esophageal adenocarcinoma. The pathogenesis of BE is believed to be driven by protracted reflux of gastric and duodenal contents into the lower esophagus through an incompetent lower esophageal sphincter (LES) in patients with gastro-esophageal reflux disease (GERD), and as a result the esophageal stem cells are transcommitted to a columnar phenotype in the presence of the reflux environment. Bile acids, along with gastric acid at pH 2-4, are the chief components of reflux. Methods: We compared expression of squamous differentiation markers in biopsies taken from normal squamous esophagus and BE. We also treated primary esophageal cells in monolayer culture with bile acid cocktail and looked at the differentially expressed genes compared to control. In order to simulate gastro-esophageal reflux in vitro we developed a 3-D transwell culture model to grow primary esophageal cells in stratified culture. Subsequently, we used this model to demonstrate the morphological and molecular effects of simulated reflux condition on this stratified epithelium in vitro. Results: Our data indicate that a combination of bile acid and gastric acid at pH5 reduces squamous differentiation in primary esophageal cells as an initial step in EGFR-mediated dedifferentiation that enables mucosal repair in response to reflux injury. We have attributed the observed squamous dedifferentiation to the unionized unconjugated and thus intracellular bile acid pool. Our data demonstrate that bile acid at pH5, but not either alone, disrupts tight junction complexes and causes increased permeability of stratified squamous esophageal epithelium. These changes approximate the appearance of dilated intercellular spaces (DIS) similar to that found in GERD patients. These findings provide novel insights into the molecular mechanisms underlying the adaptive responses of the esophageal epithelium to reflux-mediated injury. Conclusion: To extend the relevance of this study in the clinical setting, acid suppression therapy likely will not prevent DIS formation and subsequent initiation of inflammatory response in esophageal epithelium as long as the patient is refluxing bile salts at a less acidic pH. In the majority of clinical situations, long term acid suppression in patients with GERD raises the gastric pH to 4-6. This facilitates the burgeoning of intestinal microflora and increases bacterial deconjugation of bile acid, causing the predominance of unconjugated bile acid refluxing into the esophagus at a pH around 5. Taking parallels from the in vitro results shown in this dissertation, we can conclude that the refluxing cocktail thus formed might be putting patients at risk to bile acid induced altered cellular differentiation