Investigations of Thioether Generation for S-D-Ribosyl-L-homocysteine Synthesis

Creation of the thioether linkage is a fundamental step for the chemical preparation of the bacterial quorum sensing molecule S-D-ribosyl-L-homocysteine (SRH). Although previous preparations of SRH and its analogues have been reported in the literature, they employ assorted methods with varied results. Therefore, a reassessment of the methodology used for synthetic preparation of the thioether bond in SRH-like molecules is here considered. This work examines four methods of thioether generation following two mechanisms, bimolecular nucleophilic substitution (SN2) and radical-initiated thiol-ene coupling, in an attempt to generate SRH in a more efficient and reproducible manner. Both mechanisms address key objectives for SRH preparation: consistent production of the target compound in an analytically pure form, synthesis from easily obtained commodity materials, and improved understanding of each variable involved in creating the thioether bond in this molecule. One application of the thiol-ene coupling reaction generates the SRH thioether at a trial scale with a much-improved yield (93% over 70%) when compared to the most successful reported method to date. Other observations include improved understanding of steric effects involved in both mechanisms, optimization of conditions, assessment of electrophile choice, and the synthesis of a diastereomer of SRH, which may be considered for use in competitive inhibition assays

A study of the effects of the neurokinin peptides on respiratory function in sheep

The respiratory responses, including changes in pulmonary resistance (RL) and dynamic compliance (Cdyn), to the neurokinin peptides substance P (SP), neurokinin A (NKA) and neurokinin B (NKB) were assessed in anaesthetised normal Suffolk-cross and conscious Texel-cross asthmatic sheepIn normal sheep (n=l 1) intravenous SP was a more potent bronchoconstrictor than NKA (n=9) and this was similar to findings in sheep with a naturally acquired airway allergy to Ascaris suum antigen (asthmatic) (n=5) where peptides were administered by inhalation. NKB (n=4) was assessed only in normal sheep and caused insignificant changes in bronchomotor tone. Intravenous SP and NKA, in normal anaesthetised sheep, caused a dose-dependent reduction in respiratory rate and this was similar for both peptides.The bronchomotor response to SP in normal sheep demonstrated age-related changes. In sheep under 6 months of age there was a pronounced bronchoconstriction, with a subsequent reduction in the response as animals approach maturity. In old sheep, aged approximately four years, there was minimal bronchomotor response, however, there was dose-dependent apnoea.The bronchomotor response to SP in anaesthetised normal sheep was significantly antagonised after pre-treatment with atropine (lmg/kg; n=6), hexamethonium (20mg/Kg; n=3) and the NK-1 antagonist CP 96,345 (0.1 and 0.5mg/Kg; n=5), but not by the HI receptor antagonist chlorpheniramine (2mg/Kg; n=5) or the neurokinin antagonist spantide (lOug/kg/min; n=3). The anti-asthma drug nedocromil sodium (0.1 and l.Omg/kg; n=4) had a variable effect on the response. In the isolated sheep trachealis muscle preparation the contractile effect of SP was inhibited by atropine (n=4) and the Ml receptor antagonist pirenzepine (n=8), with IC₅₀ values of 5.6xl0⁻⁸ and 5xl0⁻¹⁰ M respectively, while spantide (n=7) and the NK-2 receptor antagonist L-659,874 (n=6) were ineffective.In several normal sheep (n=10) intravenous SP consistently caused augmented breaths. Bilateral vagotomy (n=7) abolished, and cooling of the right cervical vagus (n=7), after section of the left vagus, to temperatures below 7° C significantly attenuated the bronchomotor response to SP in normal sheep.The conclusion of this study is that the order of potency for the bronchomotor effects of the neurokinins is similar to rabbits and pigs but different from that reported for most other species, including man. The mechanism of action of SP is largely indirect, involving activation of vagal reflex mechanisms and/or modulation of ganglionic neurotransmission and acetylcholine release from cholinergic nerve endings

Comparative transcriptomic profiling of myxomatous mitral valve disease in the Cavalier King Charles Spaniel

BACKGROUND: Almost all elderly dogs develop myxomatous mitral valve disease by the end of their life, but the cavalier King Charles spaniel (CKCS) has a heightened susceptibility, frequently resulting in death at a young age and suggesting that there is a genetic component to the condition in this breed. Transcriptional profiling can reveal the impact of genetic variation through differences in gene expression levels. The aim of this study was to determine whether expression patterns were different in mitral valves showing myxomatous degeneration from CKCS dogs compared to valves from non-CKCS dogs. RESULTS: Gene expression patterns in three groups of canine valves resulted in distinct separation of normal valves, diseased valves from CKCS and diseased valves from other breeds; the latter were more similar to the normal valves than were the valves from CKCS. Gene expression patterns in diseased valves from CKCS dogs were quite different from those in the valves from other dogs, both affected and normal. Patterns in all diseased valves (from CKCS and other breeds) were also somewhat different from normal non-diseased samples. Analysis of differentially expressed genes showed enrichment in GO terms relating to cardiac development and function and to calcium signalling canonical pathway in the genes down-regulated in the diseased valves from CKCS, compared to normal valves and to diseased valves from other breeds. F2 (prothrombin) (CKCS diseased valves compared to normal) and MEF2C pathway activation (CKCS diseased valves compared to non-CKCS diseased valves) had the strongest association with the gene changes. A large number of genes that were differentially expressed in the CKCS diseased valves compared with normal valves and diseased valves from other breeds were associated with cardiomyocytes including CASQ2, TNNI3 and RYR2. CONCLUSION: Transcriptomic profiling identified gene expression changes in CKCS diseased valves that were not present in age and disease severity-matched non-CKCS valves. These genes are associated with cardiomyocytes, coagulation and extra-cellular matrix remodelling. Identification of genes that vary in the CKCS will allow exploration of genetic variation to understand the aetiology of the disease in this breed, and ultimately development of breeding strategies to eliminate this disease from the breed

The Role of Transforming Growth Factor-β Signaling in Myxomatous Mitral Valve Degeneration

Mitral valve prolapse (MVP) due to myxomatous degeneration is one of the most important chronic degenerative cardiovascular diseases in people and dogs. It is a common cause of heart failure leading to significant morbidity and mortality in both species. Human MVP is usually classified into primary or non-syndromic, including Barlow’s Disease (BD), fibro-elastic deficiency (FED) and Filamin-A mutation, and secondary or syndromic forms (typically familial), such as Marfan syndrome (MFS), Ehlers-Danlos syndrome, and Loeys–Dietz syndrome. Despite different etiologies the diseased valves share pathological features consistent with myxomatous degeneration. To reflect this common pathology the condition is often called myxomatous mitral valve degeneration (disease) (MMVD) and this term is universally used to describe the analogous condition in the dog. MMVD in both species is characterized by leaflet thickening and deformity, disorganized extracellular matrix, increased transformation of the quiescent valve interstitial cell (qVICs) to an activated state (aVICs), also known as activated myofibroblasts. Significant alterations in these cellular activities contribute to the initiation and progression of MMVD due to the increased expression of transforming growth factor-β (TGF-β) superfamily cytokines and the dysregulation of the TGF-β signaling pathways. Further understanding the molecular mechanisms of MMVD is needed to identify pharmacological manipulation strategies of the signaling pathway that might regulate VIC differentiation and so control the disease onset and development. This review briefly summarizes current understanding of the histopathology, cellular activities, molecular mechanisms and pathogenesis of MMVD in dogs and humans, and in more detail reviews the evidence for the role of TGF-β

Myxomatous Degeneration of the Canine Mitral Valve: From Gross Changes to Molecular Events

Myxomatous mitral valve disease (MMVD) is the single most common acquired heart disease of the dog, but is also of emerging importance in human medicine, with some features of the disease shared between both species. There has been increased understanding of this disease in recent years, with most research aiming to elucidate the cellular and molecular events of disease pathogenesis. For gross and histological changes, much of our understanding is based on historical studies and there has been no comprehensive reappraisal of the pathology of MMVD. This paper reviews the gross, histological, ultrastructural, cellular and molecular changes in canine MMVD. (C) 2017 Elsevier Ltd. All rights reserved

Comparison of cellular changes in cavalier King Charles spaniel and mixed breed dogs with myxomatous mitral valve disease

Introduction: The aim of this study was to determine if there are differences in cellular changes in Cavalier King Charles spaniel (CKCS) myxomatous mitral valves compared to non-CKCS dogs. Animals: Cavalier King Charles spaniels (n = 6) and age-matched mixed breed (n = 6) with severe myxomatous mitral valve disease (MMVD), and normal mixed breed (n = 4) dogs. Materials and Methods: Immunohistochemistry staining and qualitative and quantitative analysis of mitral valves sections, examining for the presence of CD11c and CD45, vimentin, alpha smooth muscle actin (α-SMA) and embryonic smooth muscle myosin heavy chain (Smemb), von Willebrand factor and CD31 and Ki-67. Results: Vimentin positive cell numbers were increased in the MMVD dogs and distributed throughout the valve with greatest density close to the endothelium. There were no significant differences in cell marker expression for the two diseased groups, but cell numbers were significantly increased compared to controls for α-SMA (CKCS only) and Smemb (CKCS and mixed breed: p < 0.05). Alpha smooth muscle actin+ cells were primarily located at the valve edge, with Smemb+ cells similarly located, but also present throughout the valve stroma. A small number of cells close to the valve edge co-expressed α-SMA and Smemb. Endothelial von Willebrand factor expression was identified in all valves, with evidence of disrupted endothelium in the diseased, but was also found in diseased valve stroma. There was no staining for CD11c, CD45 or CD31 in any valve. Ki-67+ cells formed linear clusters at the leaflet tip and were sparsely distributed throughout both myxomatous valve groups. Conclusions: The cellular changes notes with advanced stage MMVD appear similar for CKCS when compared to mixed breed dogs

See, Like, Share, Remember: Adolescents’ Responses to Unhealthy-, Healthy- and Non-Food Advertising in Social Media

Media-saturated digital environments seek to influence social media users’ behaviour, including through marketing. The World Health Organization has identified food marketing, including advertising for unhealthy items, as detrimental to health, and in many countries, regulation restricts such marketing and advertising to younger children. Yet regulation rarely addresses adolescents and few studies have examined their responses to social media advertising. In two studies, we examined adolescents’ attention, memory and social responses to advertising posts, including interactions between product types and source of posts. We hypothesized adolescents would respond more positively to unhealthy food advertising compared to healthy food or non-food advertising, and more positively to ads shared by peers or celebrities than to ads shared by a brand. Outcomes measured were (1a) social responses (likelihood to ‘share’, attitude to peer); (1b) brand memory (recall, recognition) and (2) attention (eye-tracking fixation duration and count). Participants were 151 adolescent social media users (Study 1: n = 72; 13−14 years; M = 13.56 years, SD = 0.5; Study 2: n = 79, 13−17 years, M = 15.37 years, SD = 1.351). They viewed 36 fictitious Facebook profile feeds created to show age-typical content. In a 3 × 3 factorial design, each contained an advertising post that varied by content (healthy/unhealthy/non-food) and source (peer/celebrity/company). Generalised linear mixed models showed that advertisements for unhealthy food evoked significantly more positive responses, compared to non-food and healthy food, on 5 of 6 measures: adolescents were more likely to wish to ‘share’ unhealthy posts; rated peers more positively when they had unhealthy posts in their feeds; recalled and recognised a greater number of unhealthy food brands; and viewed unhealthy advertising posts for longer. Interactions with sources (peers, celebrities and companies) were more complex but also favoured unhealthy food advertising. Implications are that regulation of unhealthy food advertising should address adolescents and digital media