Electrical estimulation of retinal pigment epithelial cells

We investigated and characterized the effect of externally applied electric fields (EF) on retinal pigment epithelial (RPE) cells by exposing primary cultures of human RPE cells (hRPE) and those from the ARPE19 immortalized cell line to various strengths of EF (EF-treated cells) or to no EF (control cells) under different conditions including presence or absence of serum and gelatin and following wounding. We evaluated changes in RPE cell behavior in response to EF by using a computer based image capture and analysis system (Metamorph). We found that RPE cells responded to externally applied EFs by preferential orientation perpendicular to the EF vector, directed migration towards the anode, and faster translocation rate than control, untreated cells. These responses were voltage-dependent. Responses were observed even at low voltages, of 50-300 mV. Furthermore, the migration of hRPE cell sheets generated by wounding of confluent monolayers of cells at early and late confluence could be manipulated by the application of EF, with directed migration towards the anode observed at both sides of the wounded hRPE. In conclusion, RPE cell behaviour can be controlled by an externally applied EF. The potential for externally applied EF to be used as a therapeutic strategy in the management of selected retinal diseases warrants further investigation. © 2010 Elsevier Ltd.

Toxicity of mildly modified low-density lipoproteins to cultured retinal capillary endothelial cells and pericytes

To investigate the role of modified low-density lipoproteins (LDL) in the pathogenesis of diabetic retinopathy, we studied the cytotoxicity of normal and mildly modified human LDL to bovine retinal capillary endothelial cells and pericytes in vitro. Pooled LDL was incubated (in phosphate-buffered saline-EDTA, 3 days, 37 degrees C) under 1) nitrogen with additional chelating agents and 2) air, to prepare normal and minimally oxidized LDL, respectively. Similar conditions, but with the addition of 50 mM D-glucose, were used to prepare glycated and glycoxidized LDL. None of the LDL preparations was recognized by the macrophage scavenger receptor, confirming limited modification. Retinal capillary endothelial cells and pericytes were grown to confluence and then exposed for 2 or 3 days to serum-free medium (1% albumin) supplemented with normal or modified LDL (100 mg/l) or to serum-free medium alone. Cytotoxicity was assessed by cell counting (live and total cells) and by cell protein determination. Compared with normal LDL, modified LDL were cytotoxic to both cell types at both time points, causing highly significant decreases in live and total cell counts (P <0.001) (analysis of variance). Reductions in cell protein also were significant for pericytes at day 3 (P = 0.016) and of borderline significance for endothelial cells at day 2 (P = 0.05) and day 3 (P = 0.063). Cytotoxicity increased as follows: normal <glycated <or = minimally oxidized <glycoxidized LDL. We conclude that, in diabetes, mild modification of LDL resulting from separate or combined processes of glycation and oxidation may contribute to chronic retinal capillary injury and thus to the development of diabetic retinopathy.

The influence of parents, older siblings, and non-parental care on infant development at nine months of age

Background: The majority of research examining the influence of social environment on early child development suggests benefits to two-parent households, but contradictory evidence for the effects of siblings. The aims of the present study were to examine the influence of the child's proximal social environment, and the effects of interactions between socioeconomic status and social environment on developmental outcomes.

Methods: Primary caregivers of a representative sample of 10,748 nine-month-old infants in Ireland completed the Ages and Stages Questionnaire and provided information on social environment. Adjustment was made for infant and maternal characteristics, household income, and area where the child was living at the time of the study. Further analyses tested for interactions between social environment and household income.

Results: Binary logistic regressions indicated no effects for number of parents in the household. However, the presence of siblings in the household was a consistent predictor of failing to reach milestones in communication, gross motor, problem-solving, and personal-social development. Furthermore, there was a gradient of increasing likelihood of failing in gross motor, problem-solving, and personal-social development with increasing numbers of siblings. Care by a grandparent decreased the likelihood of failing in communication and personal-social development.

Conclusions:These findings do not support the majority of research that finds positive benefits for two-parent households. Similarly, the findings suggest limited effects for non-parental care. However, the observed negative effects of siblings support both the confluence and resource dilution models of sibling effect. Examination of follow-up data may elucidate current findings.

Decadent Conservatism: Politics and Aesthetics in The Senate

The relationship between late-Victorian Decadence and Aestheticism and politics has long been vexed. This article explores the hitherto under-explored confluence of conservatism and avant-garde literature in the period by introducing The Senate, a Tory-Decadent journal that ran from 1894-7. While Decadent authors occupied various political positions, this article argues that The Senate offers a crucial link between conservatism and Decadence The article presents the journal in its political and publishing context, outlining its editorial position on such issues as the Liberal Unionist-Conservative coalition governments, Britain's relationship with Europe and the threat of ‘State Socialism’, as well as its valorisation of Bollingbroke and eighteenth-century Toryism, and its relationship to, and difference from, key Decadent journals the Yellow Book and The Savoy. It then goes on to articulate its relationship to Decadence by focussing on the presence of Paul Verlaine in its pages and its vitriolic response to the press coverage of Oscar Wilde's trials. The article concludes by exploring the surprising wake of The Senate, briefly tracing the editors' influence in the development of Modernism and links with the journal BLAST.

'Regina v John Terry': The Discursive Construction of an Alleged Racist Event

This article explores the conformation in discourse of a verbal exchange and its subsequent mediatised and legal ramifications. The event concerns an allegedly racist insult directed by high profile English professional footballer John Terry towards another player, Anton Ferdinand, during a televised match in October 2011. The substance of Terry’s utterance, which included the noun phrase ‘fucking black cunt’, was found by a Chief Magistrate not to be a racist insult, although the fact that these actual words were framed within the utterance was not in dispute. The upshot of this ruling was that Terry was acquitted of a racially aggravated public order offence. A subsequent investigation by the regulatory commission of the English Football Association (FA) ruled, almost a year after the event, that Terry was guilty of racially abusing Ferdinand. Terry was banned for four matches and fined £220,000.

It is our contention that this event, played out in legal rulings, social media and print and broadcast media, constitutes a complex web of linguistic structures and strategies in discourse, and as such lends itself well to analysis with a broad range of tools from pragmatics, discourse analysis and cognitive linguistics. Amongst other things, such an analysis can help explain the seemingly anomalous - even contradictory - position adopted in the legal ruling with regard to the speech act status of ‘fucking black cunt’; namely, that the racist content of the utterance was not contested but that the speaker was found not to have issued a racist insult. Over its course, the article addresses this broader issue by making reference to the systemic-functional interpersonal function of language, particularly to the concepts of modality, polarity and modalisation. It also draws on models of verbal irony from linguistic pragmatics, notably from the theory of irony as echoic mention (c.f. Sperber and Wilson, 1981; Wilson and Sperber, 1992). Furthermore, the article makes use of the cognitive-linguistic framework, Text World Theory (c.f. Gavins, 2007; Werth, 1999) to examine the discourse positions occupied by key actors and adapts, from cognitive poetics, the theory of mind-modelling (c.f. Stockwell, 2009) to explore the conceptual means through which these actors discursively negotiate the event.

It is argued that the pragmatic and cognitive strategies that frame the entire incident go a long way towards mitigating the impact of so ostensibly stark an act of racial abuse. Moreover, it is suggested here that the reconciliation of Terry’s action was a result of the confluence of strategies of discourse with relations of power as embodied by the media, the law and perceptions of nationhood embraced by contemporary football culture. It is further proposed that the outcome of this episode, where the FA was put in the spotlight, and where both the conflict and its key antagonists were ‘intranational’, was strongly impelled by the institution of English football and its governing body both to reproduce and maintain social, cultural and ethnic cohesion and to avoid any sense that the event featured a discernable ‘out-group’.

Osteoprotegerin Expression in the Dental Pulp

Objectives: The inflammatory response to pulpal injury or infection has major clinical significance. Osteoprotegerin (OPG) is a soluble decoy receptor for Receptor Activator of NF kappa B Ligand (RANKL), preventing ligand binding to its receptor (RANK), thus inhibiting clastic cell formation. The aim of the study is to investigate the expression of OPG in human dental pulp and the effects of inflammatory mediators. This study will specifically investigate the effects of Transforming Growth Factor Beta-1 (TGF-β1) and Interleukin 1-Beta (IL-1β) on the expression of OPG on pulp fibroblasts in vitro. Method: Five primary pulp fibroblast populations were obtained by explant culture of healthy pulp tissue. Triplicate cultures were grown to confluence in 12-well plates and stimulated for 48 hours with IL-1β (10ng/ml) or TGF-β1 (10ng/ml). The conditioned media was collected and OPG levels detected by ELISA (R+D Systems, UK). Results: All fibroblast populations produced quantifiable levels of OPG in a time-dependant fashion. IL-1β significantly increased the expression of OPG (p<0.05) in all cultures. In contrast, TGF-β1 had no significant effect on OPG expression levels. In addition, previous work in our laboratory demonstrated both TGF-β1 and IL-1β stimulated OPG expression by periodontal ligament fibroblasts. Conclusion: These data indicate that IL-1β-regulated expression of OPG by pulpal fibroblasts may mediate hard tissue turnover in the inflamed dental pulp.

Osteoprotegerin expression by periodontal ligament fibroblasts

Objectives: Receptor Activator of NF-kappaB ligand (RANKL), through binding to its receptor (RANK), plays an important role in osteoclast differentiation and activation. Conversely, osteoprotegerin (OPG), a decoy receptor for RANKL, inhibits osteoclastogenesis and subsequent bone turnover. Little is known about the role of resident periodontal ligament fibroblasts in regulating bone turnover. The aim of this study was to determine (i) if periodontal ligament fibroblasts produced OPG in vitro and (ii) the effects of IL-1b and TGF-b1 on OPG expression. Methods: Three human periodontal ligament fibroblast populations, developed by explant culture, were grown to confluence in 6-well plates in DMEM supplemented with 10% FCS. Cells were washed in HBSS and then cultured for an additional 48 hours in serum-free media supplemented with IL-1b or TGF-b1 at 10ng/ml. OPG expression levels in the conditioned medium were determined by ELISA (R&D Systems, UK) and confirmed by Western blot. Results: All three fibroblast strains produced quantifiable levels of OPG. Both IL-1b and, to a lesser extent, TGF-b1 significantly stimulated OPG expression in all fibroblast strains (p<0.05). Pre-incubation of samples with N-glycosidase F prior to Western blots indicated glycosylation of expressed OPG. Conclusions: These data indicate that periodontal ligament fibroblasts can regulate osteoclast activation via the RANK/RANKL signalling pathway. These fibroblasts may play an important role in regulating bone turnover both in periodontal disease and orthodontic tooth movement.

Neuropeptide Receptors in the Dental Pulp

Objectives: The inflammatory response to pulpal injury or infection has major clinical significance. The aim of the study is to investigate the presence and regulation of expression of neuropeptide receptors on human pulp fibroblasts and whole pulp tissue. This study will investigate the expression of Substance P (NK-1) and Neuropeptide Y (NPY-Y1) receptors on pulp fibroblasts, determine the effects of Transforming Growth Factor Beta-1 (TGF-b1) and Interleukin 1-Beta (IL-1b) on the expression of NK-1 and NPY-Y1 receptors on pulp fibroblasts and examine the levels of receptor expression in whole pulp samples. Methods: Primary pulp fibroblast cell lines were obtained from patients undergoing extractions for orthodontic reasons. The cells were grown to confluence and stimulated for 5 days with IL-1b or TGF-b1. Pulp tissue fragments were obtained from freshly extracted sound and carious teeth, snap frozen in liquid nitrogen and cracked open using a vice. The monolayer was removed with cell scrapers and pelleted. The cell membranes of the cultured cells and the whole tissue were isolated using a Mem-PER® Eukaryotic Membrane Protein Extraction Reagent Kit (Pierce, UK). The membrane proteins were separated by SDS-PAGE and Western blotting was used to detect the presence of NK-1 and NPY-Y1. Results: Initial results demonstrated the presence of NK-1 and NPY-Y1 in cultured pulp fibroblasts. Following the 5 day incubation with TGF-b1, the cells appeared not to express NK-1. IL-1b had a slight stimulatory effect on NK-1 expression. The NPY-Y1 expression was not affected by either TGF-b1 or IL-1b. In whole pulp samples, levels of NK-1 were increased in carious teeth compared to caries-free teeth. The NPY-Y1 levels were similar in carious and non-carious teeth. Conclusion: These findings give an insight into how pulp cells react to inflammatory stimuli with regards to neuropeptide receptor expression and their roles in health and disease