Tubedown-1 (Tbdn-1) suppression in oxygen-induced retinopathy and in retinopathy of prematurity

Purpose: Identification of unique proteins involved in retinopathy of prematurity (ROP) may facilitate new and more effective diagnostic tools and molecular-based treatments for ROP. Tubedown-1 (Tbdn-1), a novel homeostatic protein which copurifies with an acetyltransferase activity, is expressed in normal retinal endothelium and is specifically suppressed in retinal endothelial cells from patients with proliferative diabetic retinopathy. Furthermore, recent in vivo knockdown studies in mice have revealed that Tbdn-1 is important for retinal blood vessel homeostasis and for preventing retinal neovascularization in adults. The purpose of the present study was to determine if the expression pattern of Tbdn-1 is altered during oxygen-induced retinal neovascularization in mice and in a specimen of stage 3 human ROP. Methods: Specimens of oxygen-induced retinal neovascularization in mice, and a single specimen of active stage 3 ROP were studied by immunohistochemistry and digital image analysis using antibodies raised against Tbdn-1 and other blood vessel markers. Results: The pattern of Tbdn-1 expression during the course of oxygen-induced retinal neovascularization in mice suggests a regulating role in neonatal retinopathy. Retinal lesions from oxygen-induced retinal neovascularization in mice display suppression of retinal endothelial Tbdn-1 protein expression in conjunction with an increase in expression of proliferating cell nuclear antigen (a marker of proliferation) and α smooth muscle actin (a marker of myofibroblastic cells). Abnormal blood vessels within vitreoretinal neovascular lesions in a human specimen of active stage 3 ROP did not show Tbdn-1 protein expression. Conclusions: These results suggest that the loss of retinal endothelial Tbdn-1 expression may be a contributing factor in retinal blood vessel proliferation in ROP

Osmotic pressure-adaptive responses in the eye tissues of rainbow smelt (Osmerus mordax)

Purpose: The rainbow smelt (Osmerus mordax), is a teleost fish, which avoids freezing by becoming virtually isosmotic with seawater. The effects that such massive changes in osmolarity have on both its visual system and its highly evolved and specialized circulation are not known. New knowledge about the osmotic adaptation of the rainbow smelt eye is highly relevant to the adaptation and survival of this species and to its ability to feed as a visual predator in the face of environmental pressures. Moreover, the molecular physiologic response of the smelt to osmotic stress might provide valuable insights into understanding and managing mammalian pathological hyperosmolarity conditions, such as diabetes. We undertook the present study to provide an initial assessment of gene expression in ocular vasculature during osmotic adaptation in rainbow smelt. Methods: Immunohistochemistry with species cross reactive antibodies was used to assess blood vessel protein expression in paraffin sections. Western blotting was used to further verify antibody specificity for orthologs of mammalian blood vessel proteins in rainbow smelt. Thermal hysteresis and the analysis of glycerol concentrations in vitreous fluid were used to assess the physiologic adaptive properties of cold stressed eyes. Results: Glycerol levels and osmotic pressure were significantly increased in the vitreal fluid of smelt maintained at <0.5 °C versus those maintained at 8–10 °C. Compared to the 8–10 °C adapted specimens, the rete mirabile blood vessels and connecting regions of the endothelial linings of the choroidal vessels of the <0.5 °C adapted specimens showed a higher expression level of Tubedown (Tbdn) protein, a marker of the endothelial transcellular permeability pathway. Expression of the zonula occludens protein ZO-1, a marker of the endothelial paracellular permeability pathway showed a reciprocal expression pattern and was downregulated in rete mirabile blood vessels and connecting regions in the endothelial linings of choroidal vessels in <0.5 °C adapted specimens. Smelt orthologs of the mammalian Tbdn and zoluna occludens protein 1 (ZO-1) proteins were also detected by western blotting using anti-mammalian antibodies raised against the same epitopes as those used for immunohistochemistry. Conclusions: This work provides the first evidence that molecules known to play a role in ocular vascular homeostasis are expressed and may be differentially regulated during anti-freezing cold adaptation in smelt eyes. We propose a hypothesis that in a state of cold-induced hyperosmolarity, changes in ZO-1 expression are associated with the passage of small solutes from the plasma space to ocular fluid, while changes in Tbdn expression regulate the passage of proteins between the ocular fluid and plasma space. This work also provides fundamental insight into the mechanisms underlying the adaptation of the blood-retinal barrier to metabolically relevant compounds such as glycerol

Controlled cell proliferation on an electrochemically engineered collagen scaffold

Therapies for corneal disease and injury often rely on artificial implants, but integrating cells into synthetic corneal materials remains a significant challenge. The electrochemically formed collagen-based matrix presented here is non-toxic to cells and controls the proliferation in the corneal fibroblasts seeded onto it. Histology and biomolecular studies show a behavior similar to corneal stromal cells in a native corneal environment. Not only is this result an important first step toward developing a more realistic, multi-component artificial cornea, but it also opens possibilities for using this matrix to control and contain the growth of cells in engineered tissues

The catalytic domain of pp56(lck), but not its regulatory domain, is sufficient for inducing IL-2 production

The lymphoid src kinase pp56lck has been shown to be essential for the induction of different T lymphocyte responses, including CD4-mediated enhancement of Ag-induced T cell activation, early T cell differentiation, induction of IL-2 production, and cytotoxicity. It is assumed that pp56lck acts on these processes by phosphorylating substrates. However, it has been recently reported that the NH2 regulatory domain is sufficient to mediate CD4 accessory function. In this report we address the contribution of the regulatory and catalytic domains of pp56lck to another function of this enzyme independent of CD4: TCR-induced IL-2 production. Two pp56lck mutants lacking either the entire catalytic domain or the entire NH2 regulatory domain were generated, and their abilities to trigger transactivation of the TCR-regulated nuclear factor of activated T cells (NF-AT) region of the IL-2 promoter were compared. Only the catalytic, but not the NH2 regulatory, domain of pp56lck was able to induce NF-AT region transactivation on its own and to cooperate with other intracellular signals to trigger this response. Moreover, the catalytic domain of pp56lck was able to induce IL-2 cytokine production to an extent similar to that of wild-type pp56lck. We conclude that different domains of the pp56lck molecule contribute to regulate distinct biologic functions. In fact, while the NH2 regulatory domain is sufficient to mediate CD4 accessory function, we show here that the catalytic domain of pp56lck is sufficient for induction of IL-2 production, mimicking TCR ligation.This work was supported by the CICyT, Pharmacia, Consejo Superiro de Investigaciones Científicas, and the José Carreras International Foundation.Peer Reviewe

Development and initial factor validation of the Violence Toward Athletes Questionnaire (VTAQ) in a sample of young athletes

This study sought to develop and validate a questionnaire about violence experienced by children in sport. A convenience sample of 1055 French-Canadian athletes between 14 and 17 years old was recruited to participate in an online study assessing their experiences of interpersonal violence in sport. The Violence Toward Athletes Questionnaire (VTAQ) includes three subscales: athlete version (VTAQ-A), coach version (VTAQ-C), and parent version (VTAQ-P). Exploratory structural equation modeling (ESEM) was used to identify latent factors underlying versions of the VTAQ. The VTAQ-Athlete includes nine items with three factors: psychological (4 items), physical (2 items) and sexual (3 items). The VTAQ-Coach includes 36 items with three factors: psychological/neglect (16 items), physical (9 items) and sexual (11 items). The VTAQ-Parent includes 25 items with two factors: psychological/neglect (17 items) and physical (8 items). The VTAQ provides initial validation of the first measure by questioning children directly about their experiences of interpersonal violence in sport.<br/

Symptoms and quality of life in patients with chronic obstructive pulmonary disease treated with aclidinium in a real-life setting

Introduction: Chronic obstructive pulmonary disease (COPD) is a progressive disease with symptoms that can have a major impact on patients’ physical health. The aim of this study was to evaluate quality of life (QoL), symptom severity and dyspnoea in COPD patients treated with aclidinium up to 24 weeks. Methods: In this prospective non-interventional multicentre study (198 centres in Sweden, Denmark, and Norway), COPD patients (age ≥40 years) who started treatment with aclidinium (initial therapy, change of treatment, or add-on therapy) could be included. Health-related QoL was obtained by COPD assessment test (CAT). Symptoms were evaluated on a 6-point Likert scale. The modified Medical Research Council (mMRC) Dyspnoea Scale was used as a simple grading system to assess the level of dyspnoea/shortness of breath from0 to 4. Patients on treatment with aclidinium who completed baseline and at least one follow-up visit (week 12 or 24) were included in the study population. Results: Overall, 1,093 patients were enrolled (mean 69 years, 54% females), one-third had ≥1 exacerbation the year prior to baseline. At enrolment, 48% were LAMA naïve. Mean (standard deviation, SD) CAT score decreased from 16.9 (7.7) at baseline to 14.3 (7.3) at week 24 (p<0.01) with a decrease in all individual CAT items (p<0.05). Mean difference in morning and night-time symptoms from baseline to week 24 was −0.60 (SD 2.51) and −0.44 (SD 2.48), respectively (both p<0.001). Mean (SD) mMRC Dyspnoea Scale changed from 1.6 (1.0) at baseline to 1.5 (1.0) at week 24 (p<0.001). Conclusion: In this observational study of a Nordic real-life COPD population, treatment with aclidinium was associated with a clinically important improvement in QoL and morning and night-time symptoms, most pronounced in the LAMA naïve group. However, there is still room for improvement in the management of symptomatic COPD patients