RESUMO
Best disease (BD) is an inherited degenerative disease of the human macula that results in progressive and irreversible central vision loss. It is caused by mutations in the retinal pigment epithelium (RPE) gene BESTROPHIN1 (BEST1), which, through mechanism(s) that remain unclear, lead to the accumulation of subretinal fluid and autofluorescent waste products from shed photoreceptor outer segments (POSs). We employed human iPS cell (hiPSC) technology to generate RPE from BD patients and unaffected siblings in order to examine the cellular and molecular processes underlying this disease. Consistent with the clinical phenotype of BD, RPE from mutant hiPSCs displayed disrupted fluid flux and increased accrual of autofluorescent material after long-term POS feeding when compared with hiPSC-RPE from unaffected siblings. On a molecular level, RHODOPSIN degradation after POS feeding was delayed in BD hiPSC-RPE relative to unaffected sibling hiPSC-RPE, directly implicating impaired POS handling in the pathophysiology of the disease. In addition, stimulated calcium responses differed between BD and normal sibling hiPSC-RPE, as did oxidative stress levels after chronic POS feeding. Subcellular localization, fractionation and co-immunoprecipitation experiments in hiPSC-RPE and human prenatal RPE further linked BEST1 to the regulation and release of endoplasmic reticulum calcium stores. Since calcium signaling and oxidative stress are critical regulators of fluid flow and protein degradation, these findings likely contribute to the clinical picture of BD. In a larger context, this report demonstrates the potential to use patient-specific hiPSCs to model and study maculopathies, an important class of blinding disorders in humans.
Assuntos
Células-Tronco Pluripotentes Induzidas/citologia , Distrofia Macular Viteliforme/genética , Distrofia Macular Viteliforme/fisiopatologia , Animais , Bestrofinas , Cálcio/metabolismo , Bovinos , Diferenciação Celular , Linhagem Celular , Canais de Cloreto/genética , Canais de Cloreto/metabolismo , Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Regulação da Expressão Gênica , Homeostase , Humanos , Imuno-Histoquímica , Imunoprecipitação , Macula Lutea/patologia , Microscopia Eletrônica de Transmissão , Estresse Oxidativo , Fagocitose , Segmento Externo das Células Fotorreceptoras da Retina/metabolismo , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/patologia , Distrofia Macular Viteliforme/metabolismoRESUMO
Today the majority of people uses online social networks not only to stay in contact with friends, but also to find information about relevant topics, or to spread information. While a lot of research has been conducted into opinion formation, only little is known about which factors influence whether a user of online social networks disseminates information or not. To answer this question, we created an agent-based model and simulated message spreading in social networks using a latent-process model. In our model, we varied four different content types, six different network types, and we varied between a model that includes a personality model for its agents and one that did not. We found that the network type has only a weak influence on the distribution of content, whereas the message type has a clear influence on how many users receive a message. Using a personality model helped achieved more realistic outcomes.
RESUMO
PURPOSE: Motivational interviewing (MI) is an evidence-based technique that enables clinicians to help patients modify health behaviors. Although MI is an essential tool for physician assistants (PAs), the extent to which it is addressed in PA curricula in the United States is unknown. This study is a comprehensive description of MI education in PA programs in the United States. METHODS: Data are from the 2014 Physician Assistant Education Association Annual Program Survey. Descriptive statistics were conducted on de-identified data from all 186 PA programs in the United States. RESULTS: Of the 186 PA programs surveyed, 72.58% (n = 135) reported at least one course providing MI training. Availability of courses providing training in skills essential to the MI process varied. Having a course with verbal communication training was most frequently endorsed, and having a course with training in developing discrepancy was least frequently endorsed. The most popular teaching modality was lecture (84.95%, n = 158), whereas only 41.40% (n = 77) and 58.60% (n = 109) reported role play with evaluation and standardized patient exercises with evaluation, respectively. CONCLUSIONS: More than 70% of programs included at least one course in their curriculum that provided training in MI, suggesting that PA programs recognize the importance of MI. Instruction in change talk was not provided in nearly half of the programs. Role-play and standardized patient exercises with evaluation were underused methods despite their proven efficacy in MI education. As the first comprehensive benchmark of MI education for PAs, this study shows that although most programs address MI, opportunities exist to improve MI training in PA programs in the United States.
Assuntos
Currículo/estatística & dados numéricos , Entrevista Motivacional/métodos , Assistentes Médicos/educação , Comunicação , Empatia , Comportamentos Relacionados com a Saúde , Humanos , AutoeficáciaRESUMO
Oxytocin (OXT) is a neuropeptide that activates the oxytocin receptor (OXTR), a rhodopsin family G-protein coupled receptor. Our localization of OXTR to the retinal pigment epithelium (RPE), in close proximity to OXT in the adjacent photoreceptor neurons, leads us to propose that OXT plays an important role in RPE-retinal communication. An increase of RPE [Ca2+]i in response to OXT stimulation implies that the RPE may utilize oxytocinergic signaling as a mechanism by which it accomplishes some of its many roles. In this study, we used an established human RPE cell line, a HEK293 heterologous OXTR expression system, and pharmacological inhibitors of Ca2+ signaling to demonstrate that OXTR utilizes capacitative Ca2+ entry (CCE) mechanisms to sustain an increase in cytoplasmic Ca2+. These findings demonstrate how multiple functional outcomes of OXT-OXTR signaling could be integrated via a single pathway. In addition, the activated OXTR was able to inhibit the Kir7.1 channel, an important mediator of sub retinal waste transport and K+ homeostasis.
Assuntos
Cálcio/metabolismo , Ocitocina/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Receptores de Ocitocina/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Células Cultivadas , Células HEK293 , Humanos , Camundongos Endogâmicos C57BLRESUMO
PURPOSE: Oxytocin (OXT) is recognized as an ubiquitously acting nonapeptide hormone that is involved in processes ranging from parturition to neural development. Its effects are mediated by cell signaling that occurs as a result of oxytocin receptor (OXTR) activation. We sought to determine whether the OXT-OXTR signaling pathway is also expressed within the retina. METHODS: Immunohistochemistry using cell-specific markers was used to localize OXT within the rhesus retina. Reverse transcriptase PCR and immunohistochemistry were used to assess the expression of OXTR in both human and rhesus retina. Single-cell RT-PCR and Western blot analyses were used to determine the expression of OXTR in cultured human fetal RPE (hfRPE) cells. Human fetal RPE cells loaded with FURA-2 AM were studied by ratiometric Ca(2+) imaging to assess transient mobilization of intracellular Ca(2+) ([Ca(2+)]i). RESULTS: Oxytocin was expressed in the cone photoreceptor extracellular matrix of the rhesus retina. Oxytocin mRNA and protein were expressed in the human and rhesus RPE. Oxytocin mRNA and protein expression were observed in cultured hfRPE cells, and exposure of these cells to 100 nM OXT induced a transient 79 ± 1.5 nM increase of [Ca(2+)]i. CONCLUSIONS: Oxytocin and OXTR are present in the posterior retina, and OXT induces an increase in hfRPE [Ca(2+)]i. These results suggest that the OXT-OXTR signaling pathway is active in the retina. We propose that OXT activation of the OXTR occurs in the posterior retina and that this may serve as a paracrine signaling pathway that contributes to communication between the cone photoreceptor and the RPE.