Differentiation methods for human induced pluripotent stem cells (hiPSCs) typically yield progeny from multiple tissue lineages, limiting their utility for drug testing and autologous cell transplantation. In particular, early retina and forebrain derivatives often intermingle in pluripotent stem cell cultures, owing to their shared ancestry and tightly coupled development. Here, we demonstrate that three-dimensional populations of retinal progenitor cells (RPCs) can be isolated from early forebrain populations in both human embryonic stem cell (hESC) and hiPSC cultures, providing a valuable tool for developmental, functional, and translational studies. Using our established protocol, we identified a transient population of optic vesicle-like (OV) structures that arose during a time period appropriate for normal human retinogenesis. These structures were independently cultured and analyzed to confirm their multipotent RPC status and capacity to produce physiologically responsive retinal cell types, including photoreceptors and retinal pigment epithelium (RPE). We then applied this method to hiPSCs derived from a patient with gyrate atrophy, a retinal degenerative disease affecting the RPE. RPE generated from these hiPSCs exhibited a disease-specific functional defect that could be corrected either by pharmacological means or following targeted gene repair. The production of OV-like populations from human pluripotent stem cells should facilitate the study of human retinal development and disease and advance the use of hiPSCs in personalized medicine.

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.

We demonstrate for the first time that human blood-derived iPSCs can generate retinal cell types, providing a highly convenient donor cell source for iPSC-based retinal studies. We also show that cultured TiPSC-OVs have the capacity to self-assemble into rudimentary neuroretinal structures and express markers indicative of chemical and electrical synapses.

Highly differentiated, pigmented hiPSC-RPE monolayers can undergo limited serial expansion while retaining key cytological and functional attributes. However, passaging hiPSC-RPE cultures beyond senescence leads to loss of such features. Our findings support limited, controlled passaging of patient-specific hiPSC-RPE to procure cells needed for in vitro disease modeling, drug screening, and cellular transplantation.

The prevalence and clinical consequences of gastro-oesophageal reflux disease (GERD) in chronic obstructive pulmonary disease (COPD) are not well characterised.The present study prospectively studied 42 males with COPD (forced expiratory volume in one second % predicted: 35%, range 20-49) and 16 healthy volunteers of similar age without respiratory or gastro-oesophageal symptoms. The diagnosis of GERD was confirmed using oesophageal 24 h pH monitoring. In the current study group, reflux symptoms were measured using the Vigneri score, cough and dyspnoea with the modified Medical Research Council questionnaire, and pulmonary function with bronchodilator response and health status using St George9s Respiratory Questionnaire.Pathological reflux was documented in 26 out of 42 patients (62%) and in three volunteers (19%). In patients with GERD, 15 patients (58%) did not report any reflux symptoms. There were no differences in symptoms, health status, bronchodilator treatment and pulmonary function test between patients with and without GERD. Oxygen desaturation coincided with episodes of increased oesophageal acidity in 40% of patients with GERD.Patients with severe chronic obstructive pulmonary disease have a high prevalence of asymptomatic gastro-oesophageal reflux. The association between this reflux and oxygen desaturation deserves further attention. Eur Respir J 2004; 23: 841-845.

Human induced pluripotent stem cells (hiPSCs) have been shown to differentiate along the retinal lineage in a manner that mimics normal mammalian development. Under certain culture conditions hiPSCs form optic vesicle-like structures (OVs), which contain proliferating progenitors capable of yielding all neural retina (NR) cell types over time. Such observations imply conserved roles for regulators of retinogenesis in hiPSC-derived cultures and the developing embryo. However, whether and to what extent this assumption holds true has remained largely uninvestigated. We examined the role of a key NR transcription factor, Visual System Homeobox 2 (VSX2), using hiPSCs derived from a patient with microphthalmia caused by an R200Q mutation in the VSX2 homeodomain region. No differences were noted between (R200Q)VSX2 and sibling control hiPSCs prior to OV generation. Thereafter, (R200Q)VSX2 hiPSC-OVs displayed a significant growth deficit compared to control hiPSC-OVs, as well as increased production of retinal pigmented epithelium (RPE) at the expense of NR cell derivatives. Furthermore, (R200Q)VSX2 hiPSC-OVs failed to produce bipolar cells, a distinctive feature previously observed in Vsx2 mutant mice. (R200Q)VSX2 hiPSC-OVs also demonstrated delayed photoreceptor maturation, which could be overcome via exogenous expression of wildtype VSX2 at early stages of retinal differentiation. Finally, RNAseq analysis on isolated hiPSC-OVs implicated key transcription factors and extracellular signaling pathways as potential downstream effectors of VSX2-mediated gene regulation. Our results establish hiPSC-OVs as versatile model systems to study retinal development at stages not previously accessible in humans, and support the bona fide nature of hiPSC-OV-derived retinal progeny.

BACKGROUND: Patient activation is linked to better health outcomes and lower rates of health service utilization. The role of patient activation in the rate of hospital readmission within 30 days of hospital discharge has not been examined. METHODS: A secondary analysis using data from the Project RED-LIT randomized controlled trial conducted at an urban safety net hospital. Data from 695 Englishspeaking general medical inpatient subjects were analyzed. We used an adapted, eight-item version of the validated Patient Activation Measure (PAM). Total scores were categorized, according to standardized methods, as one of four PAM levels of activation: Level 1 (lowest activation) through Level 4 (highest activation). The primary outcome measure was total 30-day post-discharge hospital utilization, defined as total emergency department (ED) visits plus hospital readmissions including observation stays. Poisson regression was used to control for confounding. RESULTS: Of the 695 subjects, 67 (9.6 %) were PAM Level 1, 123 (17.7 %) were Level 2, 193 (27.8 %) were Level 3, and 312 (44.9 %) were Level 4. Compared with highly activated patients (PAM Level 4), a higher rate of 30-day post-discharge hospital utilization was observed for patients at lower levels of activation (PAM Level 1, incident rate ratio [IRR] 1.75, 95 % CI,1.18 to 2.60) and (PAM Level 2, IRR 1.50, 95 % CI 1.06 to 2.13). The rate of returning to the hospital among patients at PAM Level 3 was not statistically different than patients with PAM Level 4 (IRR 1.30, 95 % CI, 0.94 to 1.80). The rate ratio for PAM Level 1 was also higher compared with Level 4 for ED use alone (1.68(1.07 to 2.63)) and for hospital readmissions alone (1.93 [1.22 to 3.06]). CONCLUSION: Hospitalized adult medical patients in an urban academic safety net hospital with lower levels of Patient Activation had a higher rate of post-discharge 30-day hospital utilization.

Although several neurodevelopmental and psychiatric disorders can emerge during the preschool period, there are comparatively few instruments to assess executive control. Evidence for validity of the Shape School (K. A. Espy, 1997) was examined in a sample of 219 typically developing young children. There was good evidence for validity, as Shape School performance variables were interrelated and were associated to other criterion measures considered to measure aspects of executive control. Also suggesting validity, the Shape School variables varied as a function of whether the task demands (a) were executive, (b) required inhibition of a prepotent response or context-controlled selection among relevant stimulus-response sets, and (c) included unitary or concurrent processing. The Shape School may be an effective tool by which to measure executive control in young children who have atypical developmental patterns.