Growth kinetics and transplantation of human retinal progenitor cells.

We studied the growth kinetics of human retinal progenitor cells (hRPCs) isolated from donor tissue of different gestational ages (G.A.), determined whether hRPCs can be differentiated into mature photoreceptors and assessed their ability to integrate with degenerating host retina upon transplantation. Eyes (12-18 weeks G.A.) were obtained with IRB approval and retinas were enzymatically dissociated. Cells were expanded in vitro, counted at isolation and at each passage, and characterized using immunocytochemistry and PCR. GFP positive hRPCs were co-cultured with retinal explants from rd1 and rhodopsin -/- mice, or transplanted into B6 mice with retinal photocoagulation and rhodopsin -/- mice. Eyes were harvested for histological evaluation following transplantation. Our results show that hRPCs from 16 to 18 weeks G.A. had the longest survival in vitro and yielded the maximum number of cells, proliferating over at least 6 passages. These cells expressed the retinal stem cell markers nestin, Ki-67, PAX6 and Lhx2, and stained positively for photoreceptor markers upon differentiation with serum. Some of the GFP positive cells used for transplantation studies showed evidence of migration into the degenerative host retina and expressed rhodopsin. In conclusion, we have determined the growth kinetics of hRPCs and have shown that cells from donor tissue of 16-18 weeks G.A. exhibit the best proliferative dynamics under the specified conditions, and that hRPCs can also be differentiated along the photoreceptor lineage. Further, we have also demonstrated that following transplantation, some of these cells integrate within the host retina and differentiate to express rhodopsin, thereby supporting the potential utility of hRPC transplantation in the setting of retinal degenerative disorders.

Molecular characterization of human retinal progenitor cells.

PURPOSE: To examine the molecular profile of fetal human retinal progenitor cells (hRPCs) expanded in vitro and those grown in a co-culture system with mouse retina through the analysis of protein and gene expression and neurotransmitter-stimulated calcium dynamics. METHODS: hRPCS were isolated from human retina of 14 to 18 weeks gestational age (GA) and expanded in vitro. Immunoblot, microarray, and immunocytochemistry (ICC) assays were performed on undifferentiated hRPCs and those co-cultured with mouse retinas for 2 weeks. Cell function was assessed by using calcium imaging. RESULTS: The ICC results showed a gradual decrease in the percentages of KI67-, SOX2-, and vimentin-positive cells from passages (P) 1 to P6, whereas a sustained expression of nestin and PAX6 was observed through P6. Microarray analysis of P1 hRPCs showed the expression of early retinal developmental genes: VIM (vimentin), KI67, NES (nestin), PAX6, SOX2, HES5, GNL3, OTX2, DACH1, SIX6, and CHX10 (VSX2). At P6, hRPCs continued to express VIM, KI67, NES, PAX6, SOX2, GNL3, and SIX6. On co-culture, there was a significant increase in the expression of MKI67, PAX6, SOX2, GNL3, SIX3, and RHO (rhodopsin). Calcium imaging showed a functional response to excitatory neurotransmitters. CONCLUSIONS: Fetal-derived hRPCs show molecular characteristics indicative of a retinal progenitor state up to P6 (latest passage studied). They show a progressive decrease in the expression of immature markers as they reach P6. These cells are functional, respond to excitatory neurotransmitters, and exhibit changes in expression patterns in response to co-culture with mouse retina.

Drug compliance after stroke and myocardial infarction: a comparative study.

BACKGROUND: Stroke and myocardial infarction (MI) are both life-threatening diseases of vascular origin with a tendency to recur. In both conditions, risk of recurrence is reduced through similar drug regimens. AIM: To determine if compliance with prescribed medication after stroke or MI was similar in the two populations. SETTING AND DESIGN: Retrospective data collection and cross-sectional telephonic survey of patients discharged from a single academic medical center. MATERIALS AND METHODS: Adult patients consecutively discharged over a two-year period with a diagnosis of first-ever stroke (ischemic or hemorrhagic) or first-ever MI (ST-elevation) were identified through ICD-9 codes. Clinical details were abstracted from hospital records. Medication compliance was assessed through a structured telephone interview. STATISTICAL ANALYSIS: Bivariate analysis using Chi-square and Fisher exact tests, to determine the prevalence of noncompliance in stroke versus MI patients and differences in baseline characteristics and multivariate analysis with logistic regression to determine independent predictors of noncompliance. RESULTS: Follow-up data was collected for 298 stroke and 275 MI patients. Compliance was lower in stroke patients (68% stroke patients compliant with at least half their discharge prescriptions versus 90% MI patients; P < 0.001). Literacy and post-discharge follow-up were associated with greater compliance (P < 0.05 for both). Compliance was highest with anti-hypertensive drugs (98% after MI, 78% after stroke), followed by anti-platelet agents (94% after MI, 75% after stroke) and anti-lipid agents (70% after MI, 59% after stroke). Patients reported simply not feeling the need, acquiring fresh medical advice or a perceived lack of benefit, as reasons for not complying with their discharge prescriptions. CONCLUSIONS: Although similar drugs are involved, compliance with prescribed regimens is appreciably lower after stroke than after MI. Our findings underscore the need for better patient education regarding secondary prevention after stroke.