A highly reproducible and efficient method for retinal organoid differentiation from human pluripotent stem cells.

Human pluripotent stem cell (hPSC)-derived retinal organoids are three-dimensional cellular aggregates that differentiate and self-organize to closely mimic the spatial and temporal patterning of the developing human retina. Retinal organoid models serve as reliable tools for studying human retinogenesis, yet limitations in the efficiency and reproducibility of current retinal organoid differentiation protocols have reduced the use of these models for more high-throughput applications such as disease modeling and drug screening. To address these shortcomings, the current study aimed to standardize prior differentiation protocols to yield a highly reproducible and efficient method for generating retinal organoids. Results demonstrated that through regulation of organoid size and shape using quick reaggregation methods, retinal organoids were highly reproducible compared to more traditional methods. Additionally, the timed activation of BMP signaling within developing cells generated pure populations of retinal organoids at 100% efficiency from multiple widely used cell lines, with the default forebrain fate resulting from the inhibition of BMP signaling. Furthermore, given the ability to direct retinal or forebrain fates at complete purity, mRNA-seq analyses were then utilized to identify some of the earliest transcriptional changes that occur during the specification of these two lineages from a common progenitor. These improved methods also yielded retinal organoids with expedited differentiation timelines when compared to traditional methods. Taken together, the results of this study demonstrate the development of a highly reproducible and minimally variable method for generating retinal organoids suitable for analyzing the earliest stages of human retinal cell fate specification.

Heart failure is not a determinant of central sleep apnea in the pediatric population.

BACKGROUND/OBJECTIVES: Adults with heart failure (HF) have high prevalence of central sleep apnea (CSA). While this has been repeatedly investigated in adults, there is a deficiency of similar research in pediatric populations. The goal of this study was to compare prevalence of CSA in children with and without HF and correlate central apneic events with heart function. METHODS: Retrospective analysis of data from children with and without HF was conducted. Eligible children were less than 18 years old with echocardiogram and polysomnogram within 6 months of each other. Children were separated into groups with and without HF based on left ventricular ejection fraction (LVEF). Defining CSA as central apnea-hypopnea index (CAHI) more than 1/hour, the cohort was also classified into children with and without CSA for comparative study. RESULTS: A total of 120 children (+HF: 19, -HF: 101) were included. The +HF group was younger, with higher prevalence of trisomy 21, muscular dystrophy, oromotor incoordination, and structural heart disease. The +HF group had lower apnea-hypopnea index (median: 3/hour vs. 8.6/hour) and lower central apnea index (CAI) (median: 0.2/hour vs. 0.55/hour). Prevalence of CSA was similar in both groups (p = .195). LogCAHI was inversely correlated to age (Pearson correlation coefficient: -0.245, p = .022). Children with CSA were younger and had higher prevalence of prematurity (40% vs. 5.3%). There was no significant difference in LVEF between children with and without CSA. After excluding children with prematurity, relationship between CAHI and age was no longer sustained. CONCLUSIONS: In contrast to adults, there is no difference in prevalence of CSA in children with and without HF. Unlike in adults, LVEF does not correlate with CAI in children. Overall, it appears that central apneic events may be more a function of age and prematurity rather than of heart function.