Reproducibility and staging of 3D human retinal organoids across multiple pluripotent stem cell lines.

Numerous protocols have been described for producing neural retina from human pluripotent stem cells (hPSCs), many of which are based on the culture of 3D organoids. Although nearly all such methods yield at least partial segments of retinal structure with a mature appearance, variabilities exist within and between organoids that can change over a protracted time course of differentiation. Adding to this complexity are potential differences in the composition and configuration of retinal organoids when viewed across multiple differentiations and hPSC lines. In an effort to understand better the current capabilities and limitations of these cultures, we generated retinal organoids from 16 hPSC lines and monitored their appearance and structural organization over time by light microscopy, immunocytochemistry, metabolic imaging and electron microscopy. We also employed optical coherence tomography and 3D imaging techniques to assess and compare whole or broad regions of organoids to avoid selection bias. Results from this study led to the development of a practical staging system to reduce inconsistencies in retinal organoid cultures and increase rigor when utilizing them in developmental studies, disease modeling and transplantation.

Cancer survivors' perspectives of dietary information provision after cancer treatment: A scoping review of the Australian context.

ISSUE ADDRESSED: To support survivor-centred care in Australia, this review maps current knowledge regarding adult cancer survivors' perspectives of dietary information provision post-treatment. METHODS: A scoping review of research conducted in Australia within the past decade reported using PRISMA-ScR guidelines. Seven databases were searched (01/01/2009-05/06/2020) and records were independently screened by two researchers using eligibility criteria. Papers in the peer-reviewed literature with dietary information post-treatment as a primary and secondary outcome were eligible for inclusion. Data charting included participant characteristics, study methodology and cancer survivors' reports of dietary information provision post-treatment. RESULTS: Of 531 records identified, 12 met eligibility criteria. Most studies included breast (58%) and colorectal (42%) cancer survivors within 5 years post-diagnosis (84%). Three studies were conducted amongst specific ethnic groups (Indigenous Australians, Chinese-Australians, Greek-Australians). Participants in the included studies commonly reported limited or ineffective dietary information from healthcare providers post-treatment. Cancer survivors identified a need for individualised information regarding dietary strategies to manage ongoing symptoms, professional support for weight management, and practical skills for healthy eating. Amongst ethnic groups, there was a need for dietary information that considers traditional foods and cultural beliefs, and is available in their native language. Cancer survivors valued ongoing dietary follow-up and support post-treatment, and suggested a variety of face-to-face and online delivery modes. Those residing in rural and remote areas reported barriers to accessing dietary information post-treatment including time, cost, and availability of local services. CONCLUSIONS: There is scope to improve dietary information provision after cancer treatment in Australia. SO WHAT?: Dietary guidance post-treatment should consider individual needs, cultural background, and opportunity for ongoing follow-up and support.

A Novel Approach to Single Cell RNA-Sequence Analysis Facilitates In Silico Gene Reporting of Human Pluripotent Stem Cell-Derived Retinal Cell Types.

Cell type-specific investigations commonly use gene reporters or single-cell analytical techniques. However, reporter line development is arduous and generally limited to a single gene of interest, while single-cell RNA (scRNA)-sequencing (seq) frequently yields equivocal results that preclude definitive cell identification. To examine gene expression profiles of multiple retinal cell types derived from human pluripotent stem cells (hPSCs), we performed scRNA-seq on optic vesicle (OV)-like structures cultured under cGMP-compatible conditions. However, efforts to apply traditional scRNA-seq analytical methods based on unbiased algorithms were unrevealing. Therefore, we developed a simple, versatile, and universally applicable approach that generates gene expression data akin to those obtained from reporter lines. This method ranks single cells by expression level of a bait gene and searches the transcriptome for genes whose cell-to-cell rank order expression most closely matches that of the bait. Moreover, multiple bait genes can be combined to refine datasets. Using this approach, we provide further evidence for the authenticity of hPSC-derived retinal cell types. Stem Cells 2018;36:313-324.

Best practices in the pediatric pretransplant psychosocial evaluation.

Assessment of psychosocial functioning is an often-included component of the pretransplant evaluation process. This study reviews several domains of assessment that have been related to post-transplant outcomes across solid organ transplant populations. These include evaluation of patient and family past adherence, knowledge about the transplantation process, and their neurocognitive, psychological, and family functioning. To date, few comprehensive pretransplant evaluation measures have been standardized for use with children; however, several assessment measures used to evaluate the aforementioned domains are reviewed throughout the study. Additionally, this article discusses some developmental, illness-specific, and cultural considerations in conducting the psychosocial evaluation. We also discuss ethical issues specific to the pediatric psychosocial evaluation. Recommendations are advanced to promote a comprehensive evaluation that identifies family strengths and risk factors as they begin the transplant journey.

Generation of a rod-specific NRL reporter line in human pluripotent stem cells.

Reporter lines generated in human pluripotent stem cells can be highly useful for the analysis of specific cell types and lineages in live cultures. We created the first human rod reporter line using CRISPR/Cas9 genome editing to replace one allele of the Neural Retina Leucine zipper (NRL) gene with an eGFP transgene in the WA09 human embryonic stem cell (hESC) line. After confirming successful targeting, three-dimensional optic vesicle structures were produced to examine reporter specificity and to track rod differentiation in culture. The NRL+/eGFP hESC line robustly and exclusively labeled the entirety of rods throughout differentiation, eventually revealing highly mature structural features. This line provides a valuable tool for studying human rod development and disease and testing therapeutic strategies for retinitis pigmentosa.

3D Microstructured Scaffolds to Support Photoreceptor Polarization and Maturation.

Blinding disorders of the outer retina involve dysfunction and degeneration of photoreceptors. One potential approach to treat these forms of blindness is to repopulate the outer retina via a simple bolus injection of donor photoreceptors. However, this may not be ideal due to the highly polarized organization of photoreceptors that include apical light sensing photopigments and basal axon terminals. Furthermore, bolus injections create uncertainty with regard to the area, density, and retention of donor cells. Here, a novel and robust microfabrication process is developed to create 3D, micrometer-sized complex structures in ultrathin and biocompatible elastomer films (nonbiodegradable polydimethylsiloxane and biodegradable poly(glycerol-sebacate)) that can serve as polarizable photoreceptor delivery scaffolds, consisting of an array of cup-shaped photoreceptor capture wells that funnel into a microchannel. This "wine glass" scaffold design promotes efficient capture of human pluripotent stem-cell-derived photoreceptor cell bodies and guidance of basal axon extensions, ultimately achieving a uniform level of organization and polarization that is not possible with bolus injections or previously described scaffolds. In addition to future therapeutic applications, our scaffold design and materials provide a platform to generate reproducible and scalable in vitro models of photoreceptor-based diseases.

Predictors of parent-child interaction style in dyads with autism.

Parent synchrony has been shown to be developmentally important for the growth of communication skills in young children with autism. Understanding individual-differences in parent synchrony and other associated features of dyadic interaction therefore presents as an important step toward the goal of appreciating how and why some parent-child dyads come to adopt more optimal interaction styles, while for others, parent interaction is more asynchronous and less developmentally facilitative. Within the large, well-characterized Preschool Autism Communication Trial (PACT) cohort, baseline parent-child interaction samples were coded for three key aspects of dyadic interaction style; - Parent Synchrony, Child Initiation, and Shared Attention. We explored associations among these measures, demographic characteristics and standardized child assessment scores. While various child factors were associated with each of the interaction measures, very few associations were observed with parent/familial factors. Child language age-equivalence was a significant positive predictor of variation in each interaction measure, while child repetitive symptoms predicted reduced Shared Attention. The three interaction measures were moderately positively inter-related. In the context of childhood autism, variation in dyadic interaction style appears to be driven more by child language and repetitive behaviors than age, social-communication symptoms and non-verbal ability. Parent/family factors contributed little to explaining variability in parent-child interaction, in the current study.