ABSTRACT
Background: Reproducibility of human cortical organoid (hCO) phenotypes remains a concern for modeling neurodevelopmental disorders. While guided hCO protocols reproducibly generate cortical cell types in multiple cell lines at one site, variability across sites using a harmonized protocol has not yet been evaluated. We present an hCO cross-site reproducibility study examining multiple phenotypes. Methods: Three independent research groups generated hCOs from one induced pluripotent stem cell (iPSC) line using a harmonized miniaturized spinning bioreactor protocol. scRNA-seq, 3D fluorescent imaging, phase contrast imaging, qPCR, and flow cytometry were used to characterize the 3 month differentiations across sites. Results: In all sites, hCOs were mostly cortical progenitor and neuronal cell types in reproducible proportions with moderate to high fidelity to the in vivo brain that were consistently organized in cortical wall-like buds. Cross-site differences were detected in hCO size and morphology. Differential gene expression showed differences in metabolism and cellular stress across sites. Although iPSC culture conditions were consistent and iPSCs remained undifferentiated, primed stem cell marker expression prior to differentiation correlated with cell type proportions in hCOs. Conclusions: We identified hCO phenotypes that are reproducible across sites using a harmonized differentiation protocol. Previously described limitations of hCO models were also reproduced including off-target differentiations, necrotic cores, and cellular stress. Improving our understanding of how stem cell states influence early hCO cell types may increase reliability of hCO differentiations. Cross-site reproducibility of hCO cell type proportions and organization lays the foundation for future collaborative prospective meta-analytic studies modeling neurodevelopmental disorders in hCOs.
ABSTRACT
Background: Trans reproduction is often met with reproach when compared with the norm of the cisgender, heterosexual, nuclear family (referred to here as the 'traditional' family). This is notably seen in birth registration where a 'traditional' family is more likely to access, and agree with, the terms provided on the birth certificate (that is, 'mother' and 'father'). Aims: This study aimed to explore the differential treatment between 'traditional' and trans families in England and Wales within birth registration in case law, legislation, and primary research. Methods: Case law and legislation were analyzed to identify any differential treatment within birth registration, particularly how parental statuses are ascribed. An online survey (with 310 responses) was conducted, seeking views on constructions of 'mothers', respondents' reactions to trans families, and understandings of sex/gender and parenthood generally. Results: Themes developed from the survey data focused on the significance of the 'traditional' family as the optimal structure, manifesting in the need to ensure 'coherency and certainty' of the birth registration system regarding how parents should be registered. This need for coherency and certainty functioned to undermine trans families (who were presumed to be discreet about their child's conception), and also raised concerns that the child might not be able to locate their (biogenetic) 'origins'. Discussion: The paper concludes by discussing the implications of the research for the acceptance of trans families in law and society, and suggests the need to unmoor the birth registration system from its (cis-hetero)normative underpinnings, allowing trans parents to be registered as they wish on the birth certificate.
