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Recent advances in stem cell biology have allowed researchers to efficiently produce large numbers of cardiomyocytes from various pluripotent cell sources. Unfortunately, these cells exhibit properties that are characteristics of immature cardiomyocytes such as poor sarcomere organization, limited calcium handling, and reduced cell size and alignment. Specifically, the actin–myosin motor proteins that form sarcomeres within these cardiomyocytes fail to produce large, highly ordered repeating structures that are distinctive for adult myocytes. Instead, these cells produce heterogeneous sarcomeres that vary in thickness, alignment, and level of organization. Additionally, a large number of cardiomyopathies have been linked to mutations in genes encoding for sarcomeric proteins, resulting in disrupted sarcomere organization. This research focuses on a series of algorithms that provide a quantitative analysis technique to characterize the alignment and organization of sarcomere structures within aggregates and single cardiomyocytes. The scanning gradient Fourier transform (SGFT) method incorporates gradient analysis along with fast Fourier transforms to determine regions of sarcomere organization within individual and a population of cells, yielding a quantitative method of determining sarcomere organization and alignment at the sub-cellular scale. The utility of the SGFT technique is also demonstrated for additional applications, such as breast cancer collagen microstructure and neural rosette patterning. [ABSTRACT FROM AUTHOR]

Neural organoids have revolutionized how human neurodevelopmental disorders (NDDs) are studied. Yet, their utility for screening complex NDD etiologies and in drug discovery is limited by a lack of scalable and quantifiable derivation formats. Here, we describe the RosetteArray ® platform's ability to be used as an off-the-shelf, 96-well plate assay that standardizes incipient forebrain and spinal cord organoid morphogenesis as micropatterned, 3-D, singularly polarized neural rosette tissues (>9000 per plate). RosetteArrays are seeded from cryopreserved human pluripotent stem cells, cultured over 6-8 days, and immunostained images can be quantified using artificial intelligence-based software. We demonstrate the platform's suitability for screening developmental neurotoxicity and genetic and environmental factors known to cause neural tube defect risk. Given the presence of rosette morphogenesis perturbation in neural organoid models of NDDs and neurodegenerative disorders, the RosetteArray platform could enable quantitative high-throughput screening (qHTS) of human neurodevelopmental risk across regulatory and precision medicine applications., Competing Interests: DECLARATION OF INTERESTS G.T.K, R.W., and R.S.A are co-founders and co-owners of Neurosetta LLC, which focuses on commercializing RosetteArray™ technology for human neurodevelopmental risk assessment. N.J.F. and K.K. are employees at Neurosetta LLC.