Haploinsufficiency describes a phenomenon where one functional allele of a gene in a diploid cell or organism is insufficient for a normal phenotype. There are several neurodevelopmental disorders (NDD) affected by the haploinsufficiency phenomenon, and many of them are related to autism spectrum disorder (ASD). Here, we aim to identify genes involved in the early stages of neural differentiation when one of the two alleles is lost. We thus differentiated a genome-wide heterozygous loss-of-function CRISPR library into neural progenitor cells (NPCs) and defined about 250 genes essential for neural differentiation in a haploinsufficient manner. We were able to identify NDD-related dosage-sensitive pathways and pinpoint specific molecular processes affected by ASD-related genes. By comparing the molecular phenotypes of homozygote and heterozygote mutations, we could illuminate overlapping and distinct transcriptional pathways affected in the two mutant models, along with partial chemical rescue of some of these phenotypes. Our work provides a comprehensive framework for exploring dosage-sensitive regulation in early neural development and offers new insights into the embryonic molecular basis of ASD and other NDDs driven by gene dosage imbalance.
Roni Sarel-Gallily, Assa Sherman, Daniel Pollak et al.· Cell Death and Disease· 1 citation
This study demonstrates the successful production and injection of human induced pluripotent stem cell cardiomyocyte aggregates into infarcted cynomolgus monkey hearts, resulting in substantial, structured human grafts three months after cell transplantation. Transient graft-induced arrhythmias decreased over time. Both the arrhythmogenicity and the substantial heart function recovery in vivo notably seemed to correlate with induced pluripotent stem cell clone-dependent contractile and electrophysiological cardiomyocyte properties in vitro. Overexpression of a red fluorescent reporter protein led to a dysregulated conduction and contraction machinery in yet engraftment competent cardiomyocytes, providing an important tool to mechanistically understand and improve induced pluripotent stem cell-based heart repair in preclinical models. We demonstrate the logistically important, temporal uncoupling of cardiomyocyte production from transplantation. Cardiomyocyte aggregate transplantation yielded results comparable to the reported transplantation of 10-20-fold higher numbers of dissociated human embryonic stem cell- cardiomyocytes and suggests a higher degree of cell/ tissue maturation in cardiac grafts. Our study promotes reduced cell production costs, highlights the need for an in vitro potency assay, and shows a pragmatic new avenue for the clinical translation of human induced pluripotent stem cell-based heart repair. The study shows successful therapy with hiPSC-cardiomyocyte aggregates in infarcted non-human primates, including uncoupling of cell production from transplantation and correlation of heart recovery in vivo with cardiomyocyte properties in vitro.
I. Gruh, Andreas Martens, S. Cebotari et al.· Nature Communications· 0 citations