Biology subjects

Colas, A. R.

Publications and source records attributed to Colas, A. R..

3 recordsLinked to original sources

bioRxiv2020

Lipid availability influences the metabolic maturation of human pluripotent stem cell-derived cardiomyocytes

ObjectivesPluripotent stem cell-derived cardiomyocytes are phenotypically immature, which limits their utility in downstream applications. Metabolism is dramatically reprogramed during cardiac maturation in vivo and presents a potential avenue to drive in vitro maturation. We aimed to identify and address metabolic bottlenecks in the generation of human pluripotent stem cell (hPSC)-derived cardiomyocytes. MethodshPSCs were differentiated into cardiomyocytes using an established, chemically-defined differentiation protocol. We applied 13C metabolic flux analysis (MFA) and targeted transcriptomics to characterize cardiomyocyte metabolism in during differentiation in the presence or absence of exogenous lipids. ResultshPSC-derived cardiomyocytes induced some cardiometabolic pathways (i.e. ketone body and branched-chain amino acid oxidation) but failed to effectively activate fatty acid oxidation. MFA studies indicated that lipid availability in cultures became limited during differentiation, suggesting potential issues with nutrient availability. Exogenous supplementation of lipids improved cardiomyocyte morphology, mitochondrial function, and promoted increased fatty acid oxidation in hPSC-derivatives. ConclusionhPSC-derived cardiomyocytes are dependent upon exogenous sources of lipids for metabolic maturation. Proper supplementation removes a potential roadblock in the generation of metabolically mature cardiomyocytes. These studies further highlight the importance of considering and exploiting metabolic phenotypes in the in vitro production and utilization of functional hPSC-derivatives.

cell biology↗

bioRxiv2020

PGC1/PPAR Drive Cardiomyocyte Maturation through Regulation of Yap1 and SF3B2

Cardiomyocytes undergo significant levels of structural and functional changes after birth--fundamental processes essential for the heart to produce the volume and contractility to pump blood to the growing body. However, due to the challenges in isolating single postnatal/adult myocytes, how individual newborn cardiomyocytes acquire multiple aspects of mature phenotypes remains poorly understood. Here we implemented large-particle sorting and analyzed single myocytes from neonatal to adult hearts. Early myocytes exhibited a wide-ranging transcriptomic and size heterogeneity, maintained until adulthood with a continuous transcriptomic shift. Gene regulatory network analysis followed by mosaic gene deletion revealed that peroxisome proliferator-activated receptor coactivator-1 signaling--activated in vivo but inactive in pluripotent stem cell-derived cardiomyocytes--mediates the shift. The signaling regulated key aspects of cardiomyocyte maturation simultaneously through previously unrecognized regulators, including Yap1 and SF3B2. Our study provides a single-cell roadmap of heterogeneous transitions coupled to cellular features and unveils a multifaceted regulator controlling cardiomyocyte maturation. Significance StatementHow the individual single myocytes achieve full maturity remains a black box, largely due to the challenges with the isolation of single mature myocytes. Understanding this process is particularly important as the immaturity and early developmental arrest of pluripotent stem cell-derived myocytes has emerged a major concern in the field. Here we present the first study of high-quality single-cell transcriptomic analysis of cardiac muscle cells from neonatal to adult hearts. We identify a central transcription factor and its novel targets that control key aspects of myocyte maturation, including cellular hypertrophy, contractility, and mitochondrial activity.

developmental biology↗

bioRxiv2019

Patient-specific functional genomics and disease modeling suggest a role for LRP2 in Hypoplastic Left Heart Syndrome

Congenital heart diseases (CHD), such as hypoplastic left heart syndrome (HLHS), are considered to have complex genetic underpinnings that are poorly understood. Here, an integrated multi-disciplinary approach was applied to identify novel genes and underlying mechanisms associated with HLHS. A family-based strategy was employed that coupled whole genome sequencing (WGS) with RNA sequencing of patient-derived induced pluripotent stem cells (iPSCs) from a sporadic HLHS proband-parent trio to identify, prioritize and functionally evaluate candidate genes in model systems. Consistent with the hypoplastic phenotype, the probands iPSCs had reduced proliferation capacity. Filtering WGS for rare de novo, recessive, and loss-of-function variants revealed 10 candidate genes with recessive variants and altered expression compared to the parents iPSCs. siRNA/RNAi-mediated knockdown in generic human iPSC-derived cardiac progenitors and in the in vivo Drosophila heart model revealed that LDL receptor related protein LRP2 and apolipoprotein APOB are required for robust hiPSC-derived cardiomyocyte proliferation and normal hear structure and function, possibly involving an oligogenic mechanism via growth-promoting WNT and SHH signaling. LRP2 was further validated as a CHD gene in a zebrafish heart model and rare variant burden testing in an HLHS cohort. Collectively, this cross-functional genetic approach to complex congenital heart disease revealed LRP2 dysfunction as a likely novel genetic driver of HLHS, and hereby established a scalable approach to decipher the oligogenic underpinnings of maladaptive left heart development. One sentence summaryWhole genome sequencing and a multi-model system candidate gene validation - human iPSC-derived cardiomyocytes and Drosophila and zebrafish hearts - identified lipoprotein LRP2 as a new potential driver in congenital heart disease and suggests a deficit in proliferation as a hallmark of hypoplastic left heart syndrome.

genetics↗