Stern, J. H. | bioRxiv Science

bioRxiv2022

Identifying Biomarkers of Retinal Pigment Epithelial Cell Stem Cell-derived RPE Cell Heterogeneity and Transplantation Efficacy

Transplantation of retinal pigment epithelial (RPE) cells holds great promise for patients with retinal degenerative diseases such as age-related macular degeneration. In-depth characterization of RPE cell product identity and critical quality attributes are needed to enhance efficacy and safety of replacement therapy strategies. Here we characterized an adult RPE stem cell-derived (RPESC-RPE) cell product using bulk and single cell RNA sequencing (sc-RNA-seq), assessing functional cell integration in vitro into a mature RPE monolayer and in vivo efficacy by vision rescue in the Royal College of Surgeons rats. scRNA-seq revealed several distinct subpopulations in the RPESC-RPE product, some with progenitor markers. We identified RPE clusters expressing genes associated with in vivo efficacy and increased cell integration capability. Gene expression analysis revealed a lncRNA (TREX) as a predictive marker of in vivo efficacy. TREX knockdown decreased cell integration while overexpression increased integration in vitro and improved vision rescue in the RCS rats.

cell biology↗

bioRxiv2022

Glucagon Receptor Signaling at White Adipose Tissue Does Not Regulate Lipolysis

ObjectiveAlthough the physiologic role of glucagon receptor signaling in the liver is well defined, the impact of glucagon receptor (Gcgr) signaling at white adipose tissue (WAT) continues to be debated. While numerous studies propose glucagon stimulates WAT lipolysis, we lack evidence that physiological concentrations of glucagon regulate WAT lipolysis. Glucagon receptor antagonists are proposed as a treatment to lower blood glucose in people with type 2 diabetes, yet concerns on how these treatments may affect lipid homeostasis have led to questions regarding the potential safety and efficacy of such therapeutics. Tight regulation of adipose tissue lipolysis is critical for whole body lipid homeostasis. In turn, we used WAT Gcgr knockout mice to determine if glucagon regulates lipolysis at WAT in the mouse. MethodsWe assessed the effects of fasting and acute exogenous glucagon administration in wildtype C57BL/6J and Gcgr Adipocyte+/+ vs GcgrAdipocyte-/- mice. Using an ex vivo lipolysis protocol, we further examined the direct effects of glucagon on physiologically (fasted) and pharmacologically stimulated lipolysis. ResultsAdipocyte Gcgr expression did not affect fasting induced lipolysis or hepatic lipid accumulation in lean or diet induced obese (DIO) mice. Acute glucagon administration did not affect serum non-esterified fatty acids (NEFA), leptin, or adiponectin concentration, but did increase serum glucose and FGF21, regardless of genotype. Glucagon did not affect ex vivo lipolysis in explants from either Gcgr Adipocyte+/+ or GcgrAdipocyte-/- mice. Gcgr expression did not affect fasting-induced or isoproterenol-stimulated lipolysis from WAT explants. Moreover, glucagon receptor signaling at WAT does not affect body weight or glucose homeostasis in lean or DIO mice. ConclusionsWe have established that glucagon does not regulate WAT lipolysis, either directly or indirectly. Unlike the crucial role of hepatic glucagon receptor signaling in maintaining glucose and lipid homeostasis, we observed no metabolic consequence of WAT glucagon receptor deletion.

physiology↗