Induced Pluripotent Stem Cell (iPSC)-Derived Extracellular Vesicles Are Safer and More Effective for Cardiac Repair Than iPSCs.

RATIONALE: Extracellular vesicles (EVs) are tiny membrane-enclosed droplets released by cells through membrane budding or exocytosis. The myocardial reparative abilities of EVs derived from induced pluripotent stem cells (iPSCs) have not been directly compared with the source iPSCs. OBJECTIVE: To examine whether iPSC-derived EVs can influence the biological functions of cardiac cells in vitro and to compare the safety and efficacy of iPSC-derived EVs (iPSC-EVs) and iPSCs for cardiac repair in vivo. METHODS AND RESULTS: Murine iPSCs were generated, and EVs isolated from culture supernatants by sequential centrifugation. Atomic force microscopy, high-resolution flow cytometry, real-time quantitative RT-PCR, and mass spectrometry were used to characterize EV morphology and contents. iPSC-EVs were enriched in miRNAs and proteins with proangiogenic and cytoprotective properties. iPSC-EVs enhanced angiogenic, migratory, and antiapoptotic properties of murine cardiac endothelial cells in vitro. To compare the cardiac reparative capacities in vivo, vehicle, iPSCs, and iPSC-EVs were injected intramyocardially at 48 hours after a reperfused myocardial infarction in mice. Compared with vehicle-injected mice, both iPSC- and iPSC-EV-treated mice exhibited improved left ventricular function at 35 d after myocardial infarction, albeit iPSC-EVs rendered greater improvement. iPSC-EV injection also resulted in reduction in left ventricular mass and superior perfusion in the infarct zone. Both iPSCs and iPSC-EVs preserved viable myocardium in the infarct zone, whereas reduction in apoptosis was significant with iPSC-EVs. iPSC injection resulted in teratoma formation, whereas iPSC-EV injection was safe. CONCLUSIONS: iPSC-derived EVs impart cytoprotective properties to cardiac cells in vitro and induce superior cardiac repair in vivo with regard to left ventricular function, vascularization, and amelioration of apoptosis and hypertrophy. Because of their acellular nature, iPSC-EVs represent a safer alternative for potential therapeutic applications in patients with ischemic myocardial damage.

Essential role for Notch signaling in restricting developmental plasticity.

We report that Notch signaling is essential for the switch from developmental plasticity to commitment during Caenorhabditis elegans embryogenesis. The GLP-1 and LIN-12 Notch receptors act to set a memory state that affects commitment of cells arising from the major ectodermal progenitor (AB blastomere) several cell divisions later, thereby preventing their forced reprogramming by an endoderm-determining transcription factor. In contrast to Notch-dependent cell fate induction, this activity is autonomous to the AB lineage, is independent of the known cell fate-inducing Notch ligands, and requires a putative secreted Notch ligand, Delta Serrate Lag-3 (DSL-3). Thus, Notch signaling promotes developmental commitment by a mechanism that is distinct from that involved in specifying cell fates.

Vitamin D3 induces mesenchymal-to-endothelial transition and promotes a proangiogenic niche through IGF-1 signaling.

Although vitamin D3 (VitD3) prevents angiogenesis in cancer, VitD3 deficiency is associated with greater incidence of cardiovascular events in patients. We examined the influence of VitD3 on the angiogenic potential of mesenchymal stem cells (MSCs). VitD3 treatment increased the expression of proangiogenic molecules in MSCs, which exhibited an endothelial cell-like phenotype and promoted vascularization in vitro and in vivo. VitD3 activated the IGF-1 promoter and boosted IGF-1 receptor (IGF-1R) signaling, which was essential for the mesenchymal-to-endothelial transition (MEndoT) of MSCs. VitD3-treated MSCs created a proangiogenic microenvironment for co-cultured arterial endothelial cells, as well as aortic rings. The induction of MEndoT and angiogenesis promotion by VitD3-stimulated MSCs was attenuated by IGF-1R inhibitor picropodophyllin. We conclude that VitD3 promotes MEndoT in MSCs, and VitD3-treated MSCs augment vascularization by producing a proangiogenic niche through continued IGF-1 secretion. These results suggest a potential therapeutic role of VitD3 toward enhancing MSC-induced angiogenesis.

RNA interference: silencing in the cytoplasm and nucleus.

Although the discovery that double-stranded RNA is able to silence gene expression was only made five years ago, methods for experimentally silencing genes have already been extended into a broad diversity of organisms, including human cells. RNA interference has also been discovered to function in physiological gene silencing. RNA interference works by causing degradation of targeted mRNAs in the cytoplasm. However, recent results suggest that RNA interference may also silence gene activity in the nucleus by remodeling chromatin and repressing the transcription of targeted genes.

Using RNA interference to identify genes required for RNA interference.

RNA interference (RNAi) is a phenomenon in which double-stranded RNA (dsRNA) silences endogenous gene expression. By injecting pools of dsRNAs into Caenorhabditis elegans, we identified a dsRNA that acts as a potent suppressor of the RNAi mechanism. We have used coinjection of dsRNAs to identify four additional candidates for genes involved in the RNAi mechanism in C. elegans. Three of the genes are C. elegans mes genes, some of which encode homologs of the Drosophila chromatin-binding Polycomb-group proteins. We have used loss-of-function mutants to confirm a role for mes-3, -4, and -6 in RNAi. Interestingly, introducing very low levels of dsRNA can bypass a requirement for these genes in RNAi. The finding that genes predicted to encode proteins that associate with chromatin are involved in RNAi in C. elegans raises the possibility that chromatin may play a role in RNAi in animals, as it does in plants.