Rationale YAP, the nuclear effector of Hippo signaling, regulates cellular growth and survival in multiple organs, including the heart, by interacting with TEAD sequence specific DNA-binding proteins. Recent studies showed that YAP stimulates cardiomyocyte proliferation and survival. However, the direct transcriptional targets through which YAP exerts its effects are poorly defined. Objective To identify direct YAP targets that mediate its’ mitogenic and anti-apoptotic effects in the heart. Methods and Results We identified direct YAP targets by combining differential gene expression analysis in YAP gain- and loss-of-function with genome-wide identification of YAP bound loci using chromatin immunoprecipitation and high throughput sequencing. This screen identified Pik3cb, encoding p110β, a catalytic subunit of phosphoinositol-3-kinase (PI3K), as a candidate YAP effector that promotes cardiomyocyte proliferation and survival. YAP and TEAD occupied a conserved enhancer within the first intron of Pik3cb, and this enhancer drove YAP-dependent reporter gene expression. Yap gain- and loss-of-function studies indicated that YAP is necessary and sufficient to activate the PI3K-Akt pathway. Like Yap, Pik3cb gain-of-function stimulated cardiomyocyte proliferation, and Pik3cb knockdown dampened YAP mitogenic activity. Reciprocally, impaired heart function in Yap loss-of-function was significantly rescued by AAV-mediated Pik3cb expression. Conclusion : Pik3cb is a crucial direct target of YAP, through which the YAP activates PI3K-AKTpathway and regulates cardiomyocyte proliferation and survival.

Administration of exosomes derived from mesenchymal stromal cells (MSCs) could improve some neurologic conditions by transferring functional biomolecules to recipient cells. Furthermore, exosomes from hypoxic progenitor cells exerted better therapeutic effects in organ injury through specific cargoes. However, there are no related reports about whether exosomes derived from MSCs or hypoxia-preconditioned MSCs (PC-MSCs) could prevent memory deficits in Alzheimer disease (AD). In this study, the exosomes derived from MSCs or PC-MSCs were systemically administered to transgenic APP/PS1 mice. The expression of miR-21 in MSCs was significantly increased after hypoxic treatment. Injection of exosomes from normoxic MSCs could rescue cognition and memory impairment according to results of the Morris water maze test, reduced plaque deposition, and Aβ levels in the brain; could decrease the activation of astrocytes and microglia; could down-regulate proinflammatory cytokines (TNF-α and IL-1β); and could up-regulate anti-inflammatory cytokines (IL-4 and -10) in AD mice, as well as reduce the activation of signal transducer and activator of transcription 3 (STAT3) and NF-κB. Compared to the group administered exosomes from normoxic MSCs, in the group administered exosomes from PC-MSCs, learning and memory capabilities were significantly improved; the plaque deposition and Aβ levels were lower, and expression of growth-associated protein 43, synapsin 1, and IL-10 was increased; and the levels of glial fibrillary acidic protein, ionized calcium-binding adaptor molecule 1, TNF-α, IL-1β, and activation of STAT3 and NF-κB were sharply decreased. More importantly, exosomes from PC-MSCs effectively increased the level of miR-21 in the brain of AD mice. Additionally, replenishment of miR-21 restored the cognitive deficits in APP/PS1 mice and prevented pathologic features. Taken together, these findings suggest that exosomes from PC-MSCs could improve the learning and memory capabilities of APP/PS1 mice, and that the underlying mechanism may lie in the restoration of synaptic dysfunction and regulation of inflammatory responses through regulation of miR-21.-Cui, G.-H., Wu, J., Mou, F.-F., Xie, W.-H., Wang, F.-B., Wang, Q.-L., Fang, J., Xu, Y.-W., Dong, Y.-R., Liu, J.-R., Guo, H.-D. Exosomes derived from hypoxia-preconditioned mesenchymal stromal cells ameliorate cognitive decline by rescuing synaptic dysfunction and regulating inflammatory responses in APP/PS1 mice.

Background Exosomes are lipid-bilayer enclosed nano-sized vesicles that transfer functional cellular proteins, mRNA and miRNAs. Mesenchymal stem cells (MSCs) derived exosomes have been demonstrated to prevent memory deficits in the animal model of Alzheimer’s disease (AD). However, the intravenously injected exosomes could be abundantly tracked in other organs except for the targeted regions in the brain. Here, we proposed the use of central nervous system-specific rabies viral glycoprotein (RVG) peptide to target intravenously-infused exosomes derived from MSCs (MSC-Exo) to the brain of transgenic APP/PS1 mice. MSC-Exo were conjugated with RVG through a DOPE-NHS linker. Results RVG-tagged MSC-Exo exhibited improved targeting to the cortex and hippocampus after being administered intravenously. Compared with the group administered MSC-Exo, in the group administered RVG-conjugated MSC-Exo (MSC-RVG-Exo) plaque deposition and Aβ levels were sharply decreased and activation of astrocytes was obviously reduced. The brain targeted exosomes derived from MSCs was better than unmodified exosomes to improve cognitive function in APP/PS1 mice according to Morris water maze test. Additionally, although MSC-Exo injected intravenously reduced the expression of pro-inflammatory mediators TNF-α, IL-β, and IL-6, but the changes of anti-inflammatory factors IL-10 and IL-13 were not obvious. However, administration of MSC-RVG-Exo significantly reduced the levels of TNF-α, IL-β, and IL-6 while significantly raised the levels of IL-10, IL-4 and IL-13. Conclusions Taken together, our results demonstrated a novel method for increasing delivery of exosomes for treatment of AD. By targeting exosomes to the cortex and hippocampus of AD mouse, there was a significant improvement in learning and memory capabilities with reduced plaque deposition and Aβ levels, and normalized levels of inflammatory cytokines. Electronic supplementary material The online version of this article (10.1186/s12979-019-0150-2) contains supplementary material, which is available to authorized users.

SUMMARY Binding of the transcriptional co-activator YAP with the transcription factor TEAD stimulates growth of the heart and other organs. YAP overexpression potently stimulates fetal cardiomyocyte (CM) proliferation, but YAP's mitogenic potency declines post-natally. While investigating factors that limit YAP's postnatal mitogenic activity, we found that the CM-enriched TEAD1 binding protein VGLL4 inhibits CM proliferation by inhibiting TEAD1-YAP interaction and by targeting TEAD1 for degradation. Importantly, VGLL4 acetylation at lysine 225 negatively regulated its binding to TEAD1. This developmentally regulated acetylation event critically governs postnatal heart growth, since overexpression of an acetylation-refractory VGLL4 mutant enhanced TEAD1 degradation, limited neonatal CM proliferation, and caused CM necrosis. Our study defines an acetylation-mediated, VGLL4-dependent switch that regulates TEAD stability and YAP-TEAD activity. These insights may improve targeted modulation of TEAD-YAP activity in applications from cardiac regeneration to cancer.

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The high death rate of the transplanted stem cells in the infarcted heart and the low efficiency of differentiation toward cardiomyocytes influence the outcome of stem cell transplantation for treatment of myocardial infarction (MI). Fibrin glue (FG) has been extensively used as a cell implantation matrix to increase cell survival. However, mechanisms of the effects of FG for stem cell transplantation to improve cardiac function are unclear. We have isolated c-kit+/Sca-1+ marrow-derived cardiac stem cells (MCSCs) from rat bone marrow; the cells expressed weakly early cardiac transcription factor Nkx2.5, GATA-4, Mef2C, and Tbx5. Effects of FG on survival, proliferation, and migration of MCSCs were examined in vitro. Cytoprotective effects of FG were assessed by exposure of MCSCs to anoxia. Efficacy of MCSC transplantation in FG was evaluated in the female rat MI model. The MCSCs survived well and proliferated in FG, and they may migrate out from the edge of FG in the wound and nature state. Acridine orange/ethidium bromide staining and lactate dehydrogenase analysis showed that MCSCs in FG were more resistant to anoxia as compared with MCSCs alone. In a rat MI model, cardiac function was improved and scar area was obviously reduced in group of MCSCs in FG compared with group of MCSCs and FG alone, respectively. Y chromosome fluorescence in situ hybridization showed that there were more survived MCSCs in group of MCSCs in FG than those in group of MCSCs alone, and most Y chromosome positive cells expressed cardiac troponin T (cTnT) and connexin-43 (Cx-43). Cx-43 was located between Y chromosome positive cells and recipient cardiomyocytes. Microvessel density in the peri-infarct regions and infarct regions significantly increased in group of MCSCs in FG. These results suggest that FG provide a suitable microenvironment for survival and proliferation of MCSCs and protect cells from apoptosis and necrosis caused by anoxia. MCSCs could differentiate into cardiomyocytes after being transplanted in the border of the infarcted myocardium and form connections with native cardiomyocytes. FG is helpful for MCSC transplantation to repair myocardium and improve cardiac function through promoting the survival, migration, and cardiomyogenic differentiation of MCSCs and inducing angiogenesis.

The critical role played by IgE in allergic asthma is well-documented and clinically precedented, but some patients in whom IgE neutralization may still offer clinical benefit are excluded from treatment with the existing anti-IgE therapy, omalizumab, due to high total IgE levels or body mass. In this study, we sought to generate a novel high affinity anti-IgE antibody (MEDI4212) with potential to treat a broad severe asthma patient population. Analysis of body mass, total and allergen-specific IgE levels in a cohort of severe asthmatics was used to support the rationale for development of a high affinity IgE-targeted antibody therapeutic. Phage display technology was used to generate a human IgG1 lead antibody, MEDI4212, which was characterized in vitro using binding, signaling and functional assay systems. Protein crystallography was used to determine the details of the interaction between MEDI4212 and IgE. MEDI4212 bound human IgE with an affinity of 1.95 pM and was shown to target critical residues in the IgE Cε3 domain critical for interaction with FcεRI. MEDI4212 potently inhibited responses through FcεRI and also prevented the binding of IgE to CD23. When used ex vivo at identical concentration, MEDI4212 depleted free-IgE from human sera to levels ~1 log lower than omalizumab. Our results thus indicate that MEDI4212 is a novel, high affinity antibody that binds specifically to IgE and prevents IgE binding to its receptors. MEDI4212 effectively depleted free-IgE from human sera ex vivo to a level (1 IU/mL) anticipated to provide optimal IgE suppression in severe asthma patients.

GATA4 and GATA6 are central cardiac transcriptional regulators. The postnatal, stage-specific function of the cardiac transcription factors GATA4 and GATA6 have not been evaluated. In part, this is because current Cre-loxP approaches to cardiac gene inactivation require time consuming and costly breeding of Cre-expressing and “floxed” mouse lines, often with limited control of the extent or timing of gene inactivation. We investigated the stage-specific functions of GATA4 and GATA6 in the postnatal heart by using adeno-associated virus serotype 9 to control the timing and extent of gene inactivation by Cre. Systemic delivery of recombinant, adeno-associated virus 9 (AAV9) expressing Cre from the cardiac specific Tnnt2 promoter was well tolerated and selectively and efficiently recombined floxed target genes in cardiomyocytes. AAV9:Tnnt2-Cre efficiently inactivated Gata4 and Gata6. Neonatal Gata4/6 inactivation caused severe, rapidly lethal systolic heart failure. In contrast, Gata4/6 inactivation in adult heart caused only mild systolic dysfunction but severe diastolic dysfunction. Reducing the dose of AAV9:Tnnt2-Cre generated mosaics in which scattered cardiomyocytes lacked Gata4/6. This mosaic knockout revealed that Gata4/6 are required cell autonomously for physiological cardiomyocyte growth. Our results define novel roles of GATA4 and GATA6 in the neonatal and adult heart. Furthermore, our data demonstrate that evaluation of gene function hinges on controlling the timing and extent of gene inactivation. AAV9:Tnnt2-Cre is a powerful tool for controlling these parameters.