Metabolic labeling of cardiomyocyte-derived small extracellular-vesicle (sEV) miRNAs identifies miR-208a in cardiac regulation of lung gene expression.

Toxoplasma gondii uracil phosphoribosyltransferase (UPRT) converts 4-thiouracil (4TUc) into 4-thiouridine (4TUd), which is incorporated into nascent RNAs and can be biotinylated, then labelled with streptavidin conjugates or isolated via streptavidin-affinity methods. Here, we generated mice that expressed T. gondii UPRT only in cardiomyocytes (CM UPRT mice) and tested our hypothesis that CM-derived miRNAs (CM miRs) are transferred into remote organs after myocardial infarction (MI) by small extracellular vesicles (sEV) that are released from the heart into the peripheral blood (PB sEV). We found that 4TUd was incorporated with high specificity and sensitivity into RNAs isolated from the hearts and PB sEV of CM UPRT mice 6 h after 4TUc injection. In PB sEV, 4TUd was incorporated into CM-specific/enriched miRs including miR-208a, but not into miRs with other organ or tissue-type specificities. 4TUd-labelled miR208a was also present in lung tissues, especially lung endothelial cells (ECs), and CM-derived miR-208a (CM miR-208a) levels peaked 12 h after experimentally induced MI in PB sEV and 24 h after MI in the lung. Notably, miR-208a is expressed from intron 29 of α myosin heavy chain (αMHC), but αMHC transcripts were nearly undetectable in the lung. When PB sEV from mice that underwent MI (MI-PB sEV) or sham surgery (Sham-PB sEV) were injected into intact mice, the expression of Tmbim6 and NLK, which are suppressed by miR-208a and cooperatively regulate inflammation via the NF-κB pathway, was lower in the lungs of MI-PB sEV-treated animals than the lungs of animals treated with Sham-PB sEV or saline. In MI mice, Tmbim6 and NLK were downregulated, whereas endothelial adhesion molecules and pro-inflammatory cells were upregulated in the lung; these changes were significantly attenuated when the mice were treated with miR-208a antagomirs prior to MI surgery. Thus, CM UPRT mice enables us to track PB sEV-mediated transport of CM miRs and identify an miR-208a-mediated mechanism by which myocardial injury alters the expression of genes and inflammatory response in the lung.

Self-Assembling Peptide-Based Hydrogels in Angiogenesis.

Ischemic diseases, especially in the heart and the brain, have become a serious threat to human health. Growth factor and cell therapy are emerging as promising therapeutic strategies; however, their retention and sustainable functions in the injured tissue are limited. Self-assembling peptide (SAP)-based hydrogels, mimicking the extracellular matrix, are therefore introduced to encapsulate and controllably release cells, cell-derived exosomes or growth factors, thus promoting angiogenesis and tissue recovery after ischemia. We will summarize the classification, composition and structure of SAPs, and the influencing factors for SAP gelation. Moreover, we will describe the functionalized SAPs, and the combinatorial therapy of cells, exosomes or growth factors with functionalized SAPs for angiogenic process as well as its advantage in immunogenicity and injectability. Finally, an outlook on future directions and challenges is provided.

Human umbilical cord mesenchymal stem cell derived exosomes encapsulated in functional peptide hydrogels promote cardiac repair.

Stem cell-derived exosomes have been recognized as a potential therapy for cardiovascular disease. However, the low retention rate of exosomes after transplantation in vivo remains a major challenge in clinical applications. The aim of this study is to investigate whether human umbilical cord mesenchymal stem cell derived exosomes (UMSC-Exo) encapsulated in functional peptide hydrogels could increase the retention and stability of exosomes and improve heart function in a rat myocardial infarction model. Our results demonstrated that the PA-GHRPS peptide protected H9C2 cells from H2O2-induced oxidative stress. The gelatinization ability of PA-GHRPS can be enhanced by peptide NapFF. Therefore, these two peptides were mixed to form the PGN hydrogel, which was used to encapsulate exosomes. Our data showed that the PGN hydrogel was able to encapsulate exosomes effectively and ensured a stable and sustained release of exosomes. The exosome/PGN hydrogel mixture was injected into the infarcted border zone of rat hearts. Compared to the exosome treatment alone, the mixture improved the myocardial function by reducing inflammation, fibrosis and apoptosis, and by promoting angiogenesis. The strategy used in this study provided a practical and effective method to harness exosomes for myocardial regeneration.

Delivery of miR-675 by stem cell-derived exosomes encapsulated in silk fibroin hydrogel prevents aging-induced vascular dysfunction in mouse hindlimb.

Vascular disease is a major complication of aging, but the molecular mechanisms underlying the aging-induced vascular dysfunction remain unclear, and there is no effective treatment to prevent aging induced diseases. The objectives of the present study are to identify the signaling pathway mediating aging-induced vascular dysfunction and to develop an exosome based therapy to inhibit aging process. We used 11-month-old C57BL6 mice as pre-aging animal model and H2O2 treated H9C2 cells as an in vitro aging model to examine the therapeutic effect of miR-675. We found decreased expression of the potential aging modulator miR-675 in aging muscle, and H2O2 treatment decreased the expression of miR-675 and upregulated the expression of the aging marker ß-gal and TGF-ß1. We also found that miR-675 mimic decreased ß-gal staining in H2O2 treated H9C2 cells. Dual-luciferase reporter assays verified TGF-ß1 as the target gene of miR-675. Moreover, senescent H9C2 cells incubated with exosomes isolated from UMSCs transfected with the miR-675 mimic showed increased expression of miR-675, reduced activity of the aging marker ß-gal and reduced protein levels of TGF-ß1. We employed silk fibroin hydrogel to encapsulate exosomes in order to prolong the half-life of exosome in vivo. Fourier transform infrared spectroscopy (FTIR) revealed that exosomes were successfully encapsulated by the hydrogel. Laser Doppler perfusion imaging showed that the miR-675 exosomes encapsulated in silk fibroin hydrogel promote blood perfusion in ischemic hindlimbs. We demonstrated that miR-675 exosomes encapsulated in silk fibroin hydrogel provided sustained release of exosomes in vitro, and increased the retention time of red fluorescent PKH26-exosome in the tissue. Taken together, this study identified miR-675 as an important regulator of cell senescence and provided a novel strategy to deliver powerful exosomes by silk fibroin hydrogel to treat aging-induced vascular dysfunction.

Vertebrate estrogen regulates the development of female characteristics in silkworm, Bombyx mori.

The vertebrate estrogens include 17-ß-estradiol (E2), which has an analog in silkworm ovaries. In this study, the Bombyx mori vitellogenin gene (BmVg) was used as a biomarker to analyze the function of the E2 in silkworm. In most oviparous animals, Vg has female-specific expression. However, BmVg expression was also detected in B. mori males. Stage specific fluctuation of BmVg expression was similar in males and females, but expression levels in males were lower than in females. E2 treatment by injection or feeding of male larvae in the final instar stage induced and stimulated male BmVg transcription and protein synthesis. When silkworm ovary primordia were transplanted into males, BmVg was induced in male fat bodies. Transplanted ovaries primordia were also able to develop into ovaries and produce mature eggs. When females were treated with E2 promoted BmVg/BmVn protein accumulation in hemolymph, ovaries and eggs. However, BmVg transcription was decreased in female fat bodies. An E2 analog was identified in the hemolymph of day 3 wandering silkworms using high-performance liquid chromatography. Estradiol titers from fifth late-instar larvae to pupal stage were determined by enzyme-linked immunosorbent assay. The results suggested that silkworms synthesized a vertebrate E2 analog. This study found that E2 promoted the synthesis of BmVg, a female typical protein in silkworms.