Exosomes are nano-sized vesicles (30–200 nm) constantly released by almost all cells. The ability of exosomes to travel between cells and deliver their cargo, which includes lipids, proteins, and nucleic acids, makes them an appealing cell-free therapy option to treat multiple diseases. Here, we investigated for the first time whether human adipose tissue-derived mesenchymal stem cell-derived exosomes (ASC-exosomes) can ameliorate atopic dermatitis (AD) in an in vivo mouse model. When injected either intravenously (IV) or subcutaneously (SC) into NC/Nga mice treated with house dust mite antigens, ASC-exosomes were found to reduce pathological symptoms such as clinical score, the levels of serum IgE, the number of eosinophils in blood, and the infiltration of mast cells, CD86+, and CD206+ cells in skin lesions. ASC-exosomes also significantly reduced mRNA expression of various inflammatory cytokines such as interleukin (IL)-4, IL-23, IL-31, and tumor necrosis factor-α (TNF-α) in AD skin lesions of Nc/Nga mice. Taken together, these results suggest that ASC-exosomes can be a novel promising cell-free therapeutic modality for AD treatment.

Atopic dermatitis (AD) is a multifactorial, heterogeneous disease associated with epidermal barrier disruption and intense systemic inflammation. Previously, we showed that exosomes derived from human adipose tissue-derived mesenchymal stem cells (ASC-exosomes) attenuate AD-like symptoms by reducing multiple inflammatory cytokine levels. Here, we investigated ASC-exosomes’ effects on skin barrier restoration by analyzing protein and lipid contents. We found that subcutaneous injection of ASC-exosomes in an oxazolone-induced dermatitis model remarkably reduced trans-epidermal water loss, while enhancing stratum corneum (SC) hydration and markedly decreasing the levels of inflammatory cytokines such as IL-4, IL-5, IL-13, TNF-α, IFN-γ, IL-17, and TSLP, all in a dose-dependent manner. Interestingly, ASC-exosomes induced the production of ceramides and dihydroceramides. Electron microscopic analysis revealed enhanced epidermal lamellar bodies and formation of lamellar layer at the interface of the SC and stratum granulosum with ASC-exosomes treatment. Deep RNA sequencing analysis of skin lesions demonstrated that ASC-exosomes restores the expression of genes involved in skin barrier, lipid metabolism, cell cycle, and inflammatory response in the diseased area. Collectively, our results suggest that ASC-exosomes effectively restore epidermal barrier functions in AD by facilitating the de novo synthesis of ceramides, resulting in a promising cell-free therapeutic option for treating AD.

Progressive cardiac hypertrophy owing to pathological stimuli, such as pressure overload, is frequently associated with the development of heart failure, a major cause of morbidity and mortality worldwide. Growing evidence has shown that miRNAs are extensively involved in the pathogenesis of cardiac hypertrophy. In the present study, we examined the hypothesis that the miR-19a/b family acts as a key regulator of cardiac hypertrophy and apoptosis. Forced overexpression of miR-19a/b was sufficient to induce hypertrophy in rat neonatal cardiomyocytes. Luciferase assays revealed that miR-19a/b directly target the anti-hypertrophic genes atrogin-1 and MuRF-1 (muscle RING-finger protein-1). The endogenous expressions of the target genes were down-regulated by miR-19a/b. Pro-hypertrophic calcineurin/NFAT (nuclear factor of activated T-cells) signalling was elevated markedly in the presence of miR-19b, and the calcineurin inhibitor CsA (cyclosporin A) and the PKC (protein kinase C) inhibitor GF10923X significantly attenuated the miR-19b-mediated increase in cell size and expression of hypertrophic markers. Furthermore, miR-19b led to increased cell survival through up-regulation of the NFAT target gene encoding α-crystallin-B and repression of the pro-apoptotic gene Bim (Bcl-2-interacting mediator of cell death) under ER (endoplasmic reticulum) stress conditions. Taken together, the results of the present study demonstrate that the miR-19a/b family regulates phenotypes of cardiomyocytes via suppression of multiple direct target genes.

Pressure overload in the heart induces pathological hypertrophy and is associated with cardiac dysfunction. Apoptosis and fibrosis signaling initiated by the endoplasmic reticulum stress (ERS) is known to contribute to these maladaptive effects. The aim of this study was to investigate whether reduction of ERS by a known chemical chaperone, tauroursodeoxycholic acid (TUDCA) can attenuate pressure overload-induced cardiac remodeling in a mouse model of transverse aortic constriction (TAC). Oral administration of TUDCA at a dose of 300 mg/kg body weight (BW) in the TUDCA-TAC group reduced ERS markers (GRP78, p-PERK, and p-eIf2α), compared to the Vehicle (Veh)-TAC group. TUDCA administration, for 4 weeks after TAC significantly reduced cardiac hypertrophy as shown by the reduced heart weight (HW) to BW ratio, and expression of hypertrophic marker genes (ANF, BNP, and α-SKA). Masson's trichrome staining showed that myocardial fibrosis and collagen deposition were also significantly reduced in the TUDCA-TAC group. We also found that TUDCA significantly decreased expression of TGF-β signaling proteins and collagen isoforms. TUDCA administration also reduced cardiac apoptosis and the related proteins in the TUDCA-TAC group. Microarray analysis followed by gene ontology (GO) and pathway analysis demonstrated that extracellular matrix genes responsible for hypertrophy and fibrosis, and mitochondrial genes responsible for apoptosis and fatty acid metabolism were significantly altered in the Veh-TAC group, but the alterations were normalized in the TUDCA-TAC group, suggesting potential of TUDCA in treatment of heart diseases related to pressure-overload.

MicroRNA (miRNA) is an endogenous non-coding RNA species that either inhibits RNA translation or promotes degradation of target mRNAs. miRNAs often regulate cellular signaling by targeting multiple genes within the pathways. In the present study, using Gene Set Analysis, a useful bioinformatics tool to identify miRNAs with multiple target genes in the same pathways, we identified miR-185 as a key candidate regulator of cardiac hypertrophy. Using a mouse model, we found that miR-185 was significantly down-regulated in myocardial cells during cardiac hypertrophy induced by transverse aortic constriction. To confirm that miR-185 is an anti-hypertrophic miRNA, genetic manipulation studies such as overexpression and knock-down of miR-185 in neonatal rat ventricular myocytes were conducted. The results showed that up-regulation of miR-185 led to anti-hypertrophic effects, while down-regulation led to pro-hypertrophic effects, suggesting that miR-185 has an anti-hypertrophic role in the heart. Our study further identified Camk2d, Ncx1, and Nfatc3 as direct targets of miR-185. The activity of Nuclear Factor of Activated T-cell (NFAT) and calcium/calmodulin-dependent protein kinase II delta (CaMKIIδ) was negatively regulated by miR-185 as assessed by NFAT-luciferase activity and western blotting. The expression of phospho-phospholamban (Thr-17), a marker of CaMKIIδ activity, was also significantly reduced by miR-185. In conclusion, miR-185 effectively blocked cardiac hypertrophy signaling through multiple targets, rendering it a potential drug target for diseases such as heart failure.

Prolonged ER stress (ERS) can be associated with the induction of apoptotic cell death in various heart diseases. In this study, we searched for microRNAs affecting ERS in the heart using in silico and in vitro methods. We found that miR-185 directly targets the 3′-untranslated region of Na+/H+ exchanger-1 (NHE-1), a protein involved in ERS. Cardiomyocyte ERS-triggered apoptosis induced by 100 ng/ml tunicamycin (TM) or 1 μM thapsigargin (TG), ERS inducers, was significantly reduced by miR-185 overexpression. Protein expression of pro-apoptotic markers such as CCAAT/enhancer-binding protein homologous protein (CHOP) and cleaved-caspase-3 was also markedly reduced by miR-185 in a dose-dependent manner. Cariporide (20 μM), a pharmacological inhibitor of NHE-1, also attenuated ERS-induced apoptosis in cardiomyocytes and CHOP protein expression, suggesting that NHE-1 plays an important role in ERS-associated apoptosis in cardiomyocytes. Collectively, the present results demonstrate that miR-185 is involved in cardio-protection against ERS-mediated apoptotic cell death. [BMB Reports 2016; 49(4): 208-213]