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Endothelial progenitor cells are known to reverse acute kidney injury by paracrine mechanisms. We previously found that microvesicles released from these progenitor cells activate an angiogenic program in endothelial cells by horizontal mRNA transfer. Here, we tested whether these microvesicles prevent acute kidney injury in a rat model of ischemia–reperfusion injury. The RNA content of microvesicles was enriched in microRNAs (miRNAs) that modulate proliferation, angiogenesis, and apoptosis. After intravenous injection following ischemia–reperfusion, the microvesicles were localized within peritubular capillaries and tubular cells. This conferred functional and morphologic protection from acute kidney injury by enhanced tubular cell proliferation, reduced apoptosis, and leukocyte infiltration. Microvesicles also protected against progression of chronic kidney damage by inhibiting capillary rarefaction, glomerulosclerosis, and tubulointerstitial fibrosis. The renoprotective effect of microvesicles was lost after treatment with RNase, nonspecific miRNA depletion of microvesicles by Dicer knock-down in the progenitor cells, or depletion of pro-angiogenic miR-126 and miR-296 by transfection with specific miR-antagomirs. Thus, microvesicles derived from endothelial progenitor cells protect the kidney from ischemic acute injury by delivering their RNA content, the miRNA cargo of which contributes to reprogramming hypoxic resident renal cells to a regenerative program.

Several studies indicate that adult stem cells may improve the recovery from acute tissue injury. It has been suggested that they may contribute to tissue regeneration by the release of paracrine factors promoting proliferation of tissue resident cells. However, the factors involved remain unknown. In the present study we found that microvesicles (MVs) derived from human liver stem cells (HLSC) induced in vitro proliferation and apoptosis resistance of human and rat hepatocytes. These effects required internalization of MVs in the hepatocytes by an α4‐integrin‐dependent mechanism. However, MVs pre‐treated with RNase, even if internalized, were unable to induce hepatocyte proliferation and apoptosis resistance, suggesting an RNA‐dependent effect. Microarray analysis and quantitative RT‐PCR demonstrated that MVs were shuttling a specific subset of cellular mRNA, such as mRNA associated in the control of transcription, translation, proliferation and apoptosis. When administered in vivo, MVs accelerated the morphological and functional recovery of liver in a model of 70% hepatectomy in rats. This effect was associated with increase in hepatocyte proliferation and was abolished by RNase pre‐treatment of MVs. Using human AGO2, as a reporter gene present in MVs, we found the expression of human AGO2 mRNA and protein in the liver of hepatectomized rats treated with MVs. These data suggested a translation of the MV shuttled mRNA into hepatocytes of treated rats. In conclusion, these results suggest that MVs derived from HLSC may activate a proliferative program in remnant hepatocytes after hepatectomy by a horizontal transfer of specific mRNA subsets.

Background/Objectives: Soluble factors and cell-derived extracellular vesicles (EVs) are crucial tissue repair mediators in cell-based therapy. In the present study, we investigate the therapeutic impact of EVs released by adipose tissue-derived stem cells (ASCs) recovered from obese subjects’ visceral and subcutaneous tissues. Methods: ASCs were recovered from 10 obese (oASCs) and 6 non-obese (nASCs) participants and characterized. In selected experiments, nASCs and oASCs were cultured with palmitic acid (PA) or high glucose (HG), respectively. EVs from obese (oEVs) and non-obese (nEVs) subjects’ visceral and subcutaneous ASCs were collected after ultracentrifugation and analyzed for their cargo: microRNA-126 (miR-126), vascular endothelial growth factor (VEGF), and matrix metalloproteinase 2 (MMP-2), and for their biological effects on endothelial cells (ECs). Western blotting analysis and loss- and gain-of function experiments were performed. Results: oEVs show impaired angiogenic potential compared with nEVs. This effect depends on EV cargo: reduced content of VEGF, MMP-2 and, more importantly, miR-126. We demonstrate, using gain- and loss-of-function experiments, that this reduced miR-126 content leads to Spred1 upregulation and the inhibition of the extracellular signal–regulated kinase 1/2 mitogen-activated protein kinase pathway in ECs. We also show that PA treatment of nASCs translates into the release of EVs that recapitulate oEV cargo. Moreover, HG treatment of oASCs further reduces miR-126 EV content and EV-mediated in vitro angiogenesis. Finally, impaired pro-angiogenic potential is also detected in EVs released from obese subcutaneous adipose tissue-derived ASCs. Conclusions: These results indicate that obesity impacts on EV pro-angiogenic potential and may raise concerns about the use of adipose tissue-derived EVs in cell-based therapy in the obese setting.

Microvesicles (MVs) are circular fragments of membrane released from the endosomal compartment as exosomes or shed from the surface membranes of most cell types. An increasing body of evidence indicates that they play a pivotal role in cell-to-cell communication. Indeed, they may directly stimulate target cells by receptor-mediated interactions or may transfer from the cell of origin to various bioactive molecules including membrane receptors, proteins, mRNAs, microRNAs, and organelles. In this review we discuss the pleiotropic biologic effects of MVs that are relevant for communication among cells in physiological and pathological conditions. In particular, we discuss their potential involvement in inflammation, renal disease, and tumor progression, and the evidence supporting a bidirectional exchange of genetic information between stem and injured cells. The transfer of gene products from injured cells may explain stem cell functional and phenotypic changes without the need of transdifferentiation into tissue cells. On the other hand, transfer of gene products from stem cells may reprogram injured cells to repair damaged tissues.

Aims/hypothesis Mesenchymal stem cells (MSCs) have been shown to abrogate in vitro the proinflammatory response in type 1 diabetes. The mechanism involves paracrine factors, which may include microvesicles (MVs). We evaluated whether MVs derived from heterologous bone-marrow MSCs exert an immunomodulatory effect on T cell responses against GAD (glutamic acid decarboxylase) antigen in type 1 diabetes. Methods MVs were purified from heterologous human MSCs by differential centrifugation. Peripheral blood mononuclear cells (PBMCs) were obtained from patients with type 1 diabetes at disease onset, and responses to GAD65 stimulation were assessed by IFN-γ enzyme-linked immunosorbent spot analysis. Levels of cytokines and prostaglandin E 2 (PGE 2 ) were measured in the supernatant fraction, and T helper 17 (Th17) and regulatory T cell analysis was performed. Results MVs were internalised by PBMCs, as assessed by confocal microscopy and flow cytometry analyses. MVs significantly decreased IFN-γ spots and levels in GAD65-stimulated PBMCs, and significantly increased transforming growth factor-β (TGF-β), IL-10, IL-6 and PGE 2 levels. Furthermore, MVs decreased the number of Th17 cells and the levels of IL-17, and increased FoxP3 + regulatory T cells in GAD65-stimulated PBMCs. Conclusions/interpretation These results provide evidence that MSC-derived MVs can inhibit in vitro a proinflammatory response to an islet antigenic stimulus in type 1 diabetes. The action of MVs involves PGE 2 and TGF-β signalling pathways and IL-10 secretion, suggesting a switch to an anti-inflammatory response of T cells.

BACKGROUND/OBJECTIVES: Soluble factors and cell-derived extracellular vesicles (EVs) are crucial tissue repair mediators in cellbased therapy. In the present study, we investigate the therapeutic impact of EVs released by adipose tissue-derived stem cells (ASCs) recovered from obese subjects' visceral and subcutaneous tissues. METHODS: ASCs were recovered from 10 obese (oASCs) and 6 non-obese (nASCs) participants and characterized. In selected experiments, nASCs and oASCs were cultured with palmitic acid (PA) or high glucose (HG), respectively. EVs from obese (oEVs) and non-obese (nEVs) subjects' visceral and subcutaneous ASCs were collected after ultracentrifugation and analyzed for their cargo: microRNA-126 (miR-126), vascular endothelial growth factor (VEGF), and matrix metalloproteinase 2 (MMP-2), and for their biological effects on endothelial cells (ECs). Western blotting analysis and loss- and gain-of function experiments were performed. RESULTS: oEVs show impaired angiogenic potential compared with nEVs. This effect depends on EV cargo: reduced content of VEGF, MMP-2 and, more importantly, miR-126. We demonstrate, using gain- and loss-of-function experiments, that this reduced miR-126 content leads to Spred1 upregulation and the inhibition of the extracellular signal-regulated kinase 1/2 mitogen-activated protein kinase pathway in ECs. We also show that PA treatment of nASCs translates into the release of EVs that recapitulate oEV cargo. Moreover, HG treatment of oASCs further reduces miR-126 EV content and EV-mediated in vitro angiogenesis. Finally, impaired pro-angiogenic potential is also detected in EVs released from obese subcutaneous adipose tissue-derived ASCs. CONCLUSIONS: These results indicate that obesity impacts on EV pro-angiogenic potential and may raise concerns about the use of adipose tissue-derived EVs in cell-based therapy in the obese setting. International Journal of Obesity (2016) 40, 102-111; doi:10.1038/ijo.2015.123

The islet microcirculation has morphological characteristics resembling those of renal glomeruli. Transcription of the nephrin gene, a highly specific barrier protein of the slit diaphragm of podocyte foot processes, has been reported in the pancreas, although its cellular localisation and function remain to be defined. In this study, we purified and characterised microvascular endothelial cells (MECs) isolated from human islets and investigated the expression and distribution of nephrin on these cells. Human islet MECs were extracted and purified using anti-CD105-coated immunomagnetic beads and their endothelial characteristics were confirmed by expression of classical endothelial markers and basal high-level expression of intercellular adhesion molecule-1 and TNF-alpha-inducible vascular cell adhesion molecule-1. Nephrin expression was assessed by immunofluorescence, flow cytometric analysis and western blotting on cell lysates, as well as by RT-PCR. Immunofluorescence studies detected nephrin in a fine, punctate, diffuse pattern on cultured islet MECs, and also in human pancreatic islet sections. In both cases nephrin colocalised with endothelial markers. TNF-alpha treatment induced a marked reduction and redistribution of the protein in one or multiple aggregates. Nephrin expression was confirmed by flow cytometry, western blotting and RT-PCR studies. In contrast, nephrin could not be detected at the protein or mRNA level in human macro- and microvascular cells from other sites. Nephrin is expressed at protein and mRNA levels in islet microendothelium, supporting the hypothesis that islet MECs exhibit distinctive morphological characteristics that indicate functional specialisation of potential pathophysiological importance.

In Potter Cove, Antarctica, newly ice-free areas appeared due to glacial retreat. Simultaneously, the inflow of sediment increased, reducing underwater photosynthetically active radiation (PAR, 400–700 nm). The aim of this study was to determine the photosynthetic characteristics of two macroalgal species colonizing three newly ice-free areas, A1, A2 and A3, with increasing degree of glacial influence from A1 to A3. Turbidity, salinity and temperature were measured, and light attenuation coefficients ( K d ) calculated and considered as a proxy for glacial sediment input. The lower depth distribution of the red alga Palmaria decipiens and the brown alga Himantothallus grandifolius was 10 m in A3, 20 m in A2 and 30 m in A1. Both species were then collected, at 5 and 10 m at all areas. Photosynthetic parameters and the daily metabolic carbon balance (CB) were determined. K d was significantly higher in A3 compared with A1 and A2. The CB of P. decipiens was significantly higher in A1 followed by A2 and A3, and significantly higher at shallower than at greater depth. For H. grandifolius CB was significantly lower in A3 and in A2 at deeper depths compared with the rest of areas and depths. The lower distribution limit of the algae was positively correlated to the light penetration. An increase in the sediment run-off due to global warming might lead to an elevation of the lower depth distribution limit but retreating glaciers can open new space for macroalgal colonization. These changes will probably affect macroalgal primary productivity in Potter Cove with consequences for the coastal ecosystem.