The Dual Nature of Mesenchymal Stem Cells (MSCs): Yin and Yang of the Inflammatory Process

The well-known reparative properties of mesenchymal stem cells (MSCs) make them an attractive source for cell-based therapy. In vitro and in vivo studies support an anti-inflammatory role of MSCs by directly targeting immune cells or via the secretion of immunomodulatory factors. MSCs have been isolated from several human normal tissues, even from pathological biopsies and blood samples; in these cases, MSCs displayed peculiar characteristics, suggesting a phenotype transition into a pathological state. Indeed, MSCs derived from inflamed tissues acquired a pro-inflammatory behaviour. In this view, MSCs may be crucial players of many pathways involved in human diseases, especially during the inflammatory cascade. The present chapter will minutely describe the basic biology of human MSCs derived from normal and pathological arteries, focusing on their dual nature as cellular switchers of the inflammatory setting. We will also discuss the emerging role of miRNAs in regulating MSC functions and their potential use as alternative strategies to manipulate MSC efficacy

The characteristics and survival potential under sub-lethal stress of Mesenchymal Stromal/Stem Cells isolated from the human vascular wall

Mesenchymal stromal/stem cells (MSCs) have been identified in multiple human tissues, including the vascular wall. High proliferative potential, multilineage, and immunomodulatory properties make vascular MSCs promising candidates for regenerative medicine. Indeed, their location is strategic for controlling vascular and extra-vascular tissue homeostasis. However, the clinical application of MSCs, and in particular vascular MSCs, is still challenging. Current studies are focused on developing strategies to improve MSC therapeutic applications, like priming MSCs with stress conditions (hypoxia, nutrient deprivation) to achieve a higher therapeutic potential. The goal of the present study is to review the main findings regarding the MSCs isolated from the human vascular wall. Further, the main priming strategies tested on MSCs from different sources are reported, together with the experience on vascular MSCs isolated from healthy cryopreserved and pathological arteries. Stress induction can be a priming approach able to improve MSC effectiveness through several mechanisms that are discussed in this review. Nevertheless, these issues have not been completely explored in vascular MSCs and potential side effects need to be investigated

Contribution of vascular resident mesenchymal stromal cells to abdominal aortic aneurysm pathogenesis: increased MMP-9 expression and ineffective immunomodulation

Background. Ageing and inflammation are critical for the occurrence of aortic diseases. Extensive inflammatory infiltrate and excessive ECM proteloysis, mediated by MMPs, are typical features of abdominal aortic aneurysm (AAA). Mesenchymal Stromal Cells (MSCs) have been detected within the vascular wall and represent attractive candidates for regenerative medicine, in virtue of mesodermal lineage differentiation and immunomodulatory activity. Meanwhile, many works have underlined an impaired MSC behaviour under pathological conditions. This study was aimed to define a potential role of vascular MSCs to AAA development. Methods. Aortic tissues were collected from AAA patients and healthy donors. Our analysis was organized on three levels: 1) histology of AAA wall; 2) detection of MSCs and evaluation of MMP-9 expression on AAA tissue; 3) MSC isolation from AAA wall and characterization for mesenchymal/stemness markers, MMP-2, MMP-9, TIMP-1, TIMP-2 and EMMPRIN. AAA-MSCs were tested for immunomodulation, when cultured together with activated peripheral blood mononuclear cells (PBMCs). In addition, a co-colture of both healthy and AAA MSCs was assessed and afterwards MMP-2/9 mRNA levels were analyzed. Results. AAA-MSCs showed basic mesenchymal properties: fibroblastic shape, MSC antigens, stemness genes. MMP-9 mRNA, protein and enzymatic activity were significantly increased in AAA-MSCs. Moreover, AAA-MSCs displayed a weak immunosuppressive activity, as shown by PBMC ongoing along cell cycle. MMP-9 was shown to be modulated at the transcriptional level through the direct contact as well as the paracrine action of healthy MSCs. Discussion. Vascular injury did not affect the MSC basic phenotype, but altered their function, a increased MMP-9 expression and ineffective immunmodulation. These data suggest that vascular MSCs can contribute to aortic disease. In this view, the study of key processes to restore MSC immunomodulation could be relevant to find a pharmacological approach for monitoring the aneurysm progression

Impact of Freeze-Drying on Cord Blood (CB), Serum (S), and Platelet-Rich Plasma (CB-PRP) Preparations on Growth Factor Content and In Vitro Cell Wound Healing

Blood-based preparations are used in clinical practice for the treatment of several eye disorders. The aim of this study is to analyze the effect of freeze-drying blood-based preparations on the levels of growth factors and wound healing behaviors in an in vitro model. Platelet-rich plasma (PRP) and serum (S) preparations from the same Cord Blood (CB) sample, prepared in both fresh frozen (FF) and freeze-dried (FD) forms (and then reconstituted), were analyzed for EGF and BDNF content (ELISA Quantikine kit). The human MIO-M1 glial cell line (Moorfield/Institute of Ophthalmology, London, UK) was incubated with FF and FD products and evaluated for cell migration with scratch-induced wounding (IncuCyte S3 Essen BioScience), proliferation with cyclin A2 and D1 gene expression, and activation with vimentin and GFAP gene expression. The FF and FD forms showed similar concentrations of EGF and BDNF in both the S and PRP preparations. The wound healing assay showed no significant difference between the FF and FD forms for both S and PRP. Additionally, cell migration, proliferation, and activation did not appear to change in the FD forms compared to the FF ones. Our study showed that reconstituted FD products maintained the growth factor concentrations and biological properties of FF products and could be used as a functional treatment option

Human glial müller and umbilical vein endothelial cell coculture as an in vitro model to investigate retinal oxidative damage. A morphological and molecular assessment

The aim of this study was to optimize a coculture in vitro model established between the human Muller glial cells and human umbilical vein endothelial cells, mimicking the inner blood-retinal barrier, and to explore its resistance to damage induced by oxidative stress. A spontaneously immortalized human Muller cell line MIO-M1 and human umbilical vein endothelial cells (HUVEC) were plated together at a density ratio 1:1 and maintained up to the 8th passage (p8). The MIO-M1/HUVECs p1 through p8 were treated with increasing concentrations (range 200-800 mu M) of H2O2 to evaluate oxidative stress induced damage and comparing data with single cell cultures. The following features were assayed p1 through p8: doubling time maintenance, cell viability using MTS assay, ultrastructure of cell-cell contacts, immunofluorescence for Vimentin and GFAP, molecular biology (q-PCR) for GFAP and CD31 mRNA. MIO-M1/HUVECs cocultures maintained distinct cell cytotype up to p8 as shown by flow cytometry analysis, without evidence of cross activation, displaying cell-cell tight junctions mimicking those found in human retina, only acquiring a slight resistance to oxidative stress induction over the passages. This MIO-M1/HUVECs coculture represents a simple, reproducible and affordable model for in vitro studies on oxidative stress-induced retinal damages

Human cadaver multipotent stromal/stem cells isolated from arteries stored in liquid nitrogen for 5 years

Introduction: Regenerative medicine challenges researchers to find noncontroversial, safe and abundant stem cell sources. In this context, harvesting from asystolic donors could represent an innovative and unlimited reservoir of different stem cells. In this study, cadaveric vascular tissues were established as an alternative source of human cadaver mesenchymal stromal/stem cells (hC-MSCs). We reported the successful cell isolation from postmortem arterial segments stored in a tissue-banking facility for at least 5 years. Methods: After thawing, hC-MSCs were isolated with a high efficiency (12 × 106) and characterized with flow cytometry, immunofluorescence, molecular and ultrastructural approaches. Results: In early passages, hC-MSCs were clonogenic, highly proliferative and expressed mesenchymal (CD44, CD73, CD90, CD105, HLA-G), stemness (Stro-1, Oct-4, Notch-1), pericyte (CD146, PDGFR-β, NG2) and neuronal (Nestin) markers; hematopoietic and vascular markers were negative. These cells had colony and spheroid-forming abilities, multipotency for their potential to differentiate in multiple mesengenic lineages and immunosuppressive activity to counteract proliferation of phytohemagglutinin-stimulated blood mononuclear cells. Conclusions: The efficient procurement of stem cells from cadaveric sources, as postmortem vascular tissues, demonstrates that such cells can survive to prolonged ischemic insult, anoxia, freezing and dehydration injuries, thus paving the way for a scientific revolution where cadaver stromal/stem cells could effectively treat patients demanding cell therapies

The PPAR-γ Agonist Pioglitazone Modulates Proliferation and Migration in HUVEC, HAOSMC and Human Arteriovenous Fistula-Derived Cells

The failure of arteriovenous fistulas (AVFs) following intimal hyperplasia (IH) increases morbidity and mortality rates in patients undergoing hemodialysis for chronic kidney disease. The peroxisome-proliferator associated receptor (PPAR-γ) may be a therapeutic target in IH regulation. In the present study, we investigated PPAR-γ expression and tested the effect of pioglitazone, a PPAR-γ agonist, in different cell types involved in IH. As cell models, we used Human Endothelial Umbilical Vein Cells (HUVEC), Human Aortic Smooth Muscle Cells (HAOSMC), and AVF cells (AVFCs) isolated from (i) normal veins collected at the first AVF establishment (T0), and (ii) failed AVF with IH (T1). PPAR-γ was downregulated in AVF T1 tissues and cells, in comparison to T0 group. HUVEC, HAOSMC, and AVFC (T0 and T1) proliferation and migration were analyzed after pioglitazone administration, alone or in combination with the PPAR-γ inhibitor, GW9662. Pioglitazone negatively regulated HUVEC and HAOSMC proliferation and migration. The effect was antagonized by GW9662. These data were confirmed in AVFCs T1, where pioglitazone induced PPAR- γ expression and downregulated the invasive genes SLUG, MMP-9, and VIMENTIN. In summary, PPAR-γ modulation may represent a promising strategy to reduce the AVF failure risk by modulating cell proliferation and migration

Was the cold European winter 2009-2010 modified by anthropogenic climate change? An attribution study

An attribution study has been performed to investigate the degree to which the unusually cold European winter 2009-2010 was modified by anthropogenic climate change. Two different methods have been included for the attribution: one based on a large HadGEM3-A ensemble and one based on a statistical surrogate method. Both methods are evaluated by comparing simulated winter temperature means, trends, standard deviations, skewness, return periods, and 5 % quantiles with observations. While the surrogate method performs well, HadGEM3-A in general underestimates the trend in winter by a factor of 2/3. It has a mean cold bias dominated by the mountainous regions and also underestimates the cold 5 % quantile in many regions of Europe. Both methods show that the probability of experiencing a winter as cold as 2009-2010 has been reduced by approximately a factor of two due to anthropogenic changes. The method based on HadGEM3-A ensembles gives somewhat larger changes than the surrogate method because of differences in the definition of the unperturbed climate. The results are based on two diagnostics: the coldest day in winter and the largest continuous area with temperatures colder than twice the local standard deviation. The results are not sensitive to the choice of bias correction except in the mountainous regions. Previous results regarding the behavior of the measures of the changed probability have been extended. The counter-intuitive behavior for heavy-tailed distributions is found to hold for a range of measures and for events that become more rare in a changed climate

Evaluation of the HadGEM3-A simulations in view of detection and attribution of human influence on extreme events in Europe

A detailed analysis is carried out to assess the HadGEM3-A global atmospheric model skill in simulating extreme temperatures, precipitation and storm surges in Europe in the view of their attribution to human influence. The analysis is performed based on an ensemble of 15 atmospheric simulations forced with observed Sea Surface Temperature of the 54 year period 1960-2013. These simulations, together with dual simulations without human influence in the forcing, are intended to be used in weather and climate event attribution. The analysis investigates the main processes leading to extreme events, including atmospheric circulation patterns, their links with temperature extremes, land-atmosphere and troposphere-stratosphere interactions. It also compares observed and simulated variability, trends and generalized extreme value theory parameters for temperature and precipitation. One of the most striking findings is the ability of the model to capture North Atlantic atmospheric weather regimes as obtained from a cluster analysis of sea level pressure fields. The model also reproduces the main observed weather patterns responsible for temperature and precipitation extreme events. However, biases are found in many physical processes. Slightly excessive drying may be the cause of an overestimated summer interannual variability and too intense heat waves, especially in central/northern Europe. However, this does not seem to hinder proper simulation of summer temperature trends. Cold extremes appear well simulated, as well as the underlying blocking frequency and stratosphere-troposphere interactions. Extreme precipitation amounts are overestimated and too variable. The atmospheric conditions leading to storm surges were also examined in the Baltics region. There, simulated weather conditions appear not to be leading to strong enough storm surges, but winds were found in very good agreement with reanalyses. The performance in reproducing atmospheric weather patterns indicates that biases mainly originate from local and regional physical processes. This makes local bias adjustment meaningful for climate change attribution

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