Adipose Mesenchymal Extracellular Vesicles as Alpha-1-Antitrypsin Physiological Delivery Systems for Lung Regeneration

Accumulating evidence shows that Mesenchymal Stem/Stromal Cells (MSCs) exert their therapeutic effects by the release of secretome, made of both soluble proteins and nano/microstructured extracellular vesicles (EVs). In this work, for the first time, we proved by a proteomic investigation that adipose-derived (AD)-MSC-secretome contains alpha-1-antitrypsin (AAT), the main elastase inhibitor in the lung, 72 other proteins involved in protease/antiprotease balance, and 46 proteins involved in the response to bacteria. By secretome fractionation, we proved that AAT is present both in the soluble fraction of secretome and aggregated and/or adsorbed on the surface of EVs, that can act as natural carriers promoting AAT in vivo stability and activity. To modulate secretome composition, AD-MSCs were cultured in different stimulating conditions, such as serum starvation or chemicals (IL-1β and/or dexamethasone) and the expression of the gene encoding for AAT was increased. By testing in vitro the anti-elastase activity of MSC-secretome, a dose-dependent effect was observed; chemical stimulation of AD-MSCs did not increase their secretome anti-elastase activity. Finally, MSC-secretome showed anti-bacterial activity on Gram-negative bacteria, especially for Klebsiella pneumoniae. These preliminary results, in addition to the already demonstrated immunomodulation, pave the way for the use of MSC-secretome in the treatment of AAT-deficiency lung diseases.

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The Therapeutic Effect of Extracellular Vesicles on Asthma in Pre-Clinical Models: A Systematic Review Protocol

Journal of Respiration ◽

10.3390/jor1010009 ◽

2021 ◽

Vol 1 (1) ◽

pp. 84-95

Author(s):

Patience O. Obi ◽

Jennifer E. Kent ◽

Maya M. Jeyaraman ◽

Nicole Askin ◽

Taiana M. Pierdoná ◽

...

Keyword(s):

Systematic Review ◽

Extracellular Vesicles ◽

Current Knowledge ◽

Grey Literature ◽

Specific Treatment ◽

Therapeutic Effects ◽

Management Options ◽

Paracrine Signalling

Asthma is the most common pediatric disease, characterized by chronic airway inflammation and airway hyperresponsiveness. There are several management options for asthma, but no specific treatment. Extracellular vesicles (EVs) are powerful cellular mediators of endocrine, autocrine and paracrine signalling, and can modulate biophysiological function in vitro and in vivo. A thorough investigation of therapeutic effects of EVs in asthma has not been conducted. Therefore, this systematic review is designed to synthesize recent literature on the therapeutic effects of EVs on physiological and biological outcomes of asthma in pre-clinical studies. An electronic search of Web of Science, EMBASE, MEDLINE, and Scopus will be conducted on manuscripts published in the last five years that adhere to standardized guidelines for EV research. Grey literature will also be included. Two reviewers will independently screen the selected studies for title and abstract, and full text based on the eligibility criteria. Data will be extracted, narratively synthesized and reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. This systematic review will summarize the current knowledge from preclinical studies investigating the therapeutic effects of EVs on asthma. The results will delineate whether EVs can mitigate biological hallmarks of asthma, and if so, describe the underlying mechanisms involved in the process. This insight is crucial for identifying key pathways that can be targeted to alleviate the burden of asthma. The data will also reveal the origin, dosage and biophysical characteristics of beneficial EVs. Overall, our results will provide a scaffold for future intervention and translational studies on asthma treatment.

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Novel and Modified Neutrophil Elastase Inhibitor Loaded in Topical Formulations for Psoriasis Management

Pharmaceutics ◽

10.3390/pharmaceutics12040358 ◽

2020 ◽

Vol 12 (4) ◽

pp. 358 ◽

Cited By ~ 1

Author(s):

Andreia Nunes ◽

Joana Marto ◽

Lídia Maria Gonçalves ◽

Sandra Simões ◽

Rita Félix ◽

...

Keyword(s):

Serine Protease ◽

Neutrophil Elastase ◽

In Vitro Cytotoxicity ◽

Human Neutrophil Elastase ◽

Therapeutic Effects ◽

Neutrophil Elastase Inhibitor ◽

Analytical Methodology ◽

Elastase Inhibitor

Human neutrophil elastase (HNE) is a serine protease that degrades matrix proteins. An excess of HNE may trigger several pathological conditions, such as psoriasis. In this work, we aimed to synthesize, characterize and formulate new HNE inhibitors with a 4-oxo-β-lactam scaffold with less toxicity, as well as therapeutic index in a psoriasis context. HNE inhibitors with 4-oxo-β-lactam scaffolds were synthesized and characterized by NMR, FTIR, melting point, mass spectrometry and elemental analysis. In vitro cytotoxicity and serine protease assays were performed. The compound with the highest cell viability (AAN-16) was selected to be incorporated in an emulsion (AAN-16 E) and in a microemulsion (AAN-16 ME). Formulations were characterized in terms of organoleptic properties, pH, rheology, droplet size distribution, in vitro drug release and in vivo psoriatic activity. All compounds were successfully synthesized according to analytical methodology, with good yields. Both formulations presented suitable physicochemical properties. AAN-16 E presented the most promising therapeutic effects in a murine model of psoriasis. Overall, new HNE inhibitors were synthesized with high and selective activity and incorporated into topical emulsions with potential to treat psoriasis.

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Therapeutic Effects of Hypoxic and Pro-Inflammatory Priming of Mesenchymal Stem Cell-Derived Extracellular Vesicles in Inflammatory Arthritis

International Journal of Molecular Sciences ◽

10.3390/ijms23010126 ◽

2021 ◽

Vol 23 (1) ◽

pp. 126

Author(s):

Alasdair G. Kay ◽

Kane Treadwell ◽

Paul Roach ◽

Rebecca Morgan ◽

Rhys Lodge ◽

...

Keyword(s):

T Cell ◽

Extracellular Vesicles ◽

Inflammatory Responses ◽

Therapeutic Potential ◽

Histopathological Analysis ◽

Therapeutic Effects ◽

Synovial Joint ◽

Paracrine Signalling

Mesenchymal stem cells (MSCs) immunomodulate inflammatory responses through paracrine signalling, including via secretion of extracellular vesicles (EVs) in the cell secretome. We evaluated the therapeutic potential of MSCs-derived small EVs in an antigen-induced model of arthritis (AIA). EVs isolated from MSCs cultured normoxically (21% O2, 5% CO2), hypoxically (2% O2, 5% CO2) or with a pro-inflammatory cytokine cocktail were applied into the AIA model. Disease pathology was assessed post-arthritis induction through swelling and histopathological analysis of synovial joint structure. Activated CD4+ T cells from healthy mice were cultured with EVs or MSCs to assess deactivation capabilities prior to application of standard EVs in vivo to assess T cell polarisation within the immune response to AIA. All EVs treatments reduced knee-joint swelling whilst only normoxic and pro-inflammatory primed EVs improved histopathological outcomes. In vitro culture with EVs did not achieve T cell deactivation. Polarisation towards CD4+ helper cells expressing IL17a (Th17) was reduced when normoxic and hypoxic EV treatments were applied in vitro. Normoxic EVs applied into the AIA model reduced Th17 polarisation and improved Regulatory T cell (Treg):Th17 homeostatic balance. Normoxic EVs present the optimal strategy for broad therapeutic benefit. EVs present an effective novel technology with the potential for cell-free therapeutic translation.

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Mesenchymal Stem Cell-Derived Extracellular Vesicles and Their Therapeutic Potential

Stem Cells International ◽

10.1155/2020/8825771 ◽

2020 ◽

Vol 2020 ◽

pp. 1-10

Author(s):

Ashley G. Zhao ◽

Kiran Shah ◽

Brett Cromer ◽

Huseyin Sumer

Keyword(s):

Tissue Engineering ◽

Regenerative Medicine ◽

Extracellular Vesicles ◽

Therapeutic Potential ◽

Target Cells ◽

Therapeutic Effects ◽

Multipotent Cells ◽

Membrane Bound

Extracellular vesicles (EVs) are cell-derived membrane-bound nanoparticles, which act as shuttles, delivering a range of biomolecules to diverse target cells. They play an important role in maintenance of biophysiological homeostasis and cellular, physiological, and pathological processes. EVs have significant diagnostic and therapeutic potentials and have been studied both in vitro and in vivo in many fields. Mesenchymal stem cells (MSCs) are multipotent cells with many therapeutic applications and have also gained much attention as prolific producers of EVs. MSC-derived EVs are being explored as a therapeutic alternative to MSCs since they may have similar therapeutic effects but are cell-free. They have applications in regenerative medicine and tissue engineering and, most importantly, confer several advantages over cells such as lower immunogenicity, capacity to cross biological barriers, and less safety concerns. In this review, we introduce the biogenesis of EVs, including exosomes and microvesicles. We then turn more specifically to investigations of MSC-derived EVs. We highlight the great therapeutic potential of MSC-derived EVs and applications in regenerative medicine and tissue engineering.

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Enhancement of therapeutic potential of mesenchymal stem cell-derived extracellular vesicles

Stem Cell Research & Therapy ◽

10.1186/s13287-019-1398-3 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 45

Author(s):

Kyong-Su Park ◽

Elga Bandeira ◽

Ganesh V. Shelke ◽

Cecilia Lässer ◽

Jan Lötvall

Keyword(s):

Cell Therapy ◽

Extracellular Vesicles ◽

Therapeutic Potential ◽

Biological Activities ◽

Therapeutic Effects ◽

In Vivo Models ◽

Secreted Factors ◽

Associated Proteins

Abstract After the initial investigations into applications of mesenchymal stem cells (MSCs) for cell therapy, there was increased interest in their secreted soluble factors. Following studies of MSCs and their secreted factors, extracellular vesicles (EVs) released from MSCs have emerged as a new mode of intercellular crosstalk. MSC-derived EVs have been identified as essential signaling mediators under both physiological and pathological conditions, and they appear to be responsible for many of the therapeutic effects of MSCs. In several in vitro and in vivo models, EVs have been observed to have supportive functions in modulating the immune system, mainly mediated by EV-associated proteins and nucleic acids. Moreover, stimulation of MSCs with biophysical or biochemical cues, including EVs from other cells, has been shown to influence the contents and biological activities of subsequent MSC-derived EVs. This review provides on overview of the contents of MSC-derived EVs in terms of their supportive effects, and it provides different perspectives on the manipulation of MSCs to improve the secretion of EVs and subsequent EV-mediated activities. In this review, we discuss the possibilities for manipulating MSCs for EV-based cell therapy and for using EVs to affect the expression of elements of interest in MSCs. In this way, we provide a clear perspective on the state of the art of EVs in cell therapy focusing on MSCs, and we raise pertinent questions and suggestions for knowledge gaps to be filled.

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Extracellular vesicles mediate the intercellular exchange of nanoparticles

10.1101/2021.02.23.432325 ◽

2021 ◽

Author(s):

Xian Wu ◽

Tang Tang ◽

Yushuang Wei ◽

Katherine A. Cummins ◽

David K. Wood ◽

...

Keyword(s):

Extracellular Vesicles ◽

Cell Contact ◽

Therapeutic Effects ◽

Cellular Receptors ◽

Transcellular Transport ◽

Direct Cell ◽

Intercellular Exchange

AbstractIn order to exert their therapeutic effects, nanoparticles (NPs) often need to travel into the tissues composed of multilayered cells. Accumulative evidence has revealed the central role of transcellular transport route (entry into one cell, exocytosis, and re-entry into another) in this process. While NP endocytosis and subcellular transport have been intensively characterized, the exocytosis and re-entry steps are poorly understood, which becomes a barrier to improve NP delivery into complex tissues. Here, we termed the exocytosis and re-entry steps together as intercellular exchange. We developed a novel collagen-based 3D cellular assay to specifically monitor and quantify the intercellular exchange events of NPs and distinguish the contributions of several potential mechanisms. Our results showed that NPs can be exocytosed freely or enclosed inside extracellular vesicles (EVs) for re-entry, while direct cell-cell contact is hardly involved. EVs account for a significant fraction of NP intercellular exchange, and its importance in NP delivery was demonstrated in vitro and in vivo. Intriguingly, while freely released NPs engage with the same cellular receptors for re-entry, EV-enclosed ones bypass this dependence. These studies provide an easy and precise system to investigate the intercellular exchange stage of NP delivery, and shed the first light in the importance of EVs in NP transport between cells and across complex tissues.

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THERAPEUTIC EFFECTS OF HYPOXIC AND PRO-INFLAMMATORY PRIMING OF MESENCHYMAL STEM CELL-DERIVED EXTRACELLULAR VESICLES IN INFLAMMATORY ARTHRITIS

10.1101/2021.06.28.450178 ◽

2021 ◽

Author(s):

Alasdair G Kay ◽

Kane Treadwell ◽

Paul Roach ◽

Rebecca Morgan ◽

Rhys Lodge ◽

...

Keyword(s):

T Cell ◽

Extracellular Vesicles ◽

Inflammatory Responses ◽

Therapeutic Potential ◽

Histopathological Analysis ◽

Therapeutic Effects ◽

Synovial Joint ◽

Paracrine Signalling

Novel biological therapies have revolutionised the management of Rheumatoid Arthritis (RA) but no cure currently exists. Mesenchymal stem cells (MSCs) immunomodulate inflammatory responses through paracrine signalling, including via secretion of extracellular vesicles (EVs) in the cell secretome. We evaluated the therapeutic potential of MSCs-derived small EVs in an antigen-induced model of arthritis (AIA). EVs isolated from MSCs cultured normoxically (21% O2, 5% CO2), hypoxically (2% O2, 5% CO2) or with a pro-inflammatory cytokine cocktail were applied into the AIA model. Disease pathology was assessed post-arthritis induction through swelling and histopathological analysis of synovial joint structure. Activated CD4+ T cells from healthy mice were cultured with EVs or MSCs to assess deactivation capabilities prior to application of standard EVs in vivo to assess T cell polarisation within the immune response to AIA. All EVs treatments reduced knee-joint swelling whilst only normoxic and pro-inflammatory primed EVs improved histopathological outcomes. In vitro culture with EVs did not achieve T cell deactivation. Polarisation towards CD4+ helper cells expressing IL17a (Th17) was reduced when normoxic and hypoxic EV treatments were applied in vitro. Normoxic EVs applied into the AIA model reduced Th17 polarisation and improved Th17:Treg homeostatic balance. Priming of MSCs in EV production can be applied to alter the therapeutic efficacy however normoxic EVs present the optimal strategy for broad therapeutic benefit. The varied outcomes observed in MSCs priming may promote EVs optimised for therapies targeted for specific therapeutic priorities. EVs present an effective novel technology with potential for cell-free therapeutic translation.

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Clinical Trials in Thrombosis: Secondary Prevention of Myocardial Infarction

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647910 ◽

1976 ◽

Vol 35 (01) ◽

pp. 049-056 ◽

Cited By ~ 4

Author(s):

Christian R Klimt ◽

P. H Doub ◽

Nancy H Doub

Keyword(s):

Myocardial Infarction ◽

Secondary Prevention ◽

Case Control ◽

Therapeutic Effects ◽

Case Control Studies ◽

Definitive Answer ◽

Inhibition Of Platelet Aggregation ◽

Prospective Trials

SummaryNumerous in vivo and in vitro experiments, investigating the inhibition of platelet aggregation and the prevention of experimentally-induced thrombosis, suggest that anti-platelet drugs, such as aspirin or the combination of aspirin and dipyridamole or sulfinpyrazone, may be effective anti-thrombotic agents in man. Since 1971, seven randomized prospective trials and two case-control studies have been referenced in the literature or are currently being conducted, which evaluate the effects of aspirin, sulfinpyrazone, or dipyridamole in combination with aspirin in the secondary prevention of myocardial infarction. A critical review of these trials indicates a range of evidence from no difference to a favorable trend that antiplatelet drugs may serve as anti-thrombotic agents in man. To date, a definitive answer concerning the therapeutic effects of these drugs in the secondary prevention of coronary heart disease is not available.

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A Systematic Review on Extracellular Vesicles-Enriched Fat Grafting: A Shifting Paradigm

Aesthetic Surgery Journal ◽

10.1093/asj/sjaa362 ◽

2020 ◽

Author(s):

Mohammad Ghiasloo ◽

Laura De Wilde ◽

Kashika Singh ◽

Patrick Tonnard ◽

Alexis Verpaele ◽

...

Keyword(s):

Systematic Review ◽

Graft Survival ◽

Extracellular Vesicles ◽

Retention Rate ◽

Fat Grafting ◽

Retention Rates ◽

Fat Graft ◽

Cochrane Central Register

Abstract Background Recent evidence confirms that mesenchymal stem cells (MSCs) facilitate angiogenesis mainly through paracrine function. Extracellular vesicles (EVs) are regarded as key components of the cell secretome, possessing functional properties of their source cells. Subsequently, MSC-EVs have emerged as a novel cell-free approach to improve fat graft retention rate. Objectives To provide a systematic review of all studies reporting the use of MSC-EVs to improve graft retention rate. Methods A systematic search was undertaken using the Embase, PubMed and the Cochrane Central Register of Controlled Trials databases. Outcome measures included donor/receptor organism of the fat graft, study model, intervention groups, evaluation intervals, EV research data, in vitro and in vivo results. Results Of the total 1717 articles, 62 full-texts were screened. Seven studies reporting on 294mice were included. Overall, EV treated groups showed higher graft retention rates compared to untreated groups. Notably, retention rate was similar following EV- and MSC-treatment. In addition to reduced inflammation, graft enrichment with EVs resulted in early revascularization and better graft integrity. Interestingly, hypoxic preconditioning of MSCs improved their beneficial paracrine effects and led to a more proangiogenic EV population, as observed by both in vitro and in vivo results. Conclusions MSC-EVs appear to offer an interesting cell-free alternative to improve fat graft survival. While their clinical relevance remains to be determined, it is clear that not the cells, but their secretome is essential for graft survival. Thus, a paradigm shift from cell-assisted lipotransfer towards ‘secretome-assisted lipotransfer’ is well on its way.

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microRNA-186 in extracellular vesicles from bone marrow mesenchymal stem cells alleviates idiopathic pulmonary fibrosis via interaction with SOX4 and DKK1

Stem Cell Research & Therapy ◽

10.1186/s13287-020-02083-x ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Jing Zhou ◽

Yang Lin ◽

Xiuhua Kang ◽

Zhicheng Liu ◽

Wei Zhang ◽

...

Keyword(s):

Bone Marrow ◽

Idiopathic Pulmonary Fibrosis ◽

Pulmonary Fibrosis ◽

Extracellular Vesicles ◽

Luciferase Assay ◽

Therapeutic Effects ◽

Loss Of Function ◽

Fibroblast Activation ◽

Promising Therapeutic Approach

Abstract Background Previous reports have identified that human bone marrow mesenchymal stem cell-derived extracellular vesicles (BMSC-EVs) with their cargo microRNAs (miRNAs) are a promising therapeutic approach for the treatment of idiopathic pulmonary fibrosis (IPF). Therefore, we explored whether delivery of microRNA-186 (miR-186), a downregulated miRNA in IPF, by BMSC EVs could interfere with the progression of IPF in a murine model. Methods In a co-culture system, we assessed whether BMSC-EVs modulated the activation of fibroblasts. We established a mouse model of PF to evaluate the in vivo therapeutic effects of BMSC-EVs and determined miR-186 expression in BMSC-EVs by polymerase chain reaction. Using a loss-of-function approach, we examined how miR-186 delivered by BMSC-EVs affected fibroblasts. The putative relationship between miR-186 and SRY-related HMG box transcription factor 4 (SOX4) was tested using luciferase assay. Next, we investigated whether EV-miR-186 affected fibroblast activation and PF by targeting SOX4 and its downstream gene, Dickkopf-1 (DKK1). Results BMSC-EVs suppressed lung fibroblast activation and delayed IPF progression in mice. miR-186 was downregulated in IPF but enriched in the BMSC-EVs. miR-186 delivered by BMSC-EVs could suppress fibroblast activation. Furthermore, miR-186 reduced the expression of SOX4, a target gene of miR-186, and hence suppressed the expression of DKK1. Finally, EV-delivered miR-186 impaired fibroblast activation and alleviated PF via downregulation of SOX4 and DKK1. Conclusion In conclusion, miR-186 delivered by BMSC-EVs suppressed SOX4 and DKK1 expression, thereby blocking fibroblast activation and ameliorating IPF, thus presenting a novel therapeutic target for IPF.

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