scholarly journals Amniotic Fluid Stem Cell-Derived Extracellular Vesicles Counteract Steroid-Induced Osteoporosis In Vitro

2020 ◽  
Vol 22 (1) ◽  
pp. 38
Author(s):  
Martina Gatti ◽  
Francesca Beretti ◽  
Manuela Zavatti ◽  
Emma Bertucci ◽  
Soraia Ribeiro Luz ◽  
...  
Keyword(s):  
Amniotic Fluid ◽  
Extracellular Vesicles ◽  
Culture Medium ◽  
Cell Potential ◽  
Osteoblast Cell Line ◽  
Local Bone ◽  
Current Therapies ◽  
Amniotic Fluid Stem Cell ◽  
Related Proteins

Background—Osteoporosis is characterized by defects in both quality and quantity of bone tissue, which imply high susceptibility to fractures with limitations of autonomy. Current therapies for osteoporosis are mostly concentrated on how to inhibit bone resorption but give serious adverse effects. Therefore, more effective and safer therapies are needed that even encourage bone formation. Here we examined the effect of extracellular vesicles secreted by human amniotic fluid stem cells (AFSC) (AFSC-EV) on a model of osteoporosis in vitro. Methods—human AFSC-EV were added to the culture medium of a human pre-osteoblast cell line (HOB) induced to differentiate, and then treated with dexamethasone as osteoporosis inducer. Aspects of differentiation and viability were assessed by immunofluorescence, Western blot, mass spectrometry, and histological assays. Since steroids induce oxidative stress, the levels of reactive oxygen species and of redox related proteins were evaluated. Results—AFSC-EV were able to ameliorate the differentiation ability of HOB both in the case of pre-osteoblasts and when the differentiation process was affected by dexamethasone. Moreover, the viability was increased and parallelly apoptotic markers were reduced. The presence of EV positively modulated the redox unbalance due to dexamethasone. Conclusion—these findings demonstrated that EV from hAFSC have the ability to recover precursor cell potential and delay local bone loss in steroid-related osteoporosis.

scholarly journals Autophagy is impaired in fetal hypoplastic lungs and rescued by administration of amniotic fluid stem cell extracellular vesicles

2022 ◽  
Author(s):  
Kasra Khalaj ◽  
Lina Antounians ◽  
Rebeca Lopes Figueira ◽  
Martin Post ◽  
Augusto Zani
Keyword(s):  
Stem Cell ◽  
Amniotic Fluid ◽  
Extracellular Vesicles ◽  
Pulmonary Hypoplasia ◽  
Fetal Lung ◽  
Branching Morphogenesis ◽  
Lung Epithelial Cells ◽  
Normal Lung ◽  
Amniotic Fluid Stem Cell ◽  
Human Fetal Lung

Rationale: Pulmonary hypoplasia secondary to congenital diaphragmatic hernia (CDH) is characterized by reduced branching morphogenesis, which is responsible for poor clinical outcomes. Administration of amniotic fluid stem cell extracellular vesicles (AFSC-EVs) rescues branching morphogenesis in rodent fetal models of pulmonary hypoplasia. Herein, we hypothesized that AFSC-EVs exert their regenerative potential by affecting autophagy, a process required for normal lung development. Objectives: To evaluate autophagy in hypoplastic lungs throughout gestation and establish whether AFSC-EV administration improves branching morphogenesis through autophagy-mediated mechanisms. Methods: EVs were isolated from c-kit+ AFSC conditioned medium by ultracentrifugation and characterized for size, morphology, and EV markers. Branching morphogenesis was inhibited in rat fetuses by nitrofen administration to dams and in human fetal lung explants by blocking RAC1 activity with NSC23766. Expression of autophagy activators (BECN1 and ATG5) and adaptor (SQSTM1/p62) was analyzed in vitro (rat and human fetal lung explants) and in vivo (rat fetal lungs). Mechanistic studies on rat fetal primary lung epithelial cells were conducted using inhibitors for microRNA-17 and -20a contained in the AFSC-EV cargo and known to regulate autophagy. Measurements and Main Results: Rat and human models of fetal pulmonary hypoplasia showed reduced autophagy mainly at pseudoglandular and canalicular stages. AFSC-EV administration restored autophagy in both pulmonary hypoplasia models by transferring miR-17~92 cluster members contained in the EV cargo. Conclusions: AFSC-EV treatment rescues branching morphogenesis partly by restoring autophagy through miRNA cargo transfer. This study enhances our understanding of pulmonary hypoplasia pathogenesis and creates new opportunities for fetal therapeutic intervention in CDH babies.

scholarly journals Extracellular Vesicles Derived From Canine Mesenchymal Stromal Cells in Serum Free Culture Medium Have Anti-inflammatory Effect on Microglial Cells

2021 ◽  
Vol 8 ◽  
Author(s):  
Yukina Kuwahara ◽  
Karin Yoshizaki ◽  
Hidetaka Nishida ◽  
Hiroaki Kamishina ◽  
Sadatoshi Maeda ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Stromal Cells ◽  
Culture Medium ◽  
Serum Free ◽  
Naturally Occurring ◽  
Serum Free Condition ◽  
Cell Sources ◽  
Fetal Bovine ◽  
Inflammatory Effect

Mesenchymal stem/stromal cells (MSCs) have been used as cell sources for treating dogs with naturally-occurring diseases. Extracellular vesicles (EVs) derived from MSCs are now recognized as pivotal to modulating the immune response and supporting tissue repair. Manufacture of MSC-EVs for clinical application mandates removal of the xeno-proteins, including fetal bovine serum. The objective of this study was to examine whether canine MSCs survived and secreted EVs in serum-free medium (SFM) conditions and to assess the immunomodulatory effect of EVs in vitro. Canine MSCs were found to survive and secrete EVs under SFM conditions. The surface markers of MSCs in the SFM were similar to MSCs in complete culture medium. Canine MSC-EVs had a diameter of ~300 nm and were positive for EV markers. MSC-derived EVs from the serum-free condition reduced the levels of IL-1β by BV-2 cells in response to LPS stimulation. These results warrant further studies of the use of SFM for producing EVs derived from canine MSCs.

scholarly journals The Regenerative Potential of Amniotic Fluid Stem Cell Extracellular Vesicles: Lessons Learned by Comparing Different Isolation Techniques

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Lina Antounians ◽  
Areti Tzanetakis ◽  
Ornella Pellerito ◽  
Vincenzo D. Catania ◽  
Adrienne Sulistyo ◽  
...  
Keyword(s):  
Stem Cell ◽  
Amniotic Fluid ◽  
Extracellular Vesicles ◽  
Lessons Learned ◽  
Isolation Techniques ◽  
Amniotic Fluid Stem Cell

scholarly journals Extracellular Vesicles from Epicardial Fat Facilitate Atrial Fibrillation

Circulation ◽  
2021 ◽  
Author(s):  
Olga Shaihov-Teper ◽  
Eilon Ram ◽  
Nimer Ballan ◽  
Rafael Y. Brzezinski ◽  
Nili Naftali-Shani ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Extracellular Vesicles ◽  
Culture Medium ◽  
Heart Surgery ◽  
Biological Effects ◽  
Induced Pluripotent Stem Cell ◽  
Epicardial Fat ◽  
Human Mscs ◽  
And Migration

Background: The role of epicardial fat (eFat)-derived extracellular vesicles (EVs) in the pathogenesis of atrial fibrillation (AF) has never been studied. We tested the hypothesis that eFat-EVs transmit proinflammatory, profibrotic, and proarrhythmic molecules that induce atrial myopathy and fibrillation. Methods: We collected eFat specimens from patients with (n=32) and without AF (n=30) during elective heart surgery. eFat samples were grown as organ cultures, and the culture medium was collected every two days. We then isolated and purified eFat-EVs from the culture medium, and analyzed the EV number, size, morphology, specific markers, encapsulated cytokines, proteome, and miRNAs. Next, we evaluated the biological effects of unpurified and purified EVs on atrial mesenchymal stromal cells (MSCs) and endothelial cells (ECs) in vitro. To establish a causal association between eFat-EVs and vulnerability to AF, we modeled AF in vitro using induced pluripotent stem cell-derived cardiomyocytes (iCMs). Results: Microscopic examination revealed excessive inflammation, fibrosis, and apoptosis in fresh and cultured eFat tissues. Cultured explants from patients with AF secreted more EVs and harbored greater amounts of proinflammatory and profibrotic cytokines, as well as profibrotic miRNA, than those without AF. The proteomic analysis confirmed the distinctive profile of purified eFat-EVs from patients with AF. In vitro, purified and unpurified eFat-EVs from patients with AF had a greater effect on proliferation and migration of human MSCs and ECs, compared to eFat-EVs from patients without AF. Finally, while eFat-EVs from patients with and without AF shortened the action potential duration of iCMs, only eFat-EVs from patients with AF induced sustained reentry (rotor) in iCMs. Conclusions: We show, for the first time, a distinctive proinflammatory, profibrotic, and proarrhythmic signature of eFat-EVs from patients with AF. Our findings uncover another pathway by which eFat promotes the development of atrial myopathy and fibrillation.

scholarly journals Environmental conditions modulate the protein content and immunomodulatory activity of extracellular vesicles produced by the probiotic Propionibacterium freudenreichii

2020 ◽  
Author(s):  
Vinícius de Rezende Rodovalho ◽  
Brenda Silva Rosa da Luz ◽  
Aurélie Nicolas ◽  
Fillipe Luiz Rosa do Carmo ◽  
Julien Jardin ◽  
...  
Keyword(s):  
Protein Content ◽  
Extracellular Vesicles ◽  
Environmental Conditions ◽  
Culture Medium ◽  
Surface Proteins ◽  
Bioactive Molecules ◽  
Propionibacterium Freudenreichii ◽  
Therapeutic Applications ◽  
Immunomodulatory Properties

Propionibacterium freudenreichii is a probiotic Gram-positive bacterium with promising immunomodulatory properties. It modulates regulatory cytokines, mitigates the inflammatory response in vitro and in vivo. These properties were initially attributed to specific bacterial surface proteins. Recently, we showed that extracellular vesicles (EVs) produced by P. freudenreichii CIRM-BIA129 mimic the immunomodulatory features of parent cells in vitro (i.e. modulating NF-κB transcription factor activity and IL-8 release) which underlies the role of EVs as mediators of the probiotic effects of the bacterium. The modulation of EV properties, and particularly of those with potential therapeutic applications such as the EVs produced by the probiotic P. freudenreichii, is one of the challenges in the field to achieve efficient yields with the desired optimal functionality. Here we evaluated whether the culture medium in which the bacteria are grown could be used as a lever to modulate the protein content and hence the properties of P. freudenreichii CIRM-BIA129 EVs. The physical, biochemical and functional properties of EVs produced from cells cultivated on laboratory Yeast Extract Lactate (YEL) medium and cow milk ultrafiltrate (UF) medium were compared. UF-derived EVs were more abundant, smaller in diameter and displayed more intense anti-inflammatory activity than YEL-derived EVs. Furthermore, the growth media modulated EV content in terms of both the identities and abundances of their protein cargos, suggesting different patterns of interaction with the host. Proteins involved in amino acid metabolism and central carbon metabolism were modulated, as were the key surface proteins mediating host-propionibacteria interactions. Importance Extracellular vesicles (EVs) are cellular membrane-derived nanosized particles that are produced by most cells in all three kingdoms of life. They play a pivotal role in cell-cell communication through their ability to transport bioactive molecules from donor to recipient cells. Bacterial EVs are important factors in host-microbe interactions. Recently we have shown that EVs produced by the probiotic P. freudenreichii exhibited immunomodulatory properties. We evaluate here the impact of environmental conditions, notably culture media, on P. freudenreichii EV production and function. We show that EVs display considerable differences in protein cargo and immunomodulation depending on the culture medium used. This work offers new perspectives for the development of probiotic EV-based molecular delivery systems, and reinforces the optimization of growth conditions as a tool to modulate the potential therapeutic applications of EVs.

scholarly journals Oxidative Stress in Alzheimer’s Disease: In Vitro Therapeutic Effect of Amniotic Fluid Stem Cells Extracellular Vesicles

2020 ◽  
Vol 2020 ◽  
pp. 1-13
Author(s):  
Martina Gatti ◽  
Manuela Zavatti ◽  
Francesca Beretti ◽  
Daniela Giuliani ◽  
Eleonora Vandini ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Stem Cells ◽  
Amniotic Fluid ◽  
Extracellular Vesicles ◽  
Therapeutic Potential ◽  
Amniotic Fluid Stem Cells ◽  
Vitro Model

Alzheimer’s disease (AD) is characterized by abnormal protein aggregation, deposition of extracellular β-amyloid proteins (Aβ), besides an increase of oxidative stress. Amniotic fluid stem cells (AFSCs) should have a therapeutic potential for neurodegenerative disorders, mainly through a paracrine effect mediated by extracellular vesicles (EV). Here, we examined the effect of EV derived from human AFSCs (AFSC-EV) on the disease phenotypes in an AD neuron primary culture. We observed a positive effect of AFSC-EV on neuron morphology, viability, and Aβ and phospho-Tau levels. This could be due to the apoptotic and autophagic pathway modulation derived from the decrease in oxidative stress. Indeed, reactive oxygen species (ROS) were reduced, while GSH levels were enhanced. This modulation could be ascribed to the presence of ROS regulating enzymes, such as SOD1 present into the AFSC-EV themselves. This study describes the ROS-modulating effects of extracellular vesicles alone, apart from their deriving stem cell, in an AD in vitro model, proposing AFSC-EV as a therapeutic tool to stop the progression of AD.

scholarly journals Supporting data on in vitro cardioprotective and proliferative paracrine effects by the human amniotic fluid stem cell secretome

Data in Brief ◽  
2019 ◽  
Vol 25 ◽  
pp. 104324 ◽  
Author(s):  
Carolina Balbi ◽  
Kirsten Lodder ◽  
Ambra Costa ◽  
Silvia Moimas ◽  
Francesco Moccia ◽  
...  
Keyword(s):  
Stem Cell ◽  
Amniotic Fluid ◽  
Human Amniotic Fluid ◽  
Paracrine Effects ◽  
Amniotic Fluid Stem Cell

scholarly journals A Chemically Defined, Xeno- and Blood-Free Culture Medium Sustains Increased Production of Small Extracellular Vesicles From Mesenchymal Stem Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Aliosha I. Figueroa-Valdés ◽  
Catalina de la Fuente ◽  
Yessia Hidalgo ◽  
Ana María Vega-Letter ◽  
Rafael Tapia-Limonchi ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Large Scale ◽  
Culture Medium ◽  
Culture Media ◽  
Cell Phenotype ◽  
Target Cells ◽  
Scale Production ◽  
Paracrine Factors

Cell therapy is witnessing a notable shift toward cell-free treatments based on paracrine factors, in particular, towards small extracellular vesicles (sEV), that mimic the functional effect of the parental cells. While numerous sEV-based applications are currently in advanced preclinical stages, their promised translation depends on overcoming the manufacturing hurdles posed by the large-scale production of purified sEV. Unquestionably, the culture medium used with the parental cells plays a key role in the sEV’s secretion rate and content. An essential requisite is the use of a serum-, xeno-, and blood-free medium to meet the regulatory entity requirements of clinical-grade sEV’s production. Here, we evaluated OxiumTMEXO, a regulatory complying medium, with respect to production capacity and conservation of the EV’s characteristics and functionality and the parental cell’s phenotype and viability. A comparative study was established with standard DMEM and a commercially available culture medium developed specifically for sEV production. Under similar conditions, OxiumTMEXO displayed a three-fold increase of sEV secretion, with an enrichment of particles ranging between 51 and 200 nm. These results were obtained through direct quantification from the conditioned medium to avoid the isolation method’s interference and variability and were compared to the two culture media under evaluation. The higher yield obtained was consistent with several harvest time points (2, 4, and 6 days) and different cell sources, incluiding umbilical cord-, menstrual blood-derived mesenchymal stromal cells and fibroblasts. Additionally, the stem cell phenotype and viability of the parental cell remained unchanged. Furthermore, OxiumTMEXO-sEV showed a similar expression pattern of the vesicular markers CD63, CD9, and CD81, with respect to sEV derived from the other conditions. The in vitro internalization assays in different target cell types and the pharmacokinetic profile of intraperitoneally administered sEV in vivo indicated that the higher EV production rate did not affect the uptake kinetics or the systemic biodistribution in healthy mice. In conclusion, the OxiumTMEXO medium sustains an efficient and robust production of large quantities of sEV, conserving the classic functional properties of internalization into acceptor target cells and biodistribution in vivo, supplying the amount and quality of EVs for the development of cell-free therapies.

Sign in / Sign up

Export Citation Format

Share Document