Immortalization strategies for human mesenchymal stromal cells for large scale production of extracellular vesicles

Cytotherapy ◽  
2021 ◽  
Vol 23 (5) ◽  
pp. S119
Author(s):  
Y. Mouloud ◽  
F. Nardi Bauer ◽  
H. Hanenberg ◽  
B. Giebel ◽  
P. Horn
Keyword(s):  
Extracellular Vesicles ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Large Scale ◽  
Scale Production ◽  
Human Mesenchymal Stromal Cells ◽  
Large Scale Production

scholarly journals Automated Large-Scale Production of Paclitaxel Loaded Mesenchymal Stromal Cells for Cell Therapy Applications

Pharmaceutics ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 411 ◽  
Author(s):  
Daniela Lisini ◽  
Sara Nava ◽  
Simona Frigerio ◽  
Simona Pogliani ◽  
Guido Maronati ◽  
...  
Keyword(s):  
Cell Therapy ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Large Scale ◽  
Good Manufacturing Practice ◽  
Scale Production ◽  
Large Scale Production ◽  
Advanced Therapies ◽  
Oncological Diseases

Mesenchymal stromal cells (MSCs) prepared as advanced therapies medicinal products (ATMPs) have been widely used for the treatment of different diseases. The latest developments concern the possibility to use MSCs as carrier of molecules, including chemotherapeutic drugs. Taking advantage of their intrinsic homing feature, MSCs may improve drugs localization in the disease area. However, for cell therapy applications, a significant number of MSCs loaded with the drug is required. We here investigate the possibility to produce a large amount of Good Manufacturing Practice (GMP)-compliant MSCs loaded with the chemotherapeutic drug Paclitaxel (MSCs-PTX), using a closed bioreactor system. Cells were obtained starting from 13 adipose tissue lipoaspirates. All samples were characterized in terms of number/viability, morphology, growth kinetics, and immunophenotype. The ability of MSCs to internalize PTX as well as the antiproliferative activity of the MSCs-PTX in vitro was also assessed. The results demonstrate that our approach allows a large scale expansion of cells within a week; the MSCs-PTX, despite a different morphology from MSCs, displayed the typical features of MSCs in terms of viability, adhesion capacity, and phenotype. In addition, MSCs showed the ability to internalize PTX and finally to kill cancer cells, inhibiting the proliferation of tumor lines in vitro. In summary our results demonstrate for the first time that it is possible to obtain, in a short time, large amounts of MSCs loaded with PTX to be used in clinical trials for the treatment of patients with oncological diseases.

scholarly journals Reproducible Large-Scale Isolation of Exosomes from Adipose Tissue-Derived Mesenchymal Stem/Stromal Cells and Their Application in Acute Kidney Injury

2020 ◽  
Vol 21 (13) ◽  
pp. 4774 ◽  
Author(s):  
Jun Ho Lee ◽  
Dae Hyun Ha ◽  
Hyeon-kyu Go ◽  
Jinkwon Youn ◽  
Hyun-keun Kim ◽  
...  
Keyword(s):  
Acute Kidney Injury ◽  
Adipose Tissue ◽  
Stromal Cells ◽  
Large Scale ◽  
Kidney Diseases ◽  
Kidney Injury ◽  
Preclinical Studies ◽  
Scale Production ◽  
Positive Effects ◽  
Large Scale Production

Acute kidney injury (AKI) is a fatal medical episode caused by sudden kidney damage or failure, leading to the death of patients within a few hours or days. Previous studies demonstrated that exosomes derived from various mesenchymal stem/stromal cells (MSC-exosomes) have positive effects on renal injuries in multiple experimental animal models of kidney diseases including AKI. However, the mass production of exosomes is a challenge not only in preclinical studies with large animals but also for successful clinical applications. In this respect, tangential flow filtration (TFF) is suitable for good manufacturing practice (GMP)-compliant large-scale production of high-quality exosomes. Until now, no studies have been reported on the use of TFF, but rather ultracentrifugation has been almost exclusively used, to isolate exosomes for AKI therapeutic application in preclinical studies. Here, we demonstrated the reproducible large-scale production of exosomes derived from adipose tissue-derived MSC (ASC-exosomes) using TFF and the lifesaving effect of the ASC-exosomes in a lethal model of cisplatin-induced rat AKI. Our results suggest the possibility of large-scale stable production of ASC-exosomes without loss of function and their successful application in life-threatening diseases.

scholarly journals Membrane Particles Derived From Adipose Tissue Mesenchymal Stromal Cells Improve Endothelial Cell Barrier Integrity

2021 ◽  
Vol 12 ◽  
Author(s):  
Ana Merino ◽  
Marta Sablik ◽  
Sander S. Korevaar ◽  
Carmen López-Iglesias ◽  
Maitane Ortiz-Virumbrales ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Large Scale ◽  
Umbilical Vein ◽  
Tube Length ◽  
Scale Production ◽  
Membrane Particles ◽  
Branching Points

Proinflammatory stimuli lead to endothelial injury, which results in pathologies such as cardiovascular diseases, autoimmune diseases, and contributes to alloimmune responses after organ transplantation. Both mesenchymal stromal cells (MSC) and the extracellular vesicles (EV) released by them are widely studied as regenerative therapy for the endothelium. However, for therapeutic application, the manipulation of living MSC and large-scale production of EV are major challenges. Membrane particles (MP) generated from MSC may be an alternative to the use of whole MSC or EV. MP are nanovesicles artificially generated from the membranes of MSC and possess some of the therapeutic properties of MSC. In the present study we investigated whether MP conserve the beneficial MSC effects on endothelial cell repair processes under inflammatory conditions. MP were generated by hypotonic shock and extrusion of MSC membranes. The average size of MP was 120 nm, and they showed a spherical shape. The effects of two ratios of MP (50,000; 100,000 MP per target cell) on human umbilical vein endothelial cells (HUVEC) were tested in a model of inflammation induced by TNFα. Confocal microscopy and flow cytometry showed that within 24 hours >90% of HUVEC had taken up MP. Moreover, MP ended up in the lysosomes of the HUVEC. In a co-culture system of monocytes and TNFα activated HUVEC, MP did not affect monocyte adherence to HUVEC, but reduced the transmigration of monocytes across the endothelial layer from 138 ± 61 monocytes per microscopic field in TNFα activated HUVEC to 61 ± 45 monocytes. TNFα stimulation induced a 2-fold increase in the permeability of the HUVEC monolayer measured by the translocation of FITC-dextran to the lower compartment of a transwell system. At a dose of 1:100,000 MP significantly decreased endothelial permeability (1.5-fold) respect to TNFα Stimulated HUVEC. Finally, MP enhanced the angiogenic potential of HUVEC in an in vitro Matrigel assay by stimulating the formation of angiogenic structures, such as percentage of covered area, total tube length, total branching points, total loops. In conclusion, MP show regenerative effects on endothelial cells, opening a new avenue for treatment of vascular diseases where inflammatory processes damage the endothelium.

scholarly journals Inhibition of Human Malignant Pleural Mesothelioma Growth by Mesenchymal Stromal Cells

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1427
Author(s):  
Valentina Coccè ◽  
Silvia La Monica ◽  
Mara Bonelli ◽  
Giulio Alessandri ◽  
Roberta Alfieri ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Mesenchymal Stromal Cells ◽  
Malignant Pleural Mesothelioma ◽  
Stromal Cells ◽  
Large Scale ◽  
Significant Inhibition ◽  
G1 Arrest ◽  
Pleural Mesothelioma ◽  
Scale Production

Background: Malignant Pleural Mesothelioma (MPM) is an aggressive tumor that has a significant incidence related to asbestos exposure with no effective therapy and poor prognosis. The role of mesenchymal stromal cells (MSCs) in cancer is controversial due to their opposite effects on tumor growth and in particular, only a few data are reported on MSCs and MPM. Methods: We investigated the in vitro efficacy of adipose tissue-derived MSCs, their lysates and secretome against different MPM cell lines. After large-scale production of MSCs in a bioreactor, their efficacy was also evaluated on a human MPM xenograft in mice. Results: MSCs, their lysate and secretome inhibited MPM cell proliferation in vitro with S or G0/G1 arrest of the cell cycle, respectively. MSC lysate induced cell death by apoptosis. The efficacy of MSC was confirmed in vivo by a significant inhibition of tumor growth, similar to that produced by systemic administration of paclitaxel. Interestingly, no tumor progression was observed after the last MSC treatment, while tumors started to grow again after stopping chemotherapeutic treatment. Conclusions: These data demonstrated for the first time that MSCs, both through paracrine and cell-to-cell interaction mechanisms, induced a significant inhibition of human mesothelioma growth. Since the prognosis for MPM patients is poor and the options of care are limited to chemotherapy, MSCs could provide a potential new therapeutic approach for this malignancy.

scholarly journals Stem Cell Extracellular Vesicles in Skin Repair

2018 ◽  
Vol 6 (1) ◽  
pp. 4 ◽  
Author(s):  
Andrea da Fonseca Ferreira ◽  
Dawidson Assis Gomes
Keyword(s):  
Stem Cell ◽  
Extracellular Vesicles ◽  
Large Scale ◽  
Therapeutic Potential ◽  
Skin Aging ◽  
Skin Wound ◽  
Scale Production ◽  
Skin Wound Healing ◽  
Skin Cell ◽  
Large Scale Production

Stem cell extracellular vesicles (EVs) have been widely studied because of their excellent therapeutic potential. EVs from different types of stem cell can improve vascularization as well as aid in the treatment of cancer and neurodegenerative diseases. The skin is a complex organ that is susceptible to various types of injury. Strategies designed to restore epithelial tissues’ integrity with stem cell EVs have shown promising results. Different populations of stem cell EVs are able to control inflammation, accelerate skin cell migration and proliferation, control wound scarring, improve angiogenesis, and even ameliorate signs of skin aging. However, large-scale production of such stem cell EVs for human therapy is still a challenge. This review focuses on recent studies that explore the potential of stem cell EVs in skin wound healing and skin rejuvenation, as well as challenges of their use in therapy.

A large (scale) advance for small RNA therapeutics

2018 ◽  
Vol 10 (465) ◽  
pp. eaav3895
Author(s):  
Steven M. Jay
Keyword(s):  
Extracellular Vesicles ◽  
Small Rna ◽  
Large Scale ◽  
Scale Production ◽  
Large Scale Production

Combining established techniques enables large-scale production of potentially therapeutic extracellular vesicles enriched with specific miRNAs.

scholarly journals Extracellular Vesicles as Drug Delivery Systems in Cancer

Pharmaceutics ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 1146
Author(s):  
Laia Hernandez-Oller ◽  
Joaquin Seras-Franzoso ◽  
Fernanda Andrade ◽  
Diana Rafael ◽  
Ibane Abasolo ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Extracellular Vesicles ◽  
Drug Delivery Systems ◽  
Large Scale ◽  
Delivery Systems ◽  
Biological Behavior ◽  
Scale Production ◽  
Cancer Resistance ◽  
Large Scale Production ◽  
Transport Complex

Within tumors, Cancer Stem Cell (CSC) subpopulation has an important role in maintaining growth and dissemination while preserving high resistance against current treatments. It has been shown that, when CSCs are eliminated, the surrounding Differentiated Cancer Cells (DCCs) may reverse their phenotype and gain CSC-like features to preserve tumor progression and ensure tumor survival. This strongly suggests the existence of paracrine communication within tumor cells. It is evidenced that the molecular crosstalk is at least partly mediated by Extracellular Vesicles (EVs), which are cell-derived membranous nanoparticles that contain and transport complex molecules that can affect and modify the biological behavior of distal cells and their molecular background. This ability of directional transport of small molecules prospects EVs as natural Drug Delivery Systems (DDS). EVs present inherent homing abilities and are less immunogenic than synthetic nanoparticles, in general. Currently, strong efforts are focused into the development and improvement of EV-based DDS. Even though EV-DDS have already reached early phases in clinical trials, their clinical application is still far from commercialization since protocols for EVs loading, modification and isolation need to be standardized for large-scale production. Here, we summarized recent knowledge regarding the use of EVs as natural DDS against CSCs and cancer resistance.

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