Enhancing the efficacy and therapeutic potential of human mesenchymal stromal cells (hMSCs) in clinically relevant sepsis models
González Esteban, Héctor
González Esteban, Héctor
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Publication Date
2022-08-30
Type
Thesis
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Abstract
Rationale: Sepsis is a systemic inflammatory process produced by the reaction of the immune system against a pathogen. This inflammatory process produces tissue damage leading into vital organ malfunction and, if not treated on time, the death of the patient. Despite the severity of the syndrome no specific treatment has been discovered yet. Mesenchymal stromal cells (MSCs) are promising contenders for sepsis therapy, but issues on the heterogeneity of cell product based on different tissue sources, donors and isolation methods need to be solved. In addition, new therapeutic formulas and delivery routes are desirable to increase therapeutic effect and reduce potential side effects of MSC delivery to patients. Methods: (1) Systemic cells lines were exposed to inflammatory damage and treated with the CD362+ hUC-MSC secretome. NF-κB pathway activation, cytokine production, cell viability and indirect antibacterial capacity was analysed. (2) Sepsis was induced in animals by CLP surgery and 4x106 CD362+ hUC-MSCs per kilogram were delivered intravenously 4h later. 48 hours after the injury induction, physiological parameters, injury markers and bacterial presence was analysed. (3) CLP animals were treated with 4x106 CD362+ hUC-MSCs per kilogram simultaneous to and 24 hours after CLP. 48 hours after the injury induction physiological parameters, injury markers and bacterial presence were analysed. (4) Lung cell lines were exposed to inflammatory damage and treated with concentrated conditioned media and EVs isolated from naïve hBM-MSCs and CD362+ hUC-MSCs. NF-κB pathway activation, cytokine production, cell viability and antibacterial capacity was analysed. (5, 7). Lung injury was induced by installation of a live bacterial culture of E. coli into the lungs. After one hour, concentrated conditioned media from hBM-MSCs was intratracheally installed or nebulised using an vibrating mesh nebuliser. 48 hours after injury induction, physiological parameters, injury markers and bacterial presence were analysed. Using the same lung injury model, therapeutic capacity of the EVs, comparative between the cell sources and delivery routes, were performed by nebulisation or intravenous administration of EVs from hBM-MSCs and CD362+ hUC-MSCs 1 hour after injury induction. 48 hours after injury induction, physiological parameters, injury markers and bacterial presence were analysed. (6) Lung cells lines were exposed to inflammatory damage and treated with nebulised concentrated conditioned media and nebulised EVs isolated from naïve hBM-MSCs and CD362+ hUC-MSCs. NF-κB pathway activation, cytokine production, cell viability and antibacterial capacity were analysed. Results: (1) CD362+ hUC-MSC secretome reduces inflammatory damage in systemic cells, reducing NF-κB activation and inflammatory cytokine production, meanwhile increasing antibacterial and healing activity. (2) Treatment with CD362+ hUC-MSCs reduces systemic inflammatory markers and bacterial presence ameliorating sepsis damage. (3) No therapeutic effect was observed when CD362+ hUC-MSCs were administrated 0 and 24 hours after sepsis induction (4) hBM-MSC and CD362+ hUC-MSC concentrated conditioned media and EVs mitigate inflammatory damage in lung cells by reducing NF-κB activation and inflammatory cytokine production meanwhile increasing antibacterial and healing activity. (5) hBM-MSC and CD362+ hUC-MSC concentrated conditioned media and EVs ameliorate the lung injury in vivo, reducing lung cell infiltration, inflammatory cytokine levels and bacterial presence meanwhile improve the tissue integrity and oxygenation exchange. (6) Nebulisation of concentrated conditioned media and EVs from hBM-MSC and CD362+ hUC-MSCs retain the therapeutic capacities to mitigate inflammatory damage on lungs cells. (7) Direct nebulisation of hBM-MSC and CD362+ hUC-MSC concentrated conditioned media and EVs did not affect the therapeutic capacity of the cell products to mitigate lung injury in vivo. Conclusion: (1) CD362+ hUC-MSC secretome reduces inflammatory damage in systemic cells by inhibition of the NF-κB pathway, reduction of inflammatory cytokine production, increase in antibacterial capacity and increase in the healing process. (2) CD362+ hUC-MSC mitigate CLP induced sepsis, reducing bad prognosis and injury markers levels and promoting systemic bacterial clearance. (3) The administration time of CD362+ hUC-MSC determinates the therapeutic capacity of the cells to mitigate the septic process. (4) MSC products from hBM MSCs and CD362+ hUC-MSCs reduce inflammatory damage through inhibiting the NF-κB pathway, reducing inflammatory cytokine production and increasing antibacterial and tissue healing capacity in lung cells. (5) MSC products from hBM-MSCs and CD362+ hUC-MSCs reduce cell infiltration, oedema, inflammatory cytokine levels and bacterial presence in pneumonia lungs, improving lung function and alveolar gas exchange. (6) Nebulised cell products retain the therapeutic capacity to mitigate the inflammatory damage on lung cells. (7) Direct nebulisation of MSC products does not affect their therapeutic capacity to mitigate lung injury in vivo and offers a potential new route for the MSCs products in pneumosepsis.
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NUI Galway