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Umbilical cord-derived CD362+ mesenchymal stromal cells for E. coli pneumonia: impact of dose regimen, passage, cryopreservation, and antibiotic therapy.

ABSTRACT: BACKGROUND:Mesenchymal stromal cells (MSCs) demonstrate considerable promise for acute respiratory distress syndrome (ARDS) and sepsis. However, standard approaches to MSC isolation generate highly heterogeneous cell populations, while bone marrow (BM) constitutes a limited and difficult to access MSC source. Furthermore, a range of cell manufacturing considerations and clinical setting practicalities remain to be explored. METHODS:Adult male rats were subject to E. coli-induced pneumonia and administered CD362+ umbilical cord (UC)-hMSCs using a variety of cell production and clinical relevance considerations. In series 1, animals were instilled with E. coli and randomized to receive heterogeneous BM or UC-hMSCs or CD362+ UC-hMSCs. Subsequent series examined the impact of concomitant antibiotic therapy, MSC therapeutic cryopreservation (cryopreserved vs fresh CD362+ UC-hMSCs), impact of cell passage on efficacy (passages 3 vs 5 vs 7 vs 10), and delay of administration of cell therapy (0?h vs 6?h post-injury vs 6?h?+?12?h) following E. coli installation. RESULTS:CD362+ UC-hMSCs were as effective as heterogonous MSCs in reducing E. coli-induced acute lung injury, improving oxygenation, decreasing bacterial load, reducing histologic injury, and ameliorating inflammatory marker levels. Cryopreserved CD362+ UC-hMSCs recapitulated this efficacy, attenuating E. coli-induced injury, but therapeutic relevance did not extend beyond passage 3 for all indices. CD362+ UC-hMSCs maintained efficacy in the presence of antibiotic therapy and rescued the animal from E. coli injury when delivered at 6?h?+?12?h, following E. coli instillation. CONCLUSIONS:These translational studies demonstrated the efficacy of CD362+ UC-hMSCs, where they decreased the severity of E. coli-induced pneumonia, maintained efficacy following cryopreservation, were more effective at early passage, were effective in the presence of antibiotic therapy, and could continue to provide benefit at later time points following E. coli injury.

SUBMITTER: Horie S

PROVIDER: S-EPMC7071745 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Umbilical cord-derived CD362<sup>+</sup> mesenchymal stromal cells for E. coli pneumonia: impact of dose regimen, passage, cryopreservation, and antibiotic therapy.

Horie Shahd S Masterson Claire C Brady Jack J Loftus Paul P Horan Emma E O'Flynn Lisa L Elliman Steve S Barry Frank F O'Brien Timothy T Laffey John G JG O'Toole Daniel D

Stem cell research & therapy 20200313 1

<h4>Background</h4>Mesenchymal stromal cells (MSCs) demonstrate considerable promise for acute respiratory distress syndrome (ARDS) and sepsis. However, standard approaches to MSC isolation generate highly heterogeneous cell populations, while bone marrow (BM) constitutes a limited and difficult to access MSC source. Furthermore, a range of cell manufacturing considerations and clinical setting practicalities remain to be explored.<h4>Methods</h4>Adult male rats were subject to E. coli-induced p ...[more]

PMID: 32169108

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Project description:BACKGROUND:Mesenchymal stromal cells (MSCs) are used in over 800 clinical trials mainly due to their immune inhibitory activity. Umbilical cord (UC), the second leading source of clinically used MSCs, is usually cut in small tissue pieces. Subsequent cultivation leads to a continuous outgrowth of MSC explant monolayers (MSC-EMs) for months. Currently, the first MSC-EM culture takes approximately 2 weeks to grow out, which is then expanded and applied to patients. The initiating tissue pieces are then discarded. However, when UC pieces are transferred to new culture dishes, MSC-EMs continue to grow out. In case the functional integrity of these cells is maintained, later induced cultures could also be expanded and used for cell therapy. This would drastically increase the number of available cells for each patient. To test the functionality of MSC-EMs from early and late induction time points, we compared the first cultures to those initiated after 2 months by investigating their clonality and immunomodulatory capacity. METHODS:We analyzed the clonal composition of MSC-EM cultures by umbilical cord piece transduction using integrating lentiviral vectors harboring genetic barcodes assessed by high-throughput sequencing. We investigated the transcriptome of these cultures by microarrays. Finally, the secretome was analyzed by multiplexed ELISAs, in vitro assays, and in vivo in mice. RESULTS:DNA barcode analysis showed polyclonal MSC-EMs even after months of induction cycles. A transcriptome and secretome analyses of early and late MSC cultures showed only minor changes over time. However, upon activation with TNF-α and IFN-γ, cells from both induction time points produced a multitude of immunomodulatory cytokines. Interestingly, the later induced MSC-EMs produced higher amounts of cytokines. To test whether the different cytokine levels were in a therapeutically relevant range, we used conditioned medium (CM) in an in vitro MLR and an in vivo killing assay. CM from late induced MSC-EMs was at least as immune inhibitory as CM from early induced MSC-EMs. CONCLUSION:Human umbilical cord maintains a microenvironment for the long-term induction of polyclonal and immune inhibitory active MSCs for months. Thus, our results would offer the possibility to drastically increase the number of therapeutically applicable MSCs for a substantial amount of patients.

Dataset Information

Umbilical cord-derived CD362+ mesenchymal stromal cells for E. coli pneumonia: impact of dose regimen, passage, cryopreservation, and antibiotic therapy.

Publications

Umbilical cord-derived CD362<sup>+</sup> mesenchymal stromal cells for E. coli pneumonia: impact of dose regimen, passage, cryopreservation, and antibiotic therapy.

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