Impact of starting material (fresh versus cryopreserved marrow) on mesenchymal stem cell culture

Kaplan, Alesia; Sackett, Katie; Sumstad, Darin; Kadidlo, Diane; McKenna, David H.

doi:10.1111/trf.14192

Cited by 19 publications

(14 citation statements)

References 17 publications

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“…As with the study by Kaplan et al, cryopreservation did not significantly affect cell proliferation, but trends demonstrated higher rates in FC MSCs on day 10 ( p = 0.07, Fig. 2b) [33]. Similarly, the clonogenic capacity was also higher in fresh MSCs as compared to FT MSCs ( p < 0.05, Fig.…”

Section: Discussionsupporting

confidence: 76%

Cryopreserved mesenchymal stem cells regain functional potency following a 24-h acclimation period

et al. 2019

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Background Mesenchymal stem cells (MSCs) are attractive cell-therapy candidates. Despite their popularity and promise, there is no uniform method of preparation of MSCs. Typically, cells are cryopreserved in liquid nitrogen, thawed, and subsequently administered to a patient with little to no information on their function post-thaw. We hypothesized that a short acclimation period post-thaw will facilitate the recovery of MSC’s functional potency. Methods Human bone-marrow-derived MSCs were divided into 3 groups: FC (fresh cells; from existing culture); TT (thawed + time; acclimated for 24 h post-thaw); and FT (freshly thawed; thawed and immediately used). The 3 groups were analyzed for their cellular and functional potency. Results Phenotypic analysis demonstrated a decrease in CD44 and CD105 surface markers in FT MSCs, with no change in the other two groups. All MSCs were able to differentiate down the osteogenic and chondrogenic lineages. In FT cells, metabolic activity and apoptosis was significantly increased with concomitant decrease in cell proliferation; clonogenic capacity; and key regenerative genes. Following 24-h acclimation, apoptosis was significantly reduced in TT cells with a concomitant upregulation in angiogenic and anti-inflammatory genes. While all MSCs significantly arrested T-cell proliferation, the TT MSCs were significantly more potent. Similarly, although all MSCs maintained their anti-inflammatory properties, IFN-γ secretion was significantly diminished in FT cells. Conclusions These data demonstrate that FT MSCs maintain their multipotent differentiation capacity, immunomodulatory function, and anti-inflammatory properties; yet, various aspects of cell characteristics and function are deleteriously affected by cryopreservation. Importantly, a 24-h acclimation period ‘reactivates’ thawed cells to recover their diminished stem-cell function.

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Section: Discussionsupporting

confidence: 76%

Cryopreserved mesenchymal stem cells regain functional potency following a 24-h acclimation period

et al. 2019

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“…Previous studies have documented success in recovering viable nucleated cells from cryopreserved bone marrow [15,16,23,24]. However, the primary focus of these earlier studies was on the culture expansion of progenitor cells for clinical use and not the quantification of the initial progenitor cell content.…”

Section: Discussionmentioning

confidence: 99%

“…Strong correlations have been observed between pre-freeze and postthaw bone marrow across several cell populations, including mononuclear cells, committed myeloid precursors and hematopoietic stem cells [14]. Additionally, connective tissue progenitor cells have been recovered from cryopreserved bone marrow and successfully culture expanded with clinical applications in mind [15,16]. Although several groups have performed the CFU-F assay with both fresh and cryopreserved bone marrow and reported no differences between the two, data are limited [15] or not shown [17].…”

Section: Introductionmentioning

confidence: 99%

Cryopreserved bone marrow aspirate concentrate as a cell source for the colony-forming unit fibroblast assay

Berger¹,

Aune²,

Centeno

et al. 2020

Cytotherapy

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The prevalence of connective tissue progenitor cells within a cell-based therapy is often quantified using the colony-forming unit fibroblast (CFU-F) assay. The present study investigates the feasibility of using cryopreserved bone marrow aspirate concentrate (BMAC) as an alternative cell source to fresh BMAC for CFU-F quantification. Methods: Freshly prepared and corresponding cryopreserved BMAC samples from patients receiving autologous cell therapy (n = 98) were analyzed using the CFU-F assay for comparison. Cultures were established by directly plating BMAC at low cell densities and maintained for a 2-week growth period. Colonies were enumerated to determine CFU-F frequency, and a subset of cultures was imaged and analyzed to quantify colony area and density. Results: A nonlinear relationship was observed between plating density and CFU-F frequency over a wide range in plating densities (~30-fold). Cryopreserved BMAC yielded recoverable (77 § 23%) and viable (73 § 9%) nucleated cells upon thawing. After cryopreservation, CFU-F frequencies were found to be significantly lower (56.6 § 34.8 vs. 50.3 § 31.7 colonies per million nucleated cells). Yet the number of CFU-F in fresh and cryopreserved BMAC were strongly correlated (r = 0.87) and had similar area and densities. Further, moderate correlations were observed between the number of CFU-F and nucleated cells, and both the mean colony area and density were negatively correlated with patient age. Notably, no relationship was found between CFU-F frequency and age, regardless of whether fresh or cryopreserved BMAC was used. Conclusions: Freshly prepared and cryopreserved BMAC yielded correlated results when analyzed using the CFU-F assay. Our findings support the cryogenic storage of patient BMAC samples for retrospective CFU-F analyses, offering a potential alternative for characterizing BMAC and furthering our understanding of progenitor cells in relation to clinical outcome.

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“…These results suggest that the administration of MSCs with a high viability is required to target an improvement in efficacy. Recent experimental data on the comparison of different cell products reports that fresh BM-MSCs are 14% more viable compared to cryopreserved ones [172]. Furthermore, delivery of MSC immediately upon thawing instead of thawing and washing the MSCs could enhance MSC viability, as observed by Matthay MA et al [71].…”

Section: Clinical Trials-demonstrating Safety In Ardsmentioning

confidence: 91%

The Safety and Efficiency of Addressing ARDS Using Stem Cell Therapies in Clinical Trials

Rezoagli

Murphy

Laffey

et al. 2019

Stem Cell-Based Therapy for Lung Disease

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Acute Respiratory Distress Syndrome (ARDS) is a complex and debilitating disease of the lungs, which continues to have a high mortality rate and huge disease burden on patients. Incidence is rising, possibly due to greater awareness leading to more diagnoses rather than a change in the underlying rate. It arises from multiple etiologies, though pathogenic infection, termed pneumonia, is the most prevalent and widely studied. The distinct pathophysiology and rapid evolution of ARDS makes it uniquely challenging with regard to therapeutics development and, to date, no medicines are licensed for specific therapy. Antibiotics, ventilation, and other organ support remain intervention standards.1. Rapid onset of symptoms that cannot be attributed to any underlying cause. 2. Bilateral infiltration of leukocytes from surrounding tissue to the airspace, as identified by chest X-ray.

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Impact of starting material (fresh versus cryopreserved marrow) on mesenchymal stem cell culture

Abstract: This study demonstrates that MSCs derived from cryopreserved marrow have the same growth characteristics as fresh marrow-derived MSCs. Further studies are needed to explore potential differences in clinical efficacy.

Cited by 19 publications

References 17 publications

Cryopreserved mesenchymal stem cells regain functional potency following a 24-h acclimation period

Cryopreserved mesenchymal stem cells regain functional potency following a 24-h acclimation period

Cryopreserved bone marrow aspirate concentrate as a cell source for the colony-forming unit fibroblast assay

The Safety and Efficiency of Addressing ARDS Using Stem Cell Therapies in Clinical Trials

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