Isolation of Adipose Tissue–Derived Stem Cells: Enzymatic Digestion in Combination with Mechanical Distortion to Increase Adipose Tissue–Derived Stem Cell Yield from Human Aspirated Fat

Alstrup, Toke; Eijken, Marco; Bohn, Anja Bille; Møller, Bjarne Kuno; Damsgaard, Tine Engberg

doi:10.1002/cpsc.68

Cited by 29 publications

(12 citation statements)

References 19 publications

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“…Some authors, showed the ability to isolate and expand adipose-derived stromal cells in stirred bioreactor using Corning R Synthemax II microcarriers (Gadelorge et al, 2017), but the majority of the studies with AT MSC are focused in tissue regeneration and its immunomodulatory properties (Zhou et al, 2011;Madonna et al, 2015). Nevertheless, reports show improvements in isolation techniques than can further optimize the process of using AT MSC in xeno-free conditions (Francis et al, 2018;Alstrup et al, 2019;Lisini et al, 2019). Although we have established a serum-free and xeno-free microcarrier-based expansion of AT MSC using a commercially available formulation Santos et al, 2011;Carmelo et al, 2015), the cell adhesion (36 ± 5.2%) and maximal cell density (1.9 × 10 5 cells/mL) achieved were significantly inferior when compared to BM MSC expanded under the same conditions (95% and 3.6 × 10 5 cells/mL), highlighting the need to improve the expansion process of this specific cell source under xenogeneic(xeno)-free culture conditions (Carmelo et al, 2015).…”

Section: Discussionmentioning

confidence: 99%

Successful Use of Human AB Serum to Support the Expansion of Adipose Tissue-Derived Mesenchymal Stem/Stromal Cell in a Microcarrier-Based Platform

Moreira

Mizukami

Souza

et al. 2020

Front. Bioeng. Biotechnol.

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Mesenchymal stem/stromal cells (MSC) are promising candidates for cell-based therapies and for the promotion of tissue repair, hence the increase of clinical trials in a worldwide scale. In particular, adipose tissue-derived stem/stromal cells (AT MSC) present easy accessibility and a rather straightforward process of isolation, providing a clear advantage over other sources. The high demand of cell doses (millions of cells/kg), needed for infusion in clinical settings, requires a scalable and efficient manufacturing of AT MSC under xenogeneic(xeno)-free culture conditions. Here we describe the successful use of human AB serum [10%(v/v)] as a culture supplement, as well as coating substrate for the expansion of these cells in microcarriers using (i) a spinner flask and (ii) a 500-mL mini-bioreactor (Applikon TM Biotechnology). Cells were characterized by immunophenotype and multilineage differentiation potential. Upon an initial cell adhesion in the spinner flask of 35 ± 2.5%, culture reached a maximal cell density of 2.6 ± 0.1 × 10 5 at day 7, obtaining a 15 ± 1-fold increase. The implementation of the culture in the 500-mL mini-bioreactor presented an initial cell adhesion of 22 ± 5%, but it reached maximal cell density of 2.7 ± 0.4 × 10 5 at day 7, obtaining a 27 ± 8-fold increase. Importantly, in both stirred systems, cells retained their immunophenotype and multilineage differentiation potential (osteo-, chondro-and adipogenic lineages). Overall, the scalability of this microcarrier-based system presented herein is of major importance for the purpose of achieving clinically meaningful cell numbers.

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Section: Discussionmentioning

confidence: 99%

Successful Use of Human AB Serum to Support the Expansion of Adipose Tissue-Derived Mesenchymal Stem/Stromal Cell in a Microcarrier-Based Platform

Moreira

Mizukami

Souza

et al. 2020

Front. Bioeng. Biotechnol.

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“…For example, harvesting large volumes of BM-MSCs is associated with increased donor-site morbidity [ 23 , 24 ]. Different buffers and culturing techniques can result in yield differences in AD-MSCs [ 25 , 26 ]. In general, AD-MSCs are metabolically active for longer durations, making them able to go through more replication cycles than BM-MSCs; however, no proposed mechanism has been given for this observation [ 13 , 27 , 28 , 29 ].…”

Section: Tissues Containing Mscsmentioning

confidence: 99%

Mesenchymal Stem Cell Mechanisms of Action and Clinical Effects in Osteoarthritis: A Narrative Review

Molnar

Pavelić²,

Vrdoljak

et al. 2022

Genes

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With the insufficient satisfaction rates and high cost of operative treatment for osteoarthritis (OA), alternatives have been sought. Furthermore, the inability of current medications to arrest disease progression has led to rapidly growing clinical research relating to mesenchymal stem cells (MSCs). The availability and function of MSCs vary according to tissue source. The three primary sources include the placenta, bone marrow, and adipose tissue, all of which offer excellent safety profiles. The primary mechanisms of action are trophic and immunomodulatory effects, which prevent the further degradation of joints. However, the function and degree to which benefits are observed vary significantly based on the exosomes secreted by MSCs. Paracrine and autocrine mechanisms prevent cell apoptosis and tissue fibrosis, initiate angiogenesis, and stimulate mitosis via growth factors. MSCs have even been shown to exhibit antimicrobial effects. Clinical results incorporating clinical scores and objective radiological imaging have been promising, but a lack of standardization in isolating MSCs prevents their incorporation in current guidelines.

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“…The minced adipose tissue was then digested on an orbital shaker at 140 rpm using 0.1% collagenase type I (Sigma Aldrich, Darmstadt, Germany) at 37 • C for 1.5 h. In total, 300 µL of 0.1% collagenase were used per gram of fat. Thereafter, the fat mixture was centrifuged at 644× g for 5 min to obtain a cell pellet [3,23].…”

Section: Fat Harvest and Culture Of Mscsmentioning

confidence: 99%

Comparison of Fat Harvested from Flank and Falciform Regions for Stem Cell Therapy in Dogs

Jifcovici¹,

Solano²,

Fitzpatrick³

et al. 2021

Veterinary Sciences

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Background: Adipose tissue has recently gained attention as a source of mesenchymal stem cells (AdMSCs) for applications in treating degenerative joint disease in veterinary patients. This study aimed to quantify the stromal vascular fractions (SVFs) and colony forming units (CFU) of AdMSCs from the falciform and flank regions and compare dogs of different ages and weights. Methods: Fat tissue was harvested from the flank (21 dogs) and falciform regions (17 dogs). The fat tissue was enzymatically digested and the number of nucleated cells in the SVF was counted. The SVF was cultured in vitro and the cell growth was assessed by counting the CFU per gram of fat and the aspect ratio of the cells. Conclusions: There was no significant difference in the number of nucleated cells in the SVF from the two sites. The CFU/g of fat from falciform was 378.9 ± 293 g and from flank was 486.8 ± 517 g, and this was also insignificant. Neither age nor weight of the patient had an impact on the SVF or CFU/g. No surgical complications were reported from either of the sites. Harvesting fat for stem cell therapy for intra-articular therapy of degenerative joint disease can be an easy and fast process when obtaining the fat either from the flank or the falciform region, and it is not age or weight dependent. The harvest site for clinical canine patients can be left to the surgeon’s discretion and comfort.

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Isolation of Adipose Tissue–Derived Stem Cells: Enzymatic Digestion in Combination with Mechanical Distortion to Increase Adipose Tissue–Derived Stem Cell Yield from Human Aspirated Fat

Cited by 29 publications

References 19 publications

Successful Use of Human AB Serum to Support the Expansion of Adipose Tissue-Derived Mesenchymal Stem/Stromal Cell in a Microcarrier-Based Platform

Successful Use of Human AB Serum to Support the Expansion of Adipose Tissue-Derived Mesenchymal Stem/Stromal Cell in a Microcarrier-Based Platform

Mesenchymal Stem Cell Mechanisms of Action and Clinical Effects in Osteoarthritis: A Narrative Review

Comparison of Fat Harvested from Flank and Falciform Regions for Stem Cell Therapy in Dogs

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