GMP-manufactured density gradient media for optimized mesenchymal stromal/stem cell isolation and expansion

Grisendi, Giulia; Annerén, Cecilia; Cafarelli, Luigi; Sternieri, Rita; Veronesi, Elena; Cervo, Gian Luca; Luminari, Stefano; Maur, M; Frassoldati, Antonio; Palazzi, Giovanni; Otsuru, Satoru; Bambi, Franco; Paolucci, Paolo; Conte, Pierfranco; Horwitz, Edwin M.; Dominici, Massimo

doi:10.3109/14653241003649510

Cited by 62 publications

(48 citation statements)

References 35 publications

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“…Even though MSC isolation and expansion by GMP manufactured gradient media have recently been reported [34], this study confirmed that directly plated BM offers a more advantageous method in terms of CFU-F number, minimal manipulation, hematopoietic contamination, and cellular growth (descriptive analysis). Our results (as has also recently been described by Capelli et al [35] and Lucchini et al [36]) show that whole BM separation methods represent a good procedure for MSC expansion for clinical use compared to MSCs obtained by gradient separation.…”

Section: Discussionsupporting

confidence: 75%

Multipotent Mesenchymal Stromal Stem Cell Expansion by Plating Whole Bone Marrow at a Low Cellular Density: A More Advantageous Method for Clinical Use

Mareschi

Rustichelli

Calabrese

et al. 2012

Stem Cells International

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Mesenchymal stem cells (MSCs) are a promising source for cell therapy due to their pluripotency and immunomodulant proprieties. As the identification of “optimal” conditions is important to identify a standard procedure for clinical use. Percoll, Ficoll and whole bone marrow directly plated were tested from the same sample as separation methods. The cells were seeded at the following densities: 100 000, 10 000, 1000, 100, 10 cells/cm2. After reaching confluence, the cells were detached, pooled and re-plated at 1000, 500, 100, and 10 cells/cm2. Statistical analyses were performed. Cumulative Population Doublings (PD) did not show significant differences for the separation methods and seeding densities but only for the plating density. Some small quantity samples plated in T25 flasks at plating densities of 10 and 100 cells/cm2 did not produce any expansion. However, directly plated whole bone marrow resulted in a more advantageous method in terms of CFU-F number, cellular growth and minimal manipulation. No differences were observed in terms of gross morphology, differentiation potential or immunophenotype. These data suggest that plating whole bone marrow at a low cellular density may represent a good procedure for MSC expansion for clinical use.

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Section: Discussionsupporting

confidence: 75%

Multipotent Mesenchymal Stromal Stem Cell Expansion by Plating Whole Bone Marrow at a Low Cellular Density: A More Advantageous Method for Clinical Use

Mareschi

Rustichelli

Calabrese

et al. 2012

Stem Cells International

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“…Previous findings indicate that MenSCs, a new source of MSCs, have been successfully differentiated into chondrogenic, adipogenic, osteogenic (20), neurogenic, and cardiogenic cell lineages (12). The results of these studies demonstrate the plasticity of MenSCs for potential research in regenerative medicine.…”

Section: Discussionmentioning

confidence: 99%

Effects of hypoxia on differentiation of menstrual blood stromal stem cells towards tenogenic cells in a co-culture system with Achilles tendon cells

Zheng

Zhou

Zhang

et al. 2017

Experimental and Therapeutic Medicine

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Achilles tendons have a very poor capacity for intrinsic regeneration. The cell-based treatment strategy for Achilles tendinitis includes the application of mesenchymal stem cells (MSCs), which have high proliferative and multipotent differentiation ability, and is a promising approach. The aim of the present study was to explore the tenogenic potential of human menstrual blood stromal stem cells (MenSCs) in a co-culture system and to compare the tenogenic capability under normoxic and hypoxic conditions. MenSCs were co-cultured indirectly with Achilles tendon cells in a Transwell co-culture system for 1, 2, or 3 weeks in two different concentrations of oxygen (20 and 2% O2), whereas the control contained only MenSCs. The extracellular matrix of MenSCs in each system was evaluated by Alcian blue staining assay, histological staining, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and western blot analysis. Alcian blue staining assay revealed a significant increase (P<0.05) in proteoglycan secretion by the differentiated MenSCs. Identical results were obtained by RT-qPCR for collagen I, which was validated by western blot analysis. Considerably increased collagen I and collagen III gene expression levels were exhibited by cells in the co-culture treatment group when compared with the control (P<0.05); however, no significant difference was detected between the normoxic (20% O2) and hypoxic treatment (2% O2) groups. RT-qPCR was utilized to determine the expression levels of thrombospondin 4, scleraxis and tenascin C in the differentiated MenSCs; a significant increase in the expression of these specific genes was indicated in the co-culture treatment group compared with the control (P<0.05). Although the expression levels were markedly higher in hypoxia than in normoxia conditions, this difference was not significant. To conclude, the present study indicated that MenSCs manifested a strong proliferative and multipotent capacity for differentiation and differentiated into Achilles tenogenic cells. Therefore, the use of MenSCs may be considered in Achilles tendinitis therapy.

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“…We separated cells on 1.073 g/mL Ficoll density gradient (GE Healthcare) to minimize loss of BMSC during separation steps. 33 The mononuclear cell fraction was collected and washed in PBS. We seeded 1-2.5 × 10 5 cells/cm 2 in α-MEM containing 10% lot selected FBS and 2 ng/ml bFGF.…”

Section: Bmsc Culturesmentioning

confidence: 99%

Silencing of RB1 and RB2/P130 during adipogenesis of bone marrow stromal cells results in dysregulated differentiation

et al. 2013

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GMP-manufactured density gradient media for optimized mesenchymal stromal/stem cell isolation and expansion

Cited by 62 publications

References 35 publications

Multipotent Mesenchymal Stromal Stem Cell Expansion by Plating Whole Bone Marrow at a Low Cellular Density: A More Advantageous Method for Clinical Use

Multipotent Mesenchymal Stromal Stem Cell Expansion by Plating Whole Bone Marrow at a Low Cellular Density: A More Advantageous Method for Clinical Use

Effects of hypoxia on differentiation of menstrual blood stromal stem cells towards tenogenic cells in a co-culture system with Achilles tendon cells

Silencing of RB1 and RB2/P130 during adipogenesis of bone marrow stromal cells results in dysregulated differentiation

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