Initial cell plating density affects properties of human primary synovial mesenchymal stem cells

Nakamura, Kaori; Tsuji, Kunikazu; Mizuno, Mitsuru; Koga, Hideyuki; Muneta, Takeshi; Sekiya, Ichiro

doi:10.1002/jor.24112

Cited by 15 publications

(14 citation statements)

References 22 publications

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“…In our recent report, we plated the synovial nucleated cells at 10 3 , 10 4 , or 10 5 cells/60 cm 2 dish and cultured them for 14 days. The fold increase was negatively related to plating density, whereas the total number of cells/dish was positively related to plating density [5]. In our current study, we did similar experiments at densities of every 300 cells/60 cm 2 dish rather than every 10 times that many cells/60 cm 2 dish, and similar results were obtained.…”

Section: Discussionsupporting

confidence: 82%

“…Third, this study did not consider chondrocyte differentiation potential. In our recent report, the cartilage pellet weight was negatively related to the initial plating density [5].…”

Section: Discussionmentioning

confidence: 78%

“…For these cell therapies, just after enzyme digestion, synovial nucleated cells are plated on a certain number of dishes and are cultured with 10% autologous human serum for 14 days. Important to remember is that the available number of nucleated cells and the serum volume can practically vary from patient to patient and that the initial plating cell density [5] and the dish number, which depends on the initial plating cell density, affect the yields (Fig. 1).…”

Section: Introductionmentioning

confidence: 99%

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Optimal initial cell density that yields the highest number of primary synovial mesenchymal stem cells in a clinical setting

Horiuchi

Mizuno

Katano

et al. 2020

Preprint

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Background Synovial mesenchymal stem cell (MSC) transplantation improved clinical outcomes in knees with cartilage defects and after repairs of degenerative torn menisci. For these, nucleated cells were cultured with 10% autologous serum for 14 days. Importantly, nucleated cell number and serum volume practically varied by patient, and initial cell density and dish number affected yield/donor. Our purpose was to examine the effect of plating density within range in clinical studies on fold increase and to estimate the MSC yield/donor, which are affected by the nucleated cell number and serum volume. Methods We obtained the data of the autologous serum volume, nucleated cell number, and initial plating cell density from 10 patients who participated in our previous clinical studies. Experimentally, synovial nucleated cells harvested from three patients with osteoarthritis during total knee arthroplasty were cultured with 10% mixed allogenic human serum for 14 days to examine the effect of initial cell density on fold increase. We used both our previous clinical data and our current experimental data to predict the yields of synovial MSCs in nine cases within the range of clinical studies. Results In the clinical studies, collected serum ranged between 87 and 186 ml, harvested nucleated cells ranged between 6.2 and 23 million, and the top five total MSC yields were obtained from donors when the initial plating cell density ranged from 919 to 1905 cells/cm 2 . Experimentally, fold increase significantly decreased between 700 and 1,900 cells/cm 2 . In cases where the cell number was low and the serum volume was high, the estimated yield/donor decreased along with the initial density, while in cases where the cell number was high and the serum volume was low, the estimated yield/donor increased along with the initial density. In cases both where some nucleated cells were discarded at low density and no cells were discarded at high density, the maximum yield/donor were obtained when the initial density was the lowest density that does not discard cells. Conclusions There was an optimal initial density that yields the highest number of primary MSCs in a clinical setting when cell numbers and human serum volumes are limited.

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Section: Discussionsupporting

confidence: 82%

“…Third, this study did not consider chondrocyte differentiation potential. In our recent report, the cartilage pellet weight was negatively related to the initial plating density [5].…”

Section: Discussionmentioning

confidence: 78%

Section: Introductionmentioning

confidence: 99%

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Optimal initial cell density that yields the highest number of primary synovial mesenchymal stem cells in a clinical setting

Horiuchi

Mizuno

Katano

et al. 2020

Preprint

Self Cite

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“…However, the MSCs do not show uniform colony-forming abilities 5–7 due to differences in their differentiation ability 8 . Interestingly, even when MSCs in the same population are cultured, the initial cell density affects the colony size, resulting in differences in cartilage differentiation ability 9 . Colony information could therefore predict the characteristics of the MSCs.…”

Section: Introductionmentioning

confidence: 99%

Time-lapse image analysis for whole colony growth curves and daily distribution of the cell number per colony during the expansion of mesenchymal stem cells

Mizuno

Katano

Shimozaki

et al. 2019

Sci Rep

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Mesenchymal stem cells from the synovium (synovial MSCs) are attractive for cartilage and meniscus regeneration therapy. We developed a software program that can distinguish individual colonies and automatically count the cell number per colony using time-lapse images. In this study, we investigated the usefulness of the software and analyzed colony formation in cultured synovial MSCs. Time-lapse image data were obtained for 14-day-expanded human synovial MSCs. The cell number per colony (for 145 colonies) was automatically counted from phase-contrast and nuclear-stained images. Colony growth curves from day 1 to day 14 (for 140 colonies) were classified using cluster analysis. Correlation analysis of the distribution of the cell number per colony at 14 days versus that number at 1–14 days revealed a correlation at 7 and 14 days. We obtained accurate cell number counts from phase-contrast images. Individual colony growth curves were classified into three main groups and subgroups. Our image analysis software has the potential to improve the evaluation of cell proliferation and to facilitate successful clinical applications using MSCs.

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“…To better understand the influence of culture conditions on stem cell potential, Nakamura et al examined the effects of initial cell plating density on the growth and multipotency of synovial stem cells and found that the initial significantly affects stem cell proliferation and chondrogenic potential.…”

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confidence: 99%

Journal of Orthopaedic Research: Special Issue on Stem Cells

Guilak

Sandell²,

Huard

2019

Journal Orthopaedic Research

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Initial cell plating density affects properties of human primary synovial mesenchymal stem cells

Cited by 15 publications

References 22 publications

Optimal initial cell density that yields the highest number of primary synovial mesenchymal stem cells in a clinical setting

Optimal initial cell density that yields the highest number of primary synovial mesenchymal stem cells in a clinical setting

Time-lapse image analysis for whole colony growth curves and daily distribution of the cell number per colony during the expansion of mesenchymal stem cells

Journal of Orthopaedic Research: Special Issue on Stem Cells

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