Expansion of human mesenchymal stem/stromal cells on temporary liquid microcarriers

BACKGROUND: Traditional large-scale culture systems for human mesenchymal stem/stromal cells (hMSCs) use solid microcarriers as attachment substrates. Although the use of such substrates is advantageous because of the high surface-to-volume ratio, cell harvest from the same substrates is a challenge as it requires enzymatic treatment, often combined with agitation. Here, we investigated a two-phase system for expansion and non-enzymatic recovery of hMSCs. Perfluorocarbon droplets were dispersed in a protein-rich growth medium and were used as temporary liquid microcarriers for hMSC culture. RESULTS: hMSCs successfully attached to these liquid microcarriers, exhibiting similar morphologies to those cultured on solid ones. Fold increases of 3.03 ± 0.98 (hMSC1) and 3.81 ± 0.29 (hMSC2) were achieved on day 9. However, the maximum expansion folds were recorded on day 4 (4.79 ± 0.47 (hMSC1) and 4.856 ± 0.7 (hMSC2)). This decrease was caused by cell aggregation upon reaching confluency due to the contraction of the interface between the two phases. Cell quality, as assessed by differentiation, cell surface marker expression and clonogenic ability, was retained post expansion on the liquid microcarriers. Cell harvesting was achieved non-enzymatically in two steps: first by inducing droplet coalescence and then aspirating the interface. Quality characteristics of hMSCs continued to be retained even after inducing droplet coalescence. CONCLUSION: The prospect of a temporary microcarrier that can be used to expand cells and then ‘disappear’ for cell release without using proteolytic enzymes is a very exciting one. Here, we have demonstrated that hMSCs can attach and proliferate on these perfluorocarbon liquid microcarriers while, very importantly, retaining their quality

Large Eddy Simulation of turbulent flowand heat transfer in a Kenics static mixer

CFD modelling of momentum and heat transfer using the Large Eddy Simulation (LES) approach hasbeen presented for a Kenics static mixer. The simulations were performed with the commercial codeANSYS Fluent 15 for turbulent flow of three values of Reynoldsnumber,Re=5 000, 10 000 and18 000. The numerical modelling began in the RANS model, where standardk−εturbulence modeland wall functions were used. Then the LES iterations started from the initial velocity and temperaturefields obtained in RANS. In LES, the Smagorinsky–Lilly modelwas used for the sub-grid scalefluctuations along with wall functions for prediction of flowand heat transfer in the near-wall region.The performed numerical study in a Kenics static mixer resulted in highly fluctuating fields of bothvelocity and temperature. Simulation results were presented and analysed in the form of velocity andtemperature contours. In addition, the surface-averaged heat transfer coefficient values for the wholeinsert length were computed and compared with the literature experimental data. Good compliance ofthe LES simulation results with the experimental correlation was obtained

Expansion of human mesenchymal stem/stromal cells on temporary liquid microcarriers

Background: Traditional large scale culture systems for human mesenchymal stem/stromal cells (hMSCs) use solid microcarriers as attachment substrates. While the use of such substrates is advantageous due to the high surface-to-volume ratio, cell harvest from the same substrates is a challenge as it requires enzymatic treatment, often combined with agitation. Here, we investigated a two-phase system for expansion and non-enzymatic recovery of hMSCs. Perfluorocarbon droplets were dispersed in a protein-rich growth medium and were used as temporary liquid microcarriers for hMSC culture. Results: hMSCs successfully attached to these liquid microcarriers exhibiting similar morphologies to those cultured on solid ones. Fold increases of 3.03±0.98 (hMSC1) and 3.81±0.29 (hMSC2) were achieved at day 9. However, the maximum expansion folds were recorded at day 4 (4.79±0.47 (hMSC1) and 4.856±0.7 (hMSC2)). This decrease was caused by cell aggregation upon reaching confluency due to the contraction of the interface between the two phases. Cell quality as assessed by differentiation, cell surface marker expression and clonogenic ability was retained post-expansion on the liquid microcarriers. Cell harvesting was achieved non-enzymatically in two steps, by firstly inducing droplet coalescence, then aspirating the interface. hMSCs’ quality characteristics continued to be retained even after inducing droplet coalescence.Conclusion: The prospect of a temporary microcarrier that can be used to expand cells and then ‘disappear’ for cell release without using proteolytic enzymes is a very exciting one. Here, we’ve demonstrated that hMSCs can attach and proliferate on these perfluorocarbon liquid microcarriers, while very importantly retaining their quality.</div