The combination of dextran sulphate and polyvinyl alcohol prevents excess aggregation and promotes proliferation of pluripotent stem cells in suspension culture

Objectives For clinical applications of cell-based therapies, a large quantity human pluripotent stem cells (hPSCs) produced in standardized and scalable culture processes is required. Currently, microcarrier-free suspension shows potential for large-scale expansion hPSCs; however, hPSCs tend to aggregate during culturing leading negative effect on cell yield. To overcome this problem, we developed novel protocol effectively control the sizes aggregates enhance proliferation suspension. Materials Methods were expanded supplemented with polyvinyl alcohol (PVA) dextran sulphate (DS), 3D formed under static or dynamic conditions. The as well pluripotency after assessed using counting, size analysis, real-time quantitative polymerase chain reaction, flow cytometry immunofluorescence staining, embryoid body formation, teratoma formation transcriptome sequencing. Results Our results demonstrated that addition DS alone prevented hPSC aggregation, while PVA significantly enhanced proliferation. combination not only promoted but also uniform size-controlled aggregates. Moreover, treated PVA, combination, maintained capable differentiating into all three germ layers. mRNA-seq analysis aggregation through improving energy metabolism-related processes, regulating growth, division, reducing adhesion among by affecting expression genes related adhesion. Conclusions represent significant step towards developing simple robust approach scale.