Accelerating development of stem cell-derived therapies using multiplexed microbioreactor arrays

Introduction: Effective use of human stem cells in regenerative medicine and drug screening depends on our ability to effectively direct both their undifferentiated expansion and differentiation into desired lineages. Exquisite control over stem cell fate is needed to efficiently produce sufficient, defined cell populations for such applications, yet this is substantially hindered by undefined culture components, signal crosstalk between multiple exogenous and endogenous factors, and spatiotemporal variations in microenvironmental composition inherent to conventional culture formats.Experimental: We have developed scalable, valveless, continuous-flow microbioreactor arrays that both provide a full-factorial set of exogenous factor compositions, and also allow controlled accumulation of paracrine factors. These arrays have been used to survey up to 8100 individual, perfused cellular microenvironments in parallel.Results: Through screens of pluripotency maintenance and mesendodermal differentiation of human pluripotent stem cells (hPSCs) as examples, we demonstrate the unique ability of this platform to separate, visualise, identify and modulate paracrine effects that are not otherwise readily accessible. In addition, we have applied this same technology platform to the optimisation of maintenance (and improvement) of potency of human mesenchymal stem cells, and the impact of small molecule agonists and antagonists on osteogenic differentiation outcomes. Most recently, with our 8100-chamber, high density microbioreactor array, we have assessed the impacts and interplay of small molecule agonists and growth factors on human cardiomyocyte proliferation. Importantly, optimization of these culture conditions with the arrays was readily translatable to improving differentiation in conventional static culture protocols, exemplifying the immediate practicality of the microbioreactor array platform in a range of stem cell screens.Conclusion: This multiplexed microbioreactor platform deciphers factor interplay and signalling hierarchies that control stem cell fate, and is applicable as a universal microenvironmental screening platform for bioprocess optimisation, media formulation design, quality control for cellular therapeutics and cell-based drug toxicity and discovery.