Top-Down Inhibition (TDi) and Baseline Activation (BLa): Controlling Signal Transduction When Endogenous Cytokines are Ruining Your Differentiation

In the 20 years since the first human pluripotent stem cell (hPSC) lines were established, there have been a plethora of protocols developed that allow us to generate a wide range of human cell types in vitro. Efforts to achieve a greater degree of specificity and efficiency in generating desired cell types have resulted in increasingly complex approaches. The magnitude and timing of signals has become key, and the concept of a “fully defined” system is a forever sought-after goal with shifting goalposts. This overview discusses two related approaches that can be used to deliver a tightly regulated, intermediate-strength signal, and which can also manage the impact of endogenous signaling variation and enable a switch away from bovine serum albumin–containing medium to a better-defined system without suffering a subsequent loss of robustness or efficiency. The approaches, referred to as top-down inhibition and baseline activation, were developed to deliver intermediate levels of BMP and WNT signaling during neural crest induction from hPSC, but could be applied to a variety of other signals and differentiation systems. © 2019 by John Wiley & Sons, Inc.