Capturing Human Naïve Pluripotency in the Embryo and in the Dish

Abstract

Although human embryonic stem cells (hESCs) were first derived almost 20 years ago, it was only recently acknowledged that they share closer molecular and functional identity to postimplantation lineage-primed murine epiblast stem cells than to naïve preimplantation inner cell mass-derived mouse ESCs (mESCs). A myriad of transcriptional, epigenetic, biochemical, and metabolic attributes have now been described that distinguish naïve and primed pluripotent states in both rodents and humans. Conventional hESCs and human induced pluripotent stem cells (hiPSCs) appear to lack many of the defining hallmarks of naïve mESCs. These include important features of the naïve ground state murine epiblast, such as an open epigenetic architecture, reduced lineage-primed gene expression, and chimera and germline competence following injection into a recipient blastocyst-stage embryo. Several transgenic and chemical methods were recently reported that appear to revert conventional human PSCs to mESC-like ground states. However, it remains unclear if subtle deviations in global transcription, cell signaling dependencies, and extent of epigenetic/metabolic shifts in these various human naïve-reverted pluripotent states represent true functional differences or alternatively the existence of distinct human pluripotent states along a spectrum. In this study, we review the current understanding and developmental features of various human pluripotency-associated phenotypes and discuss potential biological mechanisms that may support stable maintenance of an authentic epiblast-like ground state of human pluripotency.

Chemical Reversion of Conventional Human Pluripotent Stem Cells to a Naïve-like State with Improved Multilineage Differentiation Potency

“…The determinants of molecular and functional pluripotency, both in vitro and in the embryo were recently reviewed 2 . These factors include the genetic background, culture-associated acquisition of mutations for key developmental genes, and differences in hESC and hiPSC derivation and culture methodologies.…”

“…Although these markers do not discriminate between conventional and LIF-3i states, their levels inversely correlate with the frequency of spontaneous differentiation that may occur in hPSC when transitioning from conventional hPSC to LIF-3i conditions. Additional surface antigens that may more specifically mark human naïve-like states in vitro 2,12,13 can also be employed to detect effective LIF-3i hPSC reversion.…”

“…Since some of these systems have already been shown to generate hPSC populations with aberrant genomic and epigenetic configurations 2 , any naïve reversion method should be assayed with a number of N-hPSC of independent genetic backgrounds, in a manner that is sufficient to validate biological reproducibility and exclude non-developmentally relevant “pseudo-pluripotent” states ( i.e. , with apparent hallmarks of molecular pluripotency but lacking functional differentiation abilities).…”

“…, with apparent hallmarks of molecular pluripotency but lacking functional differentiation abilities). Zimmerlin et al further extended the validation of the LIF-3i culture system to include assaying the molecular and functional pluripotencies of reverted N-hiPSC derived from various reprogramming methods, which is another known putative contributor of functional variability between pluripotent states 2 .…”

“…However, 2i does not support the stable maintenance of human pluripotent stem cell (hPSC) lines 2 . The various complex small molecule, growth factor-supplemented, and transgenic approaches have recently been reported to capture putatively similar human naïve-like pluripotent molecular states 2 . However, many of the “naïve-like” states created with these methods also exhibited karyotypic instability, epigenomic defects ( e.g.…”