AMPK activation reverts mouse epiblast stem cells to naive state

Abstract

Despite increasing knowledge on primed and naive pluripotency, the cell signaling that regulates the pluripotency type in stem cells remains not fully understood. Here we show that AMP kinase (AMPK) activators can induce the reversion of primed mouse epiblast stem cells (mEpiSCs) to the naive pluripotent state. The addition of AMPK activators alone or together with leukemia inhibitory factor to primed mEpiSCs induced the appearance of naive-like cells. After passaging in naive culture conditions, the colony morphology, protein expression, and global gene expression profiles indicated the naive state, as did germline transmission ability. Loss-of-function and gain-of-function studies suggested that p38 is a critical downstream target in AMPK activation. Finally, single-cell RNA sequencing analysis revealed that the reversion process through AMPK signaling passes an intermediate naive-like population. In conclusion, the AMPK pathway is a critical driving force in the reversion of primed to naive pluripotency.

AMPK-p38 axis converts human pluripotent stem cells to naïve state

“…We have previously reported that p38 is a critical downstream of AMPK in the maintenance and induction of the naïve state of mouse PSCs (Liu et al, 2019; Liu et al, 2021). Then, we examined the involvement of p38 in the AICAR-elicited naïve conversion of human PSCs.…”

“…Recently, We have reported that the treatment with AICAR or the expression of a constitutive active form of p38 (CA-p38) can maintain the naïve state of mouse ESCs and convert mouse EpiSC to naïve mouse ESCs (Liu et al, 2019; Liu et al, 2021). Based on these data, we examined whether the treatment with AICAR or CA-p38 could convert primed human PSCs to the naïve state.…”

“…We recently demonstrated that the AMPK-p38 axis is a pathway involved in the maintenance of the naïve state of mouse ESCs (Liu et al, 2019), and it can convert primed mouse Epi-SCs to the naïve state (Liu et al, 2021). Though the core signaling that induces naïve conversion is the same between mouse and human cells, several culture conditions are different (Martin et al, 1981;Bredenkamp et al, 2019).…”

“…AICAR (Fujifilm, 1mM) was added to the PXGL medium (Liu et al, 2019; Liu et al, 2021), and VPA (Guo et al, 2017) was added to Ndiff227 basal medium with PD0325901 (Sigma, 1 μM), human Lif (Fujifilm, 10 ng/ml), and penicillin/streptomycin) for naïve state induction. Cells were passaged on MEF feeder layers every 5-10 days as single cells using TrypLE™ Select CTS™ (Gibco).…”

“…As a breakthrough for this situation, we recently demonstrated that activating the 5’ adenosine monophosphate-activated protein kinase (AMPK) pathway with the simple addition of a single chemical AMPK activators can convert primed state mouse epiblast stem cells (Epi-SCs) to the naïve state embryonic stem cells (Liu et al, 2021). AMPK is an enzyme that is activated when it senses an increase in the AMP/ATP ratio due to energy deficit caused by exercise or caloric restriction and is responsible for regulating cellular energy homeostasis.…”