# Optimized derivation and culture system of human naïve pluripotent stem cells with enhanced DNA methylation status and genomic stability

## TL;DR

Researchers developed new culture systems for human naïve stem cells that improve DNA methylation and genomic stability.

## Contribution

New optimized media formulations were developed to sustain naïve pluripotent stem cells with better genomic stability.

## Key findings

- Four optimized media (LAY, LADY, LUDY, LKPY) effectively induce and maintain naïve pluripotent stem cells.
- Cells cultured in these media show improved DNA methylation and genomic stability.
- The LAY system most closely resembles pre-implantation DNA methylation patterns.

## Abstract

Human naïve pluripotent stem cells (PSCs) hold great promise for embryonic development studies. Existing induction and culture strategies for these cells, heavily dependent on MEK inhibitors, lead to widespread DNA hypomethylation, aberrant imprinting loss, and genomic instability during extended culture. Here, employing high-content analysis alongside a bifluorescence reporter system indicative of human naïve pluripotency, we screened over 1,600 chemicals and identified seven promising candidates. From these, we developed four optimized media—LAY, LADY, LUDY, and LKPY—that effectively induce and sustain PSCs in the naïve state. Notably, cells reset or cultured in these media, especially in the LAY system, demonstrate improved genome-wide DNA methylation status closely resembling that of pre-implantation counterparts, with partially restored imprinting and significantly enhanced genomic stability. Overall, our study contributes advancements to naïve pluripotency induction and long-term maintenance, providing insights for further applications of naïve PSCs.