Epigenetic dynamics during capacitation of naïve human pluripotent stem cells [PBAT]. Epigenetic dynamics during capacitation of naïve human pluripotent stem cells [PBAT]

Human pluripotent stem cells (hPSCs) are of fundamental relevance in regenerative medicine and the primary source for many novel cellular therapies. The development of naïve culture conditions has led to the expectation that these naïve hPSCs could overcome some of the limitations found in conventional (primed) hPSCs culture conditions, including recurrent epigenetic anomalies. Recent work has shown that transition to the primed state (or capacitation) is necessary for naïve hPSCs to acquire multi-lineage differentiation competence. This pluripotent state transition may recapitulate essential features of human peri-implantation development. Here we studied epigenetic changes during the transition between naïve and primed pluripotency, examining global genomic redistribution of histone modifications, chromatin accessibility, and DNA methylation, and correlating these with gene expression. We identify CpG islands, enhancers, and retrotransposons as hotspots of epigenetic dynamics between pluripotency states. Our results further reveal that hPSC resetting and subsequent capacitation rescue X chromosome-linked epigenetic erosion and reduce the ectoderm-biased gene expression of conventional primed hPSCs. Overall design: 2 lines of human naïve pluripotent stem cells (embryo-derived HNES1 and chemically reset cR-H9-EOS) were cultured in N2B27 and 2uM XAV939 for 10 days. After that the cells were split into two conditions: N2B27 + 2uM XAV939 + 3ng/ml Activin A + 10ng/ml FGF2 (XAF), or E8 medium, for extended maintenance. The experiment was performed with the cells on day 0 and 10, when the cells were cultured in XAV939; and one time point after transfer to maintenance conditions, at not less than 22 days of culture from the start of the experiment. Conventional hPSC cell line H9-EOS, which was a parental line for the chemically reset cR-H9-EOS was used as a control.

人类多能干细胞（human pluripotent stem cells, hPSCs）在再生医学领域具有核心研究价值，亦是众多新型细胞疗法的核心来源。原始态培养体系（naïve culture conditions）的建立，使得人们期待这类原始态人类多能干细胞能够克服常规（始发态（primed））人类多能干细胞培养体系存在的部分局限，包括反复出现的表观遗传异常。近期研究表明，原始态人类多能干细胞向始发态的转变（或称致敏过程）是其获得多谱系分化潜能的必要条件。该多能状态转变或可重现人类着床前后发育的关键特征。本研究聚焦原始态与始发态多能状态转变过程中的表观遗传变化，系统分析了组蛋白修饰、染色质开放状态及DNA甲基化的全基因组分布重编程，并将其与基因表达谱进行关联分析。本研究鉴定出CpG岛、增强子及逆转录转座子为多能状态转换过程中表观遗传动态变化的热点区域。研究结果进一步表明，人类多能干细胞的重置（resetting）及后续致敏过程可挽救X染色体连锁的表观遗传侵蚀，并改善常规始发态人类多能干细胞偏向外胚层的基因表达模式。实验设计如下：两株人类原始态多能干细胞（分别为胚胎来源的HNES1及化学重置得到的cR-H9-EOS）在添加2μM XAV939的N2B27培养基中培养10天。随后将细胞分为两组进行后续维持培养：一组为添加2μM XAV939、3ng/ml激活素A（Activin A）及10ng/ml成纤维细胞生长因子2（FGF2）的N2B27培养基（简称XAF培养基），另一组为E8培养基。本实验分别在培养第0天（即初始XAV939培养阶段）、第10天采集细胞样本，并在细胞转移至维持培养体系后、距实验起始至少22天的时间点采集样本。本实验以化学重置株cR-H9-EOS的亲本细胞系——常规人类多能干细胞系H9-EOS作为对照。