部分细胞重编程可稳定恢复衰老表皮干细胞的干性。

Partial cellular reprogramming stably restores the stemness of senescent epidermal stem cells.

机构信息

State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Guangzhou, China.

出版信息

Eur Rev Med Pharmacol Sci. 2023 Jun;27(12):5397-5409. doi: 10.26355/eurrev_202306_32774.

DOI:10.26355/eurrev_202306_32774

PMID:37401274

Abstract

OBJECTIVE

Adult stem cell senescence and exhaustion are important drivers of organismal age. Restored stem cell self-renewal has revealed novel therapeutic targets for decreasing the incidence of age-associated diseases (AADs) and prolonging the human health span. Transient ectopic expression of the reprogramming factors Oct3/4, Sox2, Klf4 and c-Myc (collectively known as OSKM) in somatic cells can induce partial cellular reprogramming and effectively ameliorate their age-associated hallmarks. However, how this form of rejuvenation is applied to senescent stem cells remains unknown.

MATERIALS AND METHODS

The Integrin-α6highCD71high epidermal stem cells (ESCs) with low self-renewal ability were sorted by flow cytometry and then treated by the interrupted reprogramming induced by transient expression of OSKM. The ability of secondary clones' generation and self-proliferation in vitro, as well as stem cell marker p63, were detected to determine their self-renewal ability. Besides, gene and protein of epidermal cell markers were detected to determine whether their cell identities were retained. Finally, DNA methylation age (eAge) and DNA dehydroxymethylase/methyltransferase were analyzed to explore the alternation of their global DNA methylation pattern during this rejuvenation.

RESULTS

The partial reprogramming restored the youthful self-renewal and proliferation in senescent ESCs, including larger secondary clone generation, higher expression of stem cell marker p63 and proliferation marker Ki67, and faster proliferation speed, in each case without abolishing epithelial cellular identity. Moreover, the rejuvenation of adult stem cells could be maintained for 2 weeks after reprogramming factor withdrawal, which was more stable than that of differentiated somatic cells. Additionally, we found that partial reprogramming counteracted the acceleration of eAge in senescent epidermal stem cells and DNA methyltransferase 1 (DNMT1) may play a crucial role in this process.

CONCLUSIONS

Partial reprogramming has high therapeutic potential for reversing adult stem cell age, providing an advanced way to treat AADs.

摘要

目的

成体干细胞衰老和衰竭是机体衰老的重要驱动因素。恢复干细胞自我更新能力为降低与年龄相关疾病（AAD）的发病率和延长人类健康寿命提供了新的治疗靶点。在体细胞中瞬时异位表达重编程因子 Oct3/4、Sox2、Klf4 和 c-Myc（统称为 OSKM）可诱导部分细胞重编程，并有效改善其与年龄相关的特征。然而，这种形式的年轻化如何应用于衰老的干细胞尚不清楚。

材料和方法

通过流式细胞术分选具有低自我更新能力的整合素-α6highCD71high 表皮干细胞（ESCs），然后用 OSKM 瞬时表达诱导的中断重编程处理。通过检测二级克隆的体外生成和自我增殖能力以及干细胞标志物 p63 来确定其自我更新能力。此外，还检测了表皮细胞标志物的基因和蛋白，以确定其细胞身份是否保留。最后，分析 DNA 甲基化年龄（eAge）和 DNA 去羟甲基酶/甲基转移酶，以探讨在这种年轻化过程中其全基因组甲基化模式的改变。

结果

部分重编程恢复了衰老 ESCs 的年轻自我更新和增殖能力，包括更大的二级克隆生成、更高的干细胞标志物 p63 和增殖标志物 Ki67 的表达以及更快的增殖速度，每种情况均不破坏上皮细胞身份。此外，重编程因子撤出后，成体干细胞的年轻化可维持 2 周，比分化体细胞更稳定。此外，我们发现部分重编程可抵消衰老表皮干细胞中 eAge 的加速，并且 DNA 甲基转移酶 1（DNMT1）可能在该过程中起关键作用。