骨髓间充质干细胞逆转衰老颗粒细胞中与卵巢衰老相关的 m6A RNA 甲基化修饰谱。

Bone Marrow Mesenchymal Stem Cells Reversed Ovarian Aging-related m6A RNA Methylation Modification Profile in Aged Granulosa Cells.

机构信息

The Basic Medical Laboratory of the 920Th Hospital of Joint Logistics Support Force of PLA, The Transfer Medicine Key Laboratory of Cell Therapy Technology of Yunan Province, The Integrated Engineering Laboratory of Cell Biological Medicine of State and Regions, Kunming, 650032, Yunnan Province, China.

The Affiliated Stomatology of Kunming Medical University, Kunming, 650106, Yunnan Province, China.

出版信息

Stem Cell Rev Rep. 2023 May;19(4):953-967. doi: 10.1007/s12015-022-10485-y. Epub 2023 Jan 7.

DOI:10.1007/s12015-022-10485-y

PMID:36609903

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10185602/

Abstract

BACKGROUND

Ovarian ageing causes endocrine disturbances and the degeneration of systemic tissue and organ functions to seriously affect women's physical and mental health, and effective treatment methods are urgently needed. Based on our previous studies using juvenile rhesus monkey bone marrow mesenchymal stem cells (BMMSCs) to treat ovarian ageing in rhesus monkey, we found that BMMSCs improved ovarian structure and function. This study continues to explore the mechanism by which BMMSCs reversed granulosa cell (GC) ageing.

METHODS

A GC ageing model and coculture system of BMMSCs were established, changes in the level of the N6-methyladenosine (m6A) methylation modification were detected, m6A-modified RNA immunoprecipitation sequencing (MeRIP-seq) were performed, correlations between m6A peaks and mRNA expression were determined, and the expression of hub genes was identified using Q-PCR, immunofluorescence staining, and western blot.

RESULTS

Our results showed that HO successfully induced GC ageing and that BMMSCs reversed measures of GC ageing. BMMSCs increased the expression of the FTO protein and reduced the overall level of m6A. We identified 797 m6A peaks (348 hypomethylated and 449 hypermethylated peaks) and 817 differentially expressed genes (DEGs) (412 upregulated and 405 downregulated) after aged GCs were cocultured with BMMSCs, which significantly associated with ovarian function and epigenetic modification. The epigenetic repressive mark and important cell cycle regulator lysine demethylase 8 (KDM8) was downregulated at both the mRNA and protein levels, histone H3 was upregulated in aged GCs after BMMSC coculture, and KDM8 was upregulated after FTO was inhibited through FB23.

CONCLUSIONS

Our study revealed an essential role for m6A in BMMSCs in reversing GC ageing, and FTO regulated KDM8 mediates histone H3 changes may as a novel regulatory mechanism in BMMSCs to reverse GC ageing.

摘要

背景

卵巢衰老导致内分泌紊乱和全身组织器官功能退化，严重影响女性身心健康，急需有效的治疗方法。基于我们之前使用幼年恒河猴骨髓间充质干细胞（BMMSCs）治疗恒河猴卵巢衰老的研究，我们发现 BMMSCs 改善了卵巢结构和功能。本研究继续探索 BMMSCs 逆转颗粒细胞（GC）衰老的机制。

方法

建立 GC 衰老模型和 BMMSCs 共培养系统，检测 m6A 修饰水平变化，进行 m6A 修饰 RNA 免疫沉淀测序（MeRIP-seq），确定 m6A 峰与 mRNA 表达的相关性，采用 Q-PCR、免疫荧光染色和 Western blot 鉴定关键基因的表达。

结果

我们的结果表明，HO 成功诱导 GC 衰老，BMMSCs 逆转了 GC 衰老的指标。BMMSCs 增加了 FTO 蛋白的表达，降低了整体 m6A 水平。我们发现，与 BMMSCs 共培养的衰老 GC 后有 797 个 m6A 峰（348 个低甲基化峰和 449 个高甲基化峰）和 817 个差异表达基因（DEGs）（412 个上调基因和 405 个下调基因），这些基因与卵巢功能和表观遗传修饰显著相关。在 BMMSC 共培养的衰老 GC 中，表观遗传抑制标记物和重要的细胞周期调节因子赖氨酸去甲基酶 8（KDM8）的 mRNA 和蛋白水平均下调，组蛋白 H3 在 BMMSC 共培养后的衰老 GC 中上调，并且 FTO 抑制后 KDM8 上调通过 FB23。