Sperm DNA methylation is predominantly stable in mice offspring born after transplantation of long-term cultured spermatogonial stem cells.

IF 5.7 2区 医学 Q1 Medicine Clinical Epigenetics Pub Date : 2023-04-07 DOI:10.1186/s13148-023-01469-x
Joana B Serrano, Nils C Tabeling, Cindy M de Winter-Korver, Saskia K M van Daalen, Ans M M van Pelt, Callista L Mulder
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引用次数: 2

Abstract

Background: Spermatogonial stem cell transplantation (SSCT) is proposed as a fertility therapy for childhood cancer survivors. SSCT starts with cryopreserving a testicular biopsy prior to gonadotoxic treatments such as cancer treatments. When the childhood cancer survivor reaches adulthood and desires biological children, the biopsy is thawed and SSCs are propagated in vitro and subsequently auto-transplanted back into their testis. However, culturing stress during long-term propagation can result in epigenetic changes in the SSCs, such as DNA methylation alterations, and might be inherited by future generations born after SSCT. Therefore, SSCT requires a detailed preclinical epigenetic assessment of the derived offspring before this novel cell therapy is clinically implemented. With this aim, the DNA methylation status of sperm from SSCT-derived offspring, with in vitro propagated SSCs, was investigated in a multi-generational mouse model using reduced-representation bisulfite sequencing.

Results: Although there were some methylation differences, they represent less than 0.5% of the total CpGs and methylated regions, in all generations. Unsupervised clustering of all samples showed no distinct grouping based on their pattern of methylation differences. After selecting the few single genes that are significantly altered in multiple generations of SSCT offspring compared to control, we validated the results with quantitative Bisulfite Sanger sequencing and RT-qPCRin various organs. Differential methylation was confirmed only for Tal2, being hypomethylated in sperm of SSCT offspring and presenting higher gene expression in ovaries of SSCT F1 offspring compared to control F1.

Conclusions: We found no major differences in DNA methylation between SSCT-derived offspring and control, both in F1 and F2 sperm. The reassuring outcomes from our study are a prerequisite for promising translation of SSCT to the human situation.

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精子DNA甲基化在长期培养的精原干细胞移植后出生的小鼠后代中主要是稳定的。
背景:精原干细胞移植(SSCT)被建议作为儿童癌症幸存者的生育治疗方法。SSCT首先在促性腺毒性治疗(如癌症治疗)之前进行睾丸活检冷冻保存。当儿童癌症幸存者成年后想要生儿育女时,活检解冻,ssc在体外繁殖,随后自动移植回他们的睾丸。然而,长期繁殖过程中的培养胁迫会导致sscc发生表观遗传变化,如DNA甲基化改变,并可能被SSCT后出生的后代遗传。因此,SSCT需要在临床实施这种新型细胞疗法之前对衍生后代进行详细的临床前表观遗传学评估。为此,在多代小鼠模型中,使用减少代表性亚硫酸盐测序研究了体外繁殖的ssct衍生后代精子的DNA甲基化状态。结果:尽管存在一些甲基化差异,但在所有世代中,它们占总CpGs和甲基化区域的比例不到0.5%。所有样本的无监督聚类显示没有基于甲基化差异模式的明显分组。在选择了在多代SSCT后代中与对照相比显著改变的少数单基因后,我们用亚硫酸氢盐Sanger定量测序和RT-qPCRin各器官验证了结果。只有Tal2被证实存在差异甲基化,在SSCT后代的精子中被低甲基化,在SSCT F1后代的卵巢中与对照F1相比表达更高。结论:我们发现,在F1和F2精子中,ssct衍生的后代和对照组之间的DNA甲基化没有重大差异。我们研究的令人放心的结果是SSCT有希望转化为人类情况的先决条件。
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来源期刊
Clinical Epigenetics
Clinical Epigenetics Biochemistry, Genetics and Molecular Biology-Developmental Biology
CiteScore
8.90
自引率
5.30%
发文量
150
审稿时长
12 weeks
期刊介绍: Clinical Epigenetics, the official journal of the Clinical Epigenetics Society, is an open access, peer-reviewed journal that encompasses all aspects of epigenetic principles and mechanisms in relation to human disease, diagnosis and therapy. Clinical trials and research in disease model organisms are particularly welcome.
期刊最新文献
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