小鼠睾丸器官组织的精子发生具有睾丸特异性结构,提高了生殖细胞存活率和睾酮产量。

IF 8.2 2区 医学 Q1 ENGINEERING, BIOMEDICAL
Guillaume Richer, Cleo Goyvaerts, Lorna Marchandise, Tamara Vanhaecke, Ellen Goossens, Yoni Baert
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引用次数: 0

摘要

本研究提出了一种双相方法来克服目前睾丸类器官(TO)培养的局限性,包括组织学异质性、生精细胞缺失和精子发生缺失。我们利用琼脂糖微孔从青春期前的C57BL/6J睾丸细胞中培养出睾丸器官。首先,通过比较α-MEM + 10% KSR培养基(已知可支持小鼠TO的生成)和三种优化培养基(1-3),重点是提高生精细胞在最初两周重组阶段的存活率。在优化了生殖细胞存活和细胞组织之后,评估了生长因子和通过消耗 CD45+ 免疫细胞或补充地塞米松(DEX)进行免疫调节的效果,以在随后的分化阶段促进精子发生。睾丸细胞自我重组成类似睾丸解剖单位的器官组织,其特征是由间质包围的一个管状结构。事实证明,在重组阶段,培养基1 3更有利于类器官的生长,培养基3中的睾丸细胞(7.4 ± 4.8%)与对照组(9.3 ± 5.3%)的生殖细胞数量相当。此外,在静态条件下,32×103 个细胞中有 37 ± 30% 显示出有组织的组织学。在分化阶段转用 α-MEM + 10% KSR,形成效率提高到 85 ± 7%,同时生殖细胞数量、睾酮产量(3.1 ± 0.9 ng/mL)和 γH2AX+ 类精子细胞的生成也有所增加(第 8-11 步,占总数的 1.2 ± 2.2%)。在α-MEM中加入分化因子后,精子样细胞数量增至2.9 ± 5.9%,CREM、TP1和PNA的阳性染色证实了这一点。不过,这些细胞仍为二倍体,核成熟不规则。补充 DEX 没有额外的效果,而免疫细胞耗竭则对 TO 的形成产生不利影响。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Spermatogenesis in mouse testicular organoids with testis-specific architecture, improved germ cell survival and testosterone production.

This study presents a biphasic approach to overcome the limitations of current testicular organoid (TO) cultures, including histological heterogeneity, germ cell loss and absence of spermatogenesis. Agarose microwells were utilized to create TOs from prepubertal C57BL/6 J testicular cells. First emphasis was on improving germ cell survival during the initial 2-week reorganization phase by comparingα-MEM + 10% knockout serum replacement (KSR) medium, known to support TO generation in mice, to three optimized media (1-3). Cell densities and culture dynamics were also tested to recreate histological resemblance to testes. After optimizing germ cell survival and cell organization, the effect of growth factors and immunomodulation through CD45+immune cell depletion or dexamethasone (DEX) supplementation were assessed for enhancing spermatogenesis during the subsequent differentiation phase. Testicular cells self-reorganized into organoids resembling the testicular anatomical unit, characterized by one tubule-like structure surrounded by interstitium. Media 1-3 proved superior for organoid growth during the reorganization phase, with TOs in medium 3 exhibiting germ cell numbers (7.4% ± 4.8%) comparable to controls (9.3% ± 5.3%). Additionally, 37% ± 30% demonstrated organized histology from 32 × 103cells under static conditions. Switching toα-MEM + 10% KSR during the differentiation phase increased formation efficiency to 85 ± 7%, along with elevated germ cell numbers, testosterone production (3.1 ± 0.9 ng ml-1) and generation ofγ-H2AX+spermatid-like cells (steps 8-11, 1.2% ± 2.2% of the total). Adding differentiation factors to theα-MEM increased spermatid-like cell numbers to 2.9% ± 5.9%, confirmed through positive staining for CREM, transition protein 1, and peanut agglutinin. Although, these remained diploid with irregular nuclear maturation. DEX supplementation had no additional effect, and immune cell depletion adversely impacted TO formation. The manipulability of TOs offers advantages in studying male infertility and exploring therapies, with scalability enabling high-throughput chemical screening and reducing animal usage in reproductive toxicity and drug discovery studies.

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来源期刊
Biofabrication
Biofabrication ENGINEERING, BIOMEDICAL-MATERIALS SCIENCE, BIOMATERIALS
CiteScore
17.40
自引率
3.30%
发文量
118
审稿时长
2 months
期刊介绍: Biofabrication is dedicated to advancing cutting-edge research on the utilization of cells, proteins, biological materials, and biomaterials as fundamental components for the construction of biological systems and/or therapeutic products. Additionally, it proudly serves as the official journal of the International Society for Biofabrication (ISBF).
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