成功地将培养和富集的青春期前雄鹿睾丸生殖细胞移植到经busulfan处理的同种受体。

IF 2.9 4区 生物学 Q1 ANATOMY & MORPHOLOGY
Shiva Pratap Singh, Suresh Dinkar Kharche, Yogesh Kumar Soni, Manisha Pathak, Ravi Ranjan, Sullip Kumar Majhi, Rajveer Singh Pawaiya, Manoj Kumar Singh, Manmohan Singh Chauhan
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引用次数: 0

摘要

本研究的目的是建立一种可行的方法,在超声(USG)引导下,利用睾丸内丁硫丹处理,将培养和富集的capine -male GC (cmgc)移植到同源受体体内,以产生生殖细胞(GC)缺失的山羊受体模型。评估移植后供体cmGCs的定植和精子小管(ST)正常结构的恢复。为此,对青春期前公山羊的cmGCs进行分离和富集,用差压法培养至第三代。然后,收集细胞,并使用兔抗cd90抗体进行磁活化细胞分选。在确认CD90+ cmgc的代谢活力(mtt法)和簇形成能力(结晶紫染色)后,用亲脂性红色荧光染料(PKH26)标记细胞,然后在USG指导下直接注射到受体公山羊的纵隔睾丸内。移植后8周观察到移植的CD90+ cmGCs在受体ST中的定殖和再种群。pkh26标记的供体细胞来源的菌落在酶解ST和受体睾丸冷冻切片中被鉴定。此外,组织化学分析显示,GC移植后受体睾丸的ST结构恢复正常。因此,结果表明,通过mGC治疗可以恢复不育动物的生殖能力,因此本文提出的方法可用于在受体动物睾丸中获得供体mGC衍生的功能性雄性配子。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Successful in vivo Transplantation of Cultured and Enriched Testicular Germ Cells of Pre-Pubertal Bucks to Busulfan-Treated Homologous Recipients.

The objective of the present study was to establish a workable approach for the production of germ cell (GC)-depleted recipient goat model using intra-testicular busulfan treatment and transplantation of cultured and enriched caprine-male GC (cmGCs) into the homologous recipients under ultrasonography (USG) guidance. The evaluation of post-transplantation colonization of donor cmGCs and restoration of the normal architecture of seminiferous tubules (ST) was performed. For this, the cmGCs of pre-pubertal male goats were isolated and enriched by differential platting for culture until the third passage. Thereafter, cells were harvested and further enriched by magnetic-activated cell sorting using rabbit-anti-CD90 antibody. After confirmation of metabolic viability (MTT-assay) and cluster-forming ability (crystal violet staining) of CD90+ cmGCs, the cells were labeled with a lipophilic red-fluorescent dye (PKH26) before transplanted into the recipient male goats by injection directly into the mediastinum testes under USG guidance. The colonization and repopulation of transplanted CD90+ cmGCs into the recipient ST was observed up to 8 weeks post-transplantation. The PKH26-labeled donor cell-derived colonies were identified in enzymatically digested ST and cryosections of recipient testes. Moreover, histochemical analyses revealed the restoration of the normal architecture of ST of recipient testis after GC transplantation. Therefore, the results suggest that the reproductive competence of infertile animals can be restored through mGC therapy and thus the methodology presented herein could be useful to obtain donor mGCs-derived functional male gametes in the recipient animal testis.

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来源期刊
Cells Tissues Organs
Cells Tissues Organs 生物-发育生物学
CiteScore
4.90
自引率
3.70%
发文量
45
审稿时长
6-12 weeks
期刊介绍: ''Cells Tissues Organs'' aims at bridging the gap between cell biology and developmental biology and the emerging fields of regenerative medicine (stem cell biology, tissue engineering, artificial organs, in vitro systems and transplantation biology). CTO offers a rapid and fair peer-review and exquisite reproduction quality. Special topic issues, entire issues of the journal devoted to a single research topic within the range of interests of the journal, are published at irregular intervals.
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