Maghen Leila, Shlush Ekaterina, Gat Itai, Filice Melissa, Barretto Tanya, Jarvi Keith, Lo Kirk, Gauthier-Fisher Andree S, Librach Clifford L
L Maghen, Research, CReATe Fertility Centre, Toronto, Canada.
E Shlush, Research, CReATe Fertility Centre, Toronto, Canada.
Reproduction. 2016 Oct 25. doi: 10.1530/REP-16-0220.
The expansion of functional testicular biopsy-derived human spermatogonial stem cells (hSSC) ex-vivo may enable the restoration of fertility in pre-pubertal males having undergone gonadotoxic therapies or men with severe male factor infertility. Various somatic cells are known to regulate SSC homeostasis and spermatogenesis in the developing and adult testis. Prior attempts to recapitulate this niche demonstrated the requirement of feeder cells, such as endogenous testicular somatic cells, for germ cell expansion ex-vivo. However, this strategy has limitations for the expansion of hSSCs from tissue biopsies where spermatogenesis is absent or defective. Our aim was to evaluate first trimester human umbilical cord perivascular cells (FTM HUCPVCs), a novel source of mesenchymal stromal-like cells (MSCs), as potential human feeder cells for standardized hSSC expansion ex-vivo. Targeted RNA sequencing analysis demonstrated that CD90+ve FTM HUCPVCs expanded in vitro under germ cell culture conditions express a profile of targeted testicular-associated transcripts that is similar to cultured human CD90+ve testicular adherent cells (hTACs) and secrete LIF, FGF2 and BMP4, key growth factors known to regulate spermatogenesis. We also demonstrated that mitotically-inactivated FTM HUCPVCs support the expansion of mouse germ cells and putative SSCs ex-vivo, and that FTM HUCPVC transplantation promotes in vivo germ cell regeneration following mono-2- ethylhexyl phthalate (MEHP)-induced seminiferous tubule damage in a murine model, including a partial reconstitution of tubular cellular architecture and reestablishment of DAZL and acrosin+ve germ cell layers. Together, these data suggest that FTM HUCPVCs have phenotypical and functional properties that may support repair of the human testicular niche.
体外扩增功能性睾丸活检来源的人类精原干细胞（hSSC）可能使接受性腺毒性治疗的青春期前男性或患有严重男性因素不孕症的男性恢复生育能力。已知多种体细胞在发育中和成年睾丸中调节SSC稳态和精子发生。先前重现这种微环境的尝试表明，体外生殖细胞扩增需要饲养细胞，如内源性睾丸体细胞。然而，这种策略对于从不存在精子发生或有缺陷的组织活检中扩增hSSC存在局限性。我们的目的是评估孕早期人脐带血管周围细胞（FTM HUCPVCs），一种新型的间充质基质样细胞（MSCs）来源，作为体外标准化扩增hSSC的潜在人类饲养细胞。靶向RNA测序分析表明，在生殖细胞培养条件下体外扩增的CD90+ve FTM HUCPVCs表达与培养的人类CD90+ve睾丸贴壁细胞（hTACs）相似的靶向睾丸相关转录本谱，并分泌LIF、FGF2和BMP4，这些是已知调节精子发生的关键生长因子。我们还证明，有丝分裂失活的FTM HUCPVCs在体外支持小鼠生殖细胞和假定的SSC的扩增，并且FTM HUCPVC移植在小鼠模型中促进单-2-乙基己基邻苯二甲酸酯（MEHP）诱导的生精小管损伤后的体内生殖细胞再生，包括部分重建小管细胞结构以及重新建立DAZL和顶体素+ve生殖细胞层。总之，这些数据表明FTM HUCPVCs具有可能支持人类睾丸微环境修复的表型和功能特性。
