Dataset Information

KIF11 as a Potential Marker of Spermatogenesis Within Mouse Seminiferous Tubule Cross-sections.

ABSTRACT: The arrangement of immature germ cells changes regularly and periodically along the axis of the seminiferous tubule, and is used to describe the progression of spermatogenesis. This description is based primarily on the changes in the acrosome and the nuclear morphology of haploid spermatids. However, such criteria cannot be applied under pathological conditions with arrested spermatid differentiation. In such settings, the changes associated with the differentiation of premeiotic germ cells must be analyzed. Here, we found that the unique bipolar motor protein, KIF11 (kinesin-5/Eg5), which functions in spindle formation during mitosis and meiosis in oocytes and early embryos, is expressed in premeiotic germ cells (spermatogonia and spermatocytes). Thus, we aimed to investigate whether KIF11 could be used to describe the progression of incomplete spermatogenesis. Interestingly, KIF11 expression was barely observed in haploid spermatids and Sertoli cells. The KIF11 staining allowed us to evaluate the progression of meiotic processes, by providing the time axis of spindle formation in both normal and spermatogenesis-arrested mutant mice. Accordingly, KIF11 has the potential to serve as an excellent marker to describe spermatogenesis, even in the absence of spermatid development.

SUBMITTER: Hara-Yokoyama M

PROVIDER: S-EPMC6824007 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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KIF11 as a Potential Marker of Spermatogenesis Within Mouse Seminiferous Tubule Cross-sections.

Hara-Yokoyama Miki M Kurihara Hidetake H Ichinose Shozo S Matsuda Hironori H Ichinose Shizuko S Kurosawa Masaru M Tada Norihiro N Iwahara Chihiro C Terasawa Kazue K Podyma-Inoue Katarzyna A KA Furukawa Koichi K Iwabuchi Kazuhisa K

The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 20190819 11

The arrangement of immature germ cells changes regularly and periodically along the axis of the seminiferous tubule, and is used to describe the progression of spermatogenesis. This description is based primarily on the changes in the acrosome and the nuclear morphology of haploid spermatids. However, such criteria cannot be applied under pathological conditions with arrested spermatid differentiation. In such settings, the changes associated with the differentiation of premeiotic germ cells mus ...[more]

PMID: 31424977

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Project description:Research on spermatogonia is hampered by complex architecture of the seminiferous tubule, poor viability of testicular tissue ex vivo and lack of physiologically relevant long-term culture systems. Therefore there is a need for an in vitro model that would enable long term survival and propagation of spermatogonia. We aimed at the most simplified approach to enable all different cell types within the seminiferous tubules to contribute to the creation of a niche for spermatogonia. In the present study we describe the establishment of a co-culture of mouse testicular cells that is based on proliferative and migratory activity of seminiferous tubule cells and does not involve separation, purification or differential plating of individual cell populations. The co-culture is composed of the constituents of testicular stem cell niche: Sertoli cells [identified by expression of Wilm's tumour antigen 1 (WT1) and secretion of glial cell line-derived neurotrophic factor, GDNF], peritubular myoid cells (expressing alpha smooth muscle actin, αSMA) and spermatogonia [expressing MAGE-B4, PLZF (promyelocytic leukaemia zinc finger), LIN28, Gpr125 (G protein-coupled receptor 125), CD9, c-Kit and Nanog], and can be maintained for at least five weeks. GDNF was found in the medium at a sufficient concentration to support proliferating spermatogonial stem cells (SSCs) that were able to start spermatogenic differentiation after transplantation to an experimentally sterile recipient testis. Gdnf mRNA levels were elevated by follicle-stimulating hormone (FSH) which shows that the Sertoli cells in the co-culture respond to physiological stimuli. After approximately 2-4 weeks of culture a spontaneous formation of cord-like structures was monitored. These structures can be more than 10 mm in length and branch. They are formed by peritubular myoid cells, Sertoli cells, fibroblasts and spermatogonia as assessed by gene expression profiling. In conclusion, we have managed to establish in vitro conditions that allow spontaneous reconstruction of testicular cellular microenvironments.

Project description:Spermatogenesis in mammals is a cyclic process of spermatogenic cell development in the seminiferous epithelium that can be subdivided into 12 subsequent stages. Histological staging analysis of testis sections, specifically of seminiferous tubule cross-sections, is the only effective method to evaluate the quality of the spermatogenic process and to determine developmental defects leading to infertility. Such staging analysis, however, is tedious and time-consuming, and it may take a long time to become proficient. We now have developed a Computerized Staging system of Spermatogenesis (CSS) for mouse testis sections through learning of an expert with decades of experience in mouse testis staging. The development of the CSS system comprised three major parts: 1) Developing computational image analysis models for mouse testis sections; 2) Automated classification of each seminiferous tubule cross-section into three stage groups: Early Stages (ES: stages I-V), Middle Stages (MS: stages VI-VIII), and Late Stages (LS: stages IV-XII); 3) Automated classification of MS into distinct stages VI, VII-mVIII, and late VIII based on newly developed histomorphological features. A cohort of 40 H&E stained normal mouse testis sections was built according to three modules where 28 cross-sections were leveraged for developing tubule region segmentation, spermatogenic cells types and multi-concentric-layers segmentation models. The rest of 12 testis cross-sections, approximately 2314 tubules whose stages were manually annotated by two expert testis histologists, served as the basis for developing the CSS system. The CSS system's accuracy of mean and standard deviation (MSD) in identifying ES, MS, and LS were 0.93 ± 0.03, 0.94 ± 0.11, and 0.89 ± 0.05 and 0.85 ± 0.12, 0.88 ± 0.07, and 0.96 ± 0.04 for one with 5 years of experience, respectively. The CSS system's accuracy of MSD in identifying stages VI, VII-mVIII, and late VIII are 0.74 ± 0.03, 0.85 ± 0.04, and 0.78 ± 0.06 and 0.34 ± 0.18, 0.78 ± 0.16, and 0.44 ± 0.25 for one with 5 years of experience, respectively. In terms of time it takes to collect these data, it takes on average 3 hours for a histologist and 1.87 hours for the CSS system to finish evaluating an entire testis section (computed with a PC (I7-6800k 4.0 GHzwith 32GB of RAM & 256G SSD) and a Titan 1080Ti GPU). Therefore, the CSS system is more accurate and faster compared to a human histologist in staging, and further optimization and development will not only lead to a complete staging of all 12 stages of mouse spermatogenesis but also could aid in the future diagnosis of human infertility. Moreover, the top-ranking histomorphological features identified by the CSS classifier are consistent with the primary features used by histologists in discriminating stages VI, VII-mVIII, and late VIII.

Project description:Study questionCan transplanted primate testicular cells form seminiferous tubules de novo, supporting complete spermatogenesis?Summary answerCryopreserved testicular cells from a prepubertal monkey can reorganize in an adult monkey recipient testis forming de novo seminiferous tubular cords supporting complete spermatogenesis.What is known alreadyDe novo morphogenesis of testicular tissue using aggregated cells from non-primate species grafted either subcutaneously or in the testis can support spermatogenesis.Study design, size, durationTwo postpubertal rhesus monkeys (Macaca mulatta) were given testicular irradiation. One monkey was given GnRH-antagonist treatment from 8 to 16 weeks after irradiation, while the other received sham injections. At 16 weeks, cryopreserved testicular cells from two different prepubertal monkeys [43 × 106 viable (Trypan-blue excluding) cells in 260 μl, and 80 × 106 viable cells in 400 μl] were transplanted via ultrasound-guided injections to one of the rete testis in each recipient, and immune suppression was given. The contralateral testis was sham transplanted. Testes were analyzed 9 months after transplantation.Participants/materials, setting, methodsSpermatogenic recovery was assessed by testicular volume, weight, histology and immunofluorescence. Microsatellite genotyping of regions of testicular sections obtained by LCM determined whether the cells were derived from the host or transplanted cells.Main results and the role of chanceTransplanted testis of the GnRH-antagonist-treated recipient, but not the sham-treated recipient, contained numerous irregularly shaped seminiferous tubular cords, 89% of which had differentiating germ cells, including sperm in a few of them. The percentages of donor genotype in different regions of this testis were as follows: normal tubule, 0%; inflammatory, 0%; abnormal tubule region, 67%; whole interior of abnormal tubules, >99%; adluminal region of the abnormal tubules, 92%. Thus, these abnormal tubules, including the enclosed germ cells, were derived de novo from the donor testicular cells.Large scale dataNot applicable.Limitations, reasons for cautionThe de novo tubules were observed in only one out of the two monkeys transplanted with prepubertal donor testicular cells.Wider implications of the findingsThese findings may represent a promising strategy for restoration of fertility in male childhood cancer survivors. The approach could be particularly useful in those exposed to therapeutic agents that are detrimental to the normal development of the tubule somatic cells affecting the ability of the endogenous tubules to support spermatogenesis, even from transplanted spermatogonial stem cells.Study funding/competing interest(s)This work was supported by research grants P01 HD075795 from Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD/NIH) to K.E.O and Cancer Center Support Grant P30 CA016672 from NCI/NIH to The University of Texas MD Anderson Cancer Center. The authors declare that they have no competing interests.

Dataset Information

KIF11 as a Potential Marker of Spermatogenesis Within Mouse Seminiferous Tubule Cross-sections.

Publications

KIF11 as a Potential Marker of Spermatogenesis Within Mouse Seminiferous Tubule Cross-sections.

Similar Datasets

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