Identification of spermatogonial stem cell-specific surface molecules is important in understanding the molecular mechanisms underlying the maintenance and differentiation of these cells. We have found that spermatogonia from busulfan treated mice expressed an autoantigen that distinguishes between undifferentiated and differentiated spermatogonia. Four to six weeks after busulfan treatment, germ cells located in the basal compartment of seminiferous epithelium show isotype-specific IgG deposits that form due to autoimmunity. Before busulfan treatment, the level of testicular IgG was very low but IgG levels began to increase after week 4 and peaked at week 6. When cells from the busulfan treated testis were analyzed using laser scanning cytomeoy (LSC), the frequency of cells positive for IgG deposits, 6-integrin, and 1-integrin were 16.53.8%, 11.82.6%, and 9.0 1.4%, respectively. Immunofluorescent staining suggested that most, if not all of the cells with IgG-deposits isolated from a laminin-coated dish, were also positive for a spermatogonial stem cell marker \ulcorner6-integrins as well as for a germ cell-specific marker TRA 98. We determined serum and intratesticular IgG levels and the soundness of seminiferous tubule basement membrane from busulfan treated mice using electron microscopy, in order to study the mechanism responsible for IgG deposits in spermatogonia. We found that the basement membranes of seminiferous tubules from busulfan treated mice were severely impaired when compared to those of normal adult, neonates and w/wv mice. Furthermore, new blood cells were observed in the surface of the damaged basement membrane along the seminiferous tubules. These results suggest that the IgG in spermatogonial stem cells accumulates from circulating blood through the impaired basement membranes induced by busulfan treatment. Taken together, our study suggests that IgG can be used as a new marker for undifferentiated spermatogonia cells.

The conserved THO/TREX complex is participated in pre-mRNA processing and mRNA nuclear export. In Metazoa, it has been reported that THO/TREX is loaded on nascent RNA transcribed by RNA polymerase II in a splicing-dependent manner; however, how THO/TREX functions is poorly understood. To understand the role of THO/TREX in eukaryotic gene expression, we investigated the role of THO/TREX in Drosophila germline, and found that lack of THOC5, a component of THO/TREX, showed defects in the biogenesis of piRNA, a distinct class of small non-coding RNAs that control expression of transposable elements (TE) in the Drosophila germline. Genome wide RNA-seq showed that THOC5 and other TREX components are essential for the biogenesis of piRNA. THO/TREX components are enriched on piRNA precursors transcribed from dual-strand piRNA clusters and co-localize in distinct nuclear foci that overlap with sites of piRNA transcription. The localization of TREX in nuclear foci and its loading on piRNA precursor transcripts depends on Cutoff, a protein associated with chromatin of piRNA clusters. We also show that TREX is required for accumulation of nascent piRNA precursors, suggesting that TREX is required for their efficient transcription. In addition to the biogenesis of piRNA, both THO and piRNA mutants showed over-proliferation of germline stem cell-like cells outside of stem cell niche. Taken together our study reveals a novel splicing-independent mechanism for THO/TREX loading on nascent RNA and its importance in piRNA biogenesis as well as a role of piRNA in the differentiation of germline stem cell.

The ultrastructures of germ cells and the functions of Leydig cells and Sertoli cells during spermatogenesis in male Kareius bicoloratus (Pleuronectidae) were investigated by electron microscope observation. Each of the well-developed Leydig cells during active maturation division and before spermiation contained an ovoid vesicular nucleus, a number of smooth endoplasmic reticula, well-developed tubular or vesicular mitochondrial cristae, and several lipid droplets in the cytoplasm. It is assumed that Leydig cells are typical steroidogenic cells showing cytological characteristics associated with male steroidogenesis. No cyclic structural changes in the Leydig cells were observed through the year. However, although no clear evidence of steroidogenesis or of any transfer of nutrients from the Sertoli cells to spermatogenic cells was observed, cyclic structural changes in the Sertoli cells were observed over the year. During the period of undischarged germ cell degeneration after spermiation, the Sertoli cells evidenced a lysosomal system associated with phagocytic function in the seminiferous lobules. In this study, the Sertoli cells function in phagocytosis and the resorption of products originating from degenerating spermatids and spermatozoa after spermiation. The spermatozoon lacks an acrosome, as have been shown in all teleost fish spermatozoa. The flagellum or sperm tail of this species evidences the typical 9+2 array of microtubules.

Epigenetic change is dynamic during germ cell development. DNA methylation and histone modification are the most important epigenetic process to regulate the gene expressions. They are very close reciprocal relationship on the specific genomic regions called CpG islands (CGI). The CGIs are located on the promoter regions and recruit various epigenetic regulators including, CFP1, KDM2A, KDM2B, TET1 and MLL. They contain a common domain which is the zinc finger CxxC domain. The CxxC domain reads non-methylated CpG and recruits other regulatory elements such as SET1, PRC, COMPASS and SIN3A to modify Histone proteins. CFP1 contains a CxxC domain. CFP1 protein therefore imposes an ability to distinguish its important regulatory element, “non-methylated CpG” from the genome. After binding the CpG, CFP1 recruits SET1 complex, which is involved in the histone H3 lysin 4 (H3K4) methylation. However, the functional consequence of CFP1 in the germ cell development remains unknown. In this study, we demonstrated that CFP1 is critical for the both spermatogenesis and oogenesis using conditional knockout system.

The ultrastructural characteristics of germ cell differentiations during spermatogenesis and mature sperm morphology in male () were evaluated via transmission electron microscopic observation. The accessory cells, which contained a large quantity of glycogen particles and lipid droplets in the cytoplasm, are assumed to be involved in nutrient supply for germ cell development. Morphologically, the sperm nucleus and acrosome of this species are ovoid and conical in shape, respectively. The acrosomal vesicle, which is formed by two kinds of electron-dense or lucent materials, appears from the base to the tip: a thick and slender elliptical line, which is composed of electron-dense opaque material, appears along the outer part (region) of the acrosomal vesicle from the base to the tip, whereas the inner part (region) of the acrosomal vesicle is composed of electron-lucent material in the acrosomal vesicle. Two special characteristics, which are found in the acrosomal vesicle of A. () in Pinnidae (subclass Pteriomorphia), can be employed for phylogenetic and taxonomic analyses as a taxonomic key or a significant tool. The spermatozoa were approximately in length, including a sperm nucleus (about in length), an acrosome (about in length), and a tail flagellum (about ). The axoneme of the sperm tail evidences a 9+2 structure.

Spermatogenesis and taxonomic values of mature sperm morphology of in male Septifer (Mytilisepta) virgatus were investigated by transmission electron microscope observations. The morphologies of the sperm nucleus and the acrosome of this species are the cylinder shape and cone shape, respectively. Spermatozoa are approximately 45-50 in length including a sperm nucleus (about 1.26 long), an acrosome (about 0.99 long), and tail flagellum (about 45-47 ). Several electron-dense proacrosomal vesicles become later the definitive acrosomal vesicle by the fusion of several Golgi-derived vesicles. The acrosome of this species has two regions of differing electron density: there is a thin, outer electron-dense opaque region (part) at the anterior end, behind which is a thicker, more electron-lucent region (part). In genus Septifer in Mytilidae, an axial rod does not find and also a mid-central line hole does not appear in the sperm nucleus. However, in genus Mytilus in Mytilidae, in subclass Pteriomorphia, an axial rod and a mid-central line hole appeared in the sperm nucleus. These morphological differences of the acrosome and sperm nucleus between the genuses Septifer and Mytilus can be used for phylogenetic and taxonomic analyses as a taxonomic key or a significant tool. The number of mitochondria in the midpiece of the sperm of this species are five, as seen in subclass Pteriomorphia.

본 연구는 수컷 살조개 Protothaca (Notochione) yessoensis의 정자 형성 과정 중 생식세포들의 분화와 성숙정자의 미세구조 특징에 관한 몇 가지 특징을 투과전자현미경 관찰에 의해 조사하였다. 본 종의 정자형태는 원시형(primitive type)으로, 이매패강, 이치아강(Heterodonta)에 속하는 다른 종들과 유사하다. 생식세포에 인접하여 연결되어 있는 보조세포들은 생식세포들의 발달을 위해 영양공급에 관여한다. 본 종의 정자의 핵형은 긴 원통형이며 첨체의 형태는 모자모양이다. 정자는 길이가 대략 46～50 ㎛이며, 길다란 정핵(길이 약 2.44 ㎛)과, 첨체(길이 0.45 ㎛), 그리고 미부 편모(약 42～46 ㎛)로 이루어져 있다. 미부 편모의 악소님(axoneme)은 9+2 구조를 나타낸다. 첨체소포의 특징으로써 basal ring의 기저부 위에서 측면부위는 전자밀도가 불투명한 부위를 나타내나, 첨체소포의 앞쪽 정단부위는 전자밀도가 비교적 투명한 부위로 나타나는 특징을 보인다. 이것이 이치아강에 속하는 백합과와 또 다른 여러 과들에 속하는 종들의 정자들이 갖는 첨체소포의 공통특징이다. 따라서 이치아강이 갖는 이들 첨체소포가 갖는 공통특징은 분류의 key 또는 중요한 도구로써 계통․분류를 위해 사용될 수 있다. 정자 중편에 있는 미토콘드리아 수는 4개로 이치아강 내에서 백합과의 3종을 제외한 모든 종들과 다른 과들의 종들에서 공통으로 나타나고 있는데, 예외로, 개조개, 백합, 가무락조개 만은 중편의 미토콘드리아가 5개로 이루어져 있다. 미토콘드리아 수는 과나 또는 상과 수준에서 종들의 분류학적 분석을 할 경우, 분류 key 또는 중요한 도구로 사용될 수 있다.

난자 및 정자형성과정은 성에 따라 차이가 일부 있지만 생식줄기세포 (germ-line stem cells)의 증식을 통해 그 수를 유지하고, 그 중 일부 또는 전부가 감수분열에 진입하여 생식세포인 정자와 난자를 생산한다. 따라서 생식세포의 형성과정은 세포의 증식과 성장, 분화 기작을 연구할 수 있는 매우 훌륭한 시스템으로 여겨져 과거로부터 많은 연구가 진행되어왔다. 더욱이 인간에서 보조생식술과 동물 생명공학의 발전을 통해 그 중요성은 더욱 커지고 있으나, 그 체외배양 모델시스템이 없어 최근까지의 연구에는 많은 어려움이 있었다. 과거에 많은 연구자들에 의해 생식세포의 배양, 정소와 난소의 기관 배양 등이 시도된 바 있으나, 대부분 단기간 내 성공일 뿐 장기간 배양에는 어려움이 있었다. 하지만 교토대학의 Shinohara 교수팀에 의해 체외에서 정원줄기세포의 장기간 증식 배양법이 확립되었고, 또 배양된 정원줄기세포가 정소 세정관 내에 이식을 통해 정자형성과정을 재현할 수 있음이 확인되어 이 분야의 연구와 응용연구는 활성화되기 시작했다. 또한, 최근 들어 배아줄기세포, 성체줄기세포와 같은 여러 종류의 전분화능 줄기세포에서 생식줄기세포 또는 생식세포로의 분화가 일부 성공됨에 따라 생식세포의 증식과 분화의 조절기작이 일부 밝혀지고 있다. 따라서 이러한 연구성과 는 현재까지 치료가 불가능한 비폐쇄성 무정자증환자의 불임치료 방법으로의 발전 가능성을 제시할 뿐 아니라 동물생명공학분야로의 응용가능성을 더욱 높여주고 있다. 본 연제에서는 실험동물과 인간의 줄기세포로부터 생식세포의 분화에 관한 최근 연구를 정리하고, 이를 이용한 기초연구, 산업, 임상치료에의 적용가능성을 제시하고자 한다.

This study attempted to verify the possibility of using germ cell aspiration (GCA) method as a non-fatal technique in studying the life-history of equilateral venus, Gomphina veneriformis (Veneridae) and granular ark, Tegillarca granosa (Arcidae). Using twenty-six gauge 1/2" (12.7mm) needle, GCA was carried out in equilateral venus through external ligament. In granular ark, GCA was performed by preventing closure of the shells by inserting a tongue depressor between the shells while still open. The success rate of sex identification using the GCA method was 95.6% for the equilateral venus (n=650/680) and 94.3% for the granular ark (n=707/750). Mortality of equilateral venus, which spent 33 days under wild conditions, was 13.8% (n=90/650) while the mortality of granular ark, which spent 390 days under wild conditions, was 2.4% (n=17/707). Although we believe that GCA does not appear to cause death in equilateral venus or granular ark, the success rate in employing of this methodology may differ depending on the level of proficiency of the researcher and reproductive stage of the bivalve. This study concludes that GCA is a convenient non-fatal methodology, which can be employed to identify sex and investigate gonadal maturity in Gomphina veneriformis and Tegillarca granosa.

1998년 3월부터 1999년 2월까지 전라남도 대흑산도 연안에서 채집한 비단가리비를 대상으로 생식소중량지수, 생식세포 분화 및 난소주기를 조직, 세포학적 관찰에 의해 조사하였다. 초기 난황형성 난모세포에서, 골지체, 미토콘드리아 및 조면소포체들은 지방적 형성에 관여하였다. 후기난황형성난모세포에서 생식상피상에 존재하는 외인성 물질들 즉, 글리코겐 입자들 및 지방 과립상 물질들이 난황막의 미세융모를 통해서 난모세포의 난질로 통과해 들어갔다. 후기난황형성난

Primordial germ cell (PGC) is the progenitor cell of the germ cell lineage and eventually give rise to gametes that are responsible for creating individual organisms via a fertilization process. This means that PGC is a unique cell that can be converted into individual fish. This advantage of PGCs would make it possible to develop various applications in the field of fish bioengineering. First, PGCs may make it easier to preserve the genetic resources of fish. Cryopreservation of fish eggs or embryos has not been successfully achieved so far. Therefore, the only possible method to preserve genetic resources of fishes is to raise fish as live individuals. If PGCs isolated from various fishes could be cryopresewed, these cells could be converted into live fishes via germ-line chimera production. This is particularly useful for preserving genetic materials of endangered species. Even if the species of interest were to become extinct, it could be recovered by the transplantation of cryopreserved PGCs into the embryos of a closely related species. Another application of this technology is in what could be termed "surrogate broodstock technology". (중략)