@misc{oai:niigata-u.repo.nii.ac.jp:00005501, author = {Nishiyama, Maki}, month = {Mar}, note = {Reproduction in gnathostomes (jawed vertebrates) is controlled by a hierarchically organized endocrine system called the hypothalamic-pituitary-gonadal axis. In spite of the diverged patterns of reproductive strategies and behaviors, this endocrine network is remarkably conserved throughout gnathostomes. Gonadotropins (GTHs), in response to hypothalamic gonadotropin-releasing hormone, are secreted from the pituitary and stimulate the gonads inducing the synthesis and release of sex steroid hormones, which in turn elicit growth and maturation of the gonads. Hagfishes, which lack both jaws and vertebrae, are considered the most primitive vertebrate known, living or extinct. Accordingly, studies on hagfishes are indispensable for understanding the origin and evolution of the reproductive endocrine system in vertebrates. Recently, single GTH has been identified from the pituitary gland of brown hagfish, Paramyxine atami. It is suggested that the pituitary-gonadal system has been established during the early evolution of agnathans (jawless vertebrates). At present, little is known about gonadal sex hormones in hagfish. The present study was designed to explore sex steroid hormonal profiles and their biosynthetic enzymes in relation to gonadal developments in the brown hagfish. Plasma concentrations of estradiol-17β, testosterone and progesterone were examined with respect to developmental conditions of gonads, sexual differences, and possible function of the atretic follicles in the brown hagflsh. using a time-resolved fluoroimmunoassay. Plasma concentrations of these three hormones were low in juveniles of both sexes. In females, plasma estradiol-17β showed a significant correlation with ovarian development, with the highest concentrations in late vitellogenic adults. Plasma testosterone and progesterone also increased significantly in non-vitellogenic adult females; however, plasma testosterone showed no significant differences among adult females at different ovarian developments, while plasma progesterone was significantly lower in late vitellogenic adults than it was in non-vitellogenic adults. Vitellogenic females that possessed atretic follicles showed significantly lower concentrations of all three hormones than females that possessed only normal follicles. In males, no significant differences were found in plasma estradiol or testosterone levels among groups of different developmental stages of the testis, while plasma progesterone showed an inverse relationship with testicular development. Thus, differences were found in plasma sex steroid hormone profiles between males and females., Sex steroids are synthesized from cholesterol by a series of steroid synthetic enzymes such as cytochrome P450 enzymes (CYP11A, CYP17 and CYP19) and hydroxysteroid dehydrogenases (3β-HSD and 17β-HSD). Cytochrome P450 side-chain cleavage enzyme (CYP11A) which catalyzes the first step in the production of steroid hormones, was cloned by EST analysis of the brown hagfish testis. Phylogenetic analysis demonstrated that the hagfish CYP11A was positioned within the clade of vertebrate CYP11A, and was separated from CYP11B. By in situ hybridization, CYP11A mRNA signals were found in the theca cells of the ovarian follicles and Leydig cells and the tubule-boundary cells of the testis: these cells are established as steroid producing cells in the gonads of gnathostomes. The real-time PCR analysis was performed to examine the functional significance of hagfish CYP11A in relation to the gonadal developments. In females, the highest levels of CYP11A mRNA expression were found in the late vitellogenic adults, and clear positive correlation was noted in CYP11A mRNA expression levels in relation to ovarian development. In males, CYP11A mRNA expression levels were significantly higher in adults with medium and large GSIs than those in juveniles and adults with small GSIs. Thus, there was a clear discrepancy between males and females on the relationship between the transcriptional levels of CYP11A and plasma steroid levels. These results suggest a possibility that male hagfish uses other steroids than estradiol or testosterone as major androgens. From these results, it is suggested that CYP11A plays functional roles as a steroidogenic enzyme in the gonadal developments. Moreover, hagfish GTH stimulated the transcriptional levels ofCYP11A in the cultured testis. This result further suggested that the steroidogenic activity of the hagfish testis is under the control of the pituitary GTH. Taking above-mentioned all results into consideration, it is suggested that vertebrates, during their early evolution, have established the pituitary-gonadal axis., 脊椎動物の多様な進化は、脳の神経情報を液性情報に変換・増幅して末梢の標的器官に送る情報伝達器官としての下垂体と、そこから分泌される腺下垂体ホルモンを介した視床下部-下垂体系を獲得したことで、もたらされたと考えられている。腺下垂体ホルモンの一つである生殖腺刺激ホルモン(Gonadotropin, GTH)は、顎を持つ脊椎動物(顎口類)において標的器官である生殖腺に作用し、様々な性ステロイドホルモンの合成・分泌を促す。それらのホルモンが生殖腺の発達や配偶子の形成・成熟、産卵行動などを誘起している。一方、脊椎動物の進化の最初期に出現した顎を持たない脊椎動物(無顎類)の遺存種であるヌタウナギ類は、化石種・現存種を含めて最も原始的な脊椎動物とされているが、その生態や生理学的機能などについてはほとんど調べられていない。従って、ヌタウナギ類の生殖内分泌機構を知ることは、この動物の生殖腺機能や、配偶子形成制御の理解だけでなく、脊椎動物の生殖内分泌機構の起源や、それがどのように進化し、多様性を生じたのかを考えるうえできわめて重要である。最近、ヌタウナギ類でも機能的なGTHが同定されたが、生殖腺から分泌されるホルモン分子についてはほとんど知られていない。本研究では、新潟県産クロヌタウナギ(Paramyxine atami)の性ステロイドホルモンの血中量を測定するとともに、ステロイド合成に関連する酵素群を探索し、得られた酵素の発現動態を生殖腺機能の調節と関連づけて理解することを目的とした。まず、クロヌタウナギ尾部から採血を行った後、メスについては卵の長軸の長さ(卵径)を、オスは体重当たりの精巣重量の割合[GSI-精巣重量(g)/体重(g)×100]を指標として、生殖腺の発達段階を区分した。そして時間分解蛍光免疫測定法(Time-Resolved-Fluoroimmunoassay、TR-FIA)によるクロヌタウナギ血液中のエストラジオール17β (E2)、テストステロン(T)、プロゲステロン(P)の測定系を確立した。メスでは血中E2量と生殖腺の発達との間に相関がみられ、卵黄形成の進んだ群で最も高い値を示した。血中TとP量は卵黄形成の未熟な成体群で最も高い値を示し、 P量については卵黄発達に伴い減少した。このことから、 E2がヌタウナギのメスにおいて生殖腺の発達、特に卵黄形成に関連しており、 TやPは中間産物として存在する可能性が考えられた。また、卵黄形成の途中で退化した退化卵胞をもつ個体群では、3ホルモンとも低い値を示したことから性ステロイドホルモンの合成が抑制されていると考えられた。一方、オスでは、生殖腺の発達と血中性ステロイドホルモン量の間に有意な相関は得られなかった。, 学位の種類: 博士(理学). 報告番号: 甲第3919号. 学位記番号: 新大院博(理)甲第388号. 学位授与年月日: 平成26年3月24日, 新大院博(理)甲第388号}, title = {Evolutionary studies on sex steroid hormone in the hagfish : plasma concentrations and biosynthetic enzymes}, year = {2014} }