CAJ | 학술논문

通过黄鳝成体性腺组织的体外培养,探讨黄鳝成体性腺生殖干细胞体外分离培养与鉴定的方法.分离黄鳝性腺组织进行植块培养,倒置相差显微镜下观察细胞生长状况.利用H.E、油红O染色、碱性磷酸酶(AP)细胞化学染色等方法对培养细胞进行鉴定.黄鳝成体性腺组织原代培养3～5 d,可见组织周围有细胞迁出,并形成生长晕.15 d左右,生长晕直径可达3～3.2 mm,生长晕中的细胞主要包括:成纤维细胞、神经样细胞、类生殖干细胞三种类型.成纤维细胞呈不对称的长梭形,突起短而多,细胞核较浅,核仁明显,有2～3个核仁,是生长晕的主要细胞成分.神经样细胞具有放射丝状细胞突起,油红O染色呈阳性反应.类生殖干细胞呈圆形,单核或多核,核质比大,AP染色呈现出阳性反应.探索了适于黄鳝成体性腺组织体外培养的条件和方法,并对原代培养的细胞进行了初步鉴定,为进一步利用体外培养的黄鳝成体性腺生殖干细胞开展黄鳝性逆转分子机制研究奠定了基础.
통과황선성체성선조직적체외배양,탐토황선성체성선생식간세포체외분리배양여감정적방법.분리황선성선조직진행식괴배양,도치상차현미경하관찰세포생장상황.이용H.E、유홍O염색、감성린산매(AP)세포화학염색등방법대배양세포진행감정.황선성체성선조직원대배양3～5 d,가견조직주위유세포천출,병형성생장훈.15 d좌우,생장훈직경가체3～3.2 mm,생장훈중적세포주요포괄:성섬유세포、신경양세포、류생식간세포삼충류형.성섬유세포정불대칭적장사형,돌기단이다,세포핵교천,핵인명현,유2～3개핵인,시생장훈적주요세포성분.신경양세포구유방사사상세포돌기,유홍O염색정양성반응.류생식간세포정원형,단핵혹다핵,핵질비대,AP염색정현출양성반응.탐색료괄우황선성체성선조직체외배양적조건화방법,병대원대배양적세포진행료초보감정,위진일보이용체외배양적황선성체성선생식간세포개전황선성역전분자궤제연구전정료기출.
Objective:To culture gonad tissues of Monopterus Albus and To explore the methods of the purification,culture and identification of the germ line stem cells of Monopterus Albus in vitro.Method:The tissue block pasted culture method were used to culture gonad tissues of Monopterus Albus and the germ line stem cells of Monopterus Albus was observed under inverted phase contrast microscope.The cultured cells were identified by H.E,Oil red O staining,alkaline phosphatase(AP)cell staining.Results:The cells migrated out around tissues after 3-5 days culturing and the growth halo emerged.About 15 days,the diameter of the growth halo is up to 3～3.2 mm.the cells of the growth halo include three types:fibroblasts,nerve-like cells,and germ stem cell similar cells.Fibroblast was the main components of the growth halo.Its body was flat.The morphology was unregular and there were many short prominences and the light nuclear of cell had 2-3 obvious nucleoluses.the main components of the growth halo.The morphology of Nerve-like cells.Nerve-like cells with radiation filamentous cell protrusions were positive in oil red O staining.Germ stem eell similar cells are round,single-core or multi-core,and the ratio of the nucleus and cytoplasm is mass.AP staining shows positive.Conclusion:the suitable conditions and methods for the cultivation of gonad tissues of Monopterus Albus in vitro and preliminary identificated the cells of primary culture.Therefore,this study has laid a foundation for the cultivation of the germ line stem cells of Monopterus Albus in vitro as well as for the mechanism of sex reversal of Monopterus Albus on the molecule level.