鸡胚原始生殖细胞的建系方法及其影响因素

Establishing method and influencing factors of embryonic primordial germ cells in chick

学术背景:胚胎原始生殖细胞经体外抑制分化培养可得到胚胎生殖细胞,是胚胎干细胞的又一来源。以其作为外源基因的载体用于制作嵌合体、基因敲除等具有很高的实用价值。目的:概括性论述鸡胚胎原始生殖细胞建系方法、影响因素及应用前景等问题。检索策略:由该论文的研究人员应用计算机检索Pubmed数据库1990-01/2006-04的相关文献,检索词"embryos,primordialgermcells,embryonicstemcells,embryonicgermcells,genitalridge,Invitroculture",并限定文章语言种类为English。同时计算机检索中国期刊全文数据库2000-04/2007-04的相关文献,检索词"胚胎,原始生殖细胞,胚胎干细胞,胚胎生殖细胞,生殖嵴,体外培养",并限定文章语言种类为中文。共检索到60篇文献,对资料进行初审,纳入标准:①与鸡胚胎原始生殖细胞的建系、影响因素及应用密切相关。②同一领域选择近期发表或在权威杂志上发表的文章。排除标准:重复性研究。文献评价:文献的来源主要是通过对鸡胚胎原始生殖细胞建系方法、影响因素及应用前景方面内容进行汇总分析。所选用的30篇文献中,7篇为综述,其余均为临床或基础研究。资料综合:①原始生殖细胞离体培养的关键是保证细胞具备无限增殖能力的同时维持其未分化状态。目前主要通过选择鸡胚原始生殖细胞的适宜分离时期和方法、合理设计培养基、采用饲养层细胞培养以及添加抑制分化的细胞因子等方法来解决。②鸡胚原始生殖细胞的生物学检测主要包括形态学鉴定、碱性磷酸酶活性检测、PAS染色检测、阶段特异性胚胎表面抗原1检测、核型分析、分化试验以及嵌合体试验等方法。③鸡胚原始生殖细胞的应用研究广泛,相比较桑椹胚和囊胚体外培养获取干细胞优势明显;为基础研究提供了最原始的发育生物学模型;生产克隆动物,降低禽类的保种成本;生产药物蛋白,是生产治疗用重组蛋白极为理想的来源。结论:尽管目前有关鸡原始生殖细胞的培养与应用已取得一定进展,但在生产实践上如何设计出更适合于鸡原始生殖细胞生长的离体培养体系、如何更高效方便地获得鸡原始生殖细胞、如何提高利用原始生殖细胞制作嵌合体的成功率等问题还有待进一步深入研究。

BACKGROUND： Embryonic germ cells can be obtained by inhibiting differentiation and in vitro culture of the embryonic primordial germ cells （PGCs）, and they are another source of embryonic stem cells. As the foreign gene vector, the embryonic PGCs have highly practical value for chimera and gene knockout. OBJECTIVE： To discuss the method of establishing the chicken embryonic PGCs, and their influencing factor and application prospect. RETRIEVE STRATEGY： An online search of PubMed database was undertaken to identify the English articles published from January 1990 to April 2006 by using the key words of ＂embryos, primordial germ cells, embryonic stem cells, embryonic germ cells, genital ridge, in vitro culture＂. Meanwhile, Chinese Journal Full-Text Database was also searched for relevant articles published between April 2000 and April 2007 with the same key words in Chinese. Totally 60 articles were collected, and the data were checked in the first trial. Inclusive criteria：①literatures were related to the method of establishing the chicken embryonic PGCs, their influencing factor and application prospect. ②articles announcing in the near future or on authority magazine with the same field were selected. Exclusive criteria： reduplicative studies. LITERATURE EVALUATION： All the included articles summarized the method of establishing the chicken embryonic PGCs, their influencing factor and application prospect. Among 30 articles met the criteria, 7 ones were reviews while others were clinical or basic researches. DATA SYNTHESIS： ① The key that in vitro culture of PGCs is to insure the unlimited growth and maintain undifferentiation state of cells. The present countermeasure is mainly choosing the optimal period and method of the chicken embryo PGCs, designing medium rationally, adopting feeder layer as the culture method and adding the inhibit-differentiation cytokines. ② Biological detections of chicken embryonic PGCs mainly include the morphology identification, alkaline phosphatase activity detection, PAS dyeing detection, stage specific embryonic surface antigen-1 detection, karyotype analysis, differentiation experiment, chimera trial, and so on.③Chicken embryonic PGCs are applied widely and have the advantage about gaining stem cells compared with the mulberry embryo and the blastula culture in vitro; Provide the most primitive developing biological model for the basic research; Produce the clone animal and reduce the cost of species maintenance for domestic fowls; Produce medicine protein and become the extremely ideal source for producing recombinant protein for curing use. CONCLUSION： Although there have been an advanced progress in the culture and application of chicken PGCs, however, some practical problems need to be further explored, such as how to design the culture system of in vitro more suitable chicken PGCs, how to obtain chicken PGCs more high-effectively and more conveniently, and how to improve the success rate of making chimera from PGCs.