DERIVATION OF BABOON EMBRYONIC STEM CELL LINES

狒狒胚胎干细胞系的衍生

• 批准号: 6116388
• 负责人: Warren C LADIGES
• 金额: $ 8.67万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6116388
• 关键词: Mammalia; Primates; animal tissue

The overall objective of this protocol is to isolate and characterize baboon embryonic stem (ES) cells for the purpose of providing an in vitro model of early cell growth and proliferation similar to that in humans, and as a source of differentiation-driven cells, tissues, and organs. The baboon is used as a model of bone marrow transplantation, and an ES cell line will be a useful source of blood cell precursors, thus helping to conserve the number of animals for biomedical research. Five adult female baboons were observed on a daily basis and paired with adult male baboons at the onset of sexual receptivity, which was determined by change in the color and swelling of the perineal sex skin. Observations of mating behavior were recorded and the pair was separated when the female was no longer receptive (around the time of detumescence or decreasing color and swelling of the sex skin). Based on previous studies in the baboon, ovulation was estimated to occur three days before the onset of detumescence. Nonsurgical attempts to recover preimplantation embryos were made five to nine days after ovulation. Baboons were sedated with 0.2 ml/kg of ketamine/xylazine (0.70.3) and placed in lateral recumbency. A 14-gauge cannula (stainless steel tube) was passed through the vagina and cervix and into the uterus. Ultrasound was performed to confirm uterine placement. Sterile culture medium was flushed into the uterus and the back flow was collected into a sterile culture tube. A microscopic search for the embryo was made. Any embryos found were shipped via express mail to the Wisconsin Regional Primate Research Center for stem cell derivation. Over the past ten months, 12 flush attempts were made. Nine of these were attempted on animals that mated successfully. Four of these nine animals were flushed twice in one month, but the second flush also did not result in any embryos. One embryo was collected and sent to Wisconsin. It arrived safely and continued to do well in culture. Continued efforts will help assure the successful establishment of baboon ES cells as an important and useful in vitro cell culture system.

该方案的总体目的是隔离和表征狒狒胚胎（ES）细胞，以提供与人类相似的早期细胞生长和增殖的体外模型，以及作为分化驱动的细胞，组织和器官的来源。 狒狒被用作骨髓移植的模型，ES细胞系将是血细胞前体的有用来源，从而有助于保留生物医学研究的动物数量。 每天观察五个成年雌性狒狒，并在性接受开始时与成年雄性狒狒配对，这是由会阴性皮肤的颜色和肿胀的变化决定的。 记录了交配行为的观察结果，并在女性不再接受时（在驱散或降低性皮肤的颜色和肿胀时）时，对两对分开。 根据先前在狒狒中的研究，估计排卵发生在发作开始前三天。 排卵后五到九天，进行了恢复植入前胚胎的非外科尝试。 用0.2 mL/kg的氯胺酮/甲基嗪（0.70.3）镇静狒狒，并放置在侧向卧位中。 穿过阴道和子宫颈的14规套管（不锈钢管）进入子宫。 进行超声检查以确认子宫位置。 将无菌培养基冲入子宫，然后将后流收集到无菌培养管中。 对胚胎进行了微观搜索。 发现的任何胚胎都是通过快递邮件运送到威斯康星州的灵长类动物研究中心的。 在过去的十个月中，进行了12次冲洗尝试。 其中有九个是对成功交配的动物进行的。 这九只动物中有四只在一个月内冲洗了两次，但第二种动物也没有导致任何胚胎。 收集一个胚胎并送往威斯康星州。它安全地到达，并继续在文化中表现出色。 持续的努力将有助于确保成功建立狒狒ES细胞，这是一种重要且有用的体外细胞培养系统。