省内动态
中科院广州生物院赖良学研究组等发表大动物疾病模型研究进展
2017年1月4日，国际学术期刊《human molecular genetics》在线发表了中国科学院广州生物医药与健康研究院赖良学研究组、樊娜娜博士和南方医科大学南方医院的曾抗教授研究组合作的研究成果 ，论文题为“Generation of Hoxc13 knockout pigs recapitulates human ectodermal dysplasia–9”。论文通过基因编辑技术培育出的无毛小猪，可精准模拟人类毛发缺失的疾病病理，用于人类的毛发再生研究。 赖良学研究员及樊娜娜博士、曾抗教授为本文的共同通讯作者。
本研究利用精确基因编辑技术模拟人的纯发-甲型外胚叶发育不良症(ED-9)，针对其致病基因Hoxc13基因进行突变，成功培育出ED-9模型猪。当前，各种原因导致的脱发和少发疾病，给患者带来了很大困扰。其中，纯发-甲型外胚层发育不良症(ED-9)是一种先天性疾病，患者除毛发缺失和甲营养不良外，并无其他缺陷。在小鼠中，研究人员对ED-9致病基因Hoxc13进行敲除后发现，除全身无毛和趾发育异常外，还存在骨异常及存活率低等问题。小鼠模型不能完全模拟出人类的ED-9疾病病症。此外，小鼠短寿命短、体型小，不利于有效性和安全性长效研究。
近年来，猪被认为是一种理想的动物模型。猪皮肤在结构、厚度色素沉着及血液供应方面都与人类更为相似。在这项研究中，研究人员采用单链寡核苷酸(ssODNs)结合CRISPR/Cas9和体细胞核移植，通过在猪中模拟人类Hoxc13的无义突变，成功制备了Hoxc13缺失猪模型。该猪模型除全身毛发缺失和蹄发育异常外无小鼠模型存在的其他缺陷，忠实模拟了人类ED-9疾病病症，因此，ED-9模型猪可用于人类ED-9疾病病理和毛发再生研究。另外，该模型除毛发缺失和蹄发育不良外，无其他症状。Hoxc13缺失猪可替代无毛小鼠作为一种无毛实验动物模型，广泛应用于皮肤晒伤、光老化、伤口愈合、化妆品测试及皮肤癌等研究。
图：广州生物院在大动物疾病模型的研究中取得进展
原文摘要：Atrichia and sparse hair phenotype cause distress to many patients.Ectodermal dysplasia–9 (ED–9) is a congenital condition characterized by hypotrichosis and nail dystrophy without other disorders, and Hoxc13 is a pathogenic gene for ED–9. However, mice carrying Hoxc13 mutation present several other serious disorders, such as skeletal defects, progressive weight loss and low viability. Mouse models cannot faithfully mimic human ED–9. In this study, we generated an ED–9 pig model viaHoxc13 gene knockout through single-stranded oligonucleotides (c.396C?>?A) combined with CRISPR/Cas9 and somatic cell nuclear transfer. Eight cloned piglets with three types of biallelic mutations (five piglets with Hoxc13c.396C?>?A/c.396C?>?A, two piglets with Hoxc13c.396C?>?A/c.396C?>?A?+?1 and one piglet with Hoxc13Δ40/Δ40) were obtained.Hoxc13 was not expressed in pigs with all three mutation types, and the expression levels of Hoxc13-regulated genes, namely, Foxn1, Krt85 and Krt35, were decreased. The hair follicles displayed various abnormal phenotypes, such as reduced number of follicles and disarrayed hair follicle cable without normal hair all over the body. By contrast, the skin structure, skeleton phenotype, body weight gain and growth of Hoxc13knockout pigs were apparently normal. The phenotypes of Hoxc13mutation in pigs were similar to those in ED–9 patients. Therefore,Hoxc13 knockout pigs could be utilized as a model for ED–9 pathogenesis and as a hairless model for hair regeneration research. Moreover, the hairless pigs without other major abnormal phenotypes generated in this study could be effective models for other dermatological research because of the similarity between pig and human skins.
