Pontes Kelly Cristine de Sousa, Groenewoud Arwin, Cao Jinfeng, Ataide Livia Maria Silva, Snaar-Jagalska Ewa, Jager Martine J
Department of Ophthalmology, Leiden University Medical Center, Leiden, The Netherlands.
Institute of Biology, Leiden University, Leiden, The Netherlands.
Invest Ophthalmol Vis Sci. 2017 Dec 1;58(14):6065-6071. doi: 10.1167/iovs.17-22023.
Conjunctival melanoma (CM) is a rare malignant disease that can lead to recurrences and metastases. There is a lack of effective treatments for the metastases, and we set out to develop a new animal model to test potential therapies. Zebrafish are being used as a model for many diseases, and our goal was to test whether this animal could be used to study CM.
Three human CM cell lines (CRMM-1 and CM2005.1, which both harbor a B-RAF mutation, and CRMM-2, which has an N-RAS mutation) were injected into the yolk sac, around the eye, and into the duct of Cuvier of transgenic (fli:GFP) Casper zebrafish embryos. Fluorescent and confocal images were taken to assess the phenotype and the behavior of engrafted cells and to test the effect of Vemurafenib as a treatment against CM.
While the cells that had been injected inside the yolk sac died and those injected around the eye sporadically went into the circulation, the cells that had been injected into the duct of Cuvier colonized the zebrafish: cells from all three cell lines proliferated and disseminated to the eyes, where they formed clusters, and to the tail, where we noticed extravasation and micrometastases. Vemurafenib, a potent agent for treatment of B-RAF V600E-positive melanoma, inhibited outgrowth of CRMM-1 and CM2005.1 cells in a mutation-dependent way.
The (fli:GFP) Casper zebrafish embryo can be used as an efficient animal model to study metastatic behavior of human CM cells and warrants further testing of drug efficacy to aid care of CM patients.
结膜黑色素瘤（CM）是一种罕见的恶性疾病，可导致复发和转移。目前对于转移灶缺乏有效的治疗方法，我们着手开发一种新的动物模型来测试潜在的治疗方法。斑马鱼正被用作多种疾病的模型，我们的目标是测试这种动物是否可用于研究CM。
将三种人CM细胞系（CRMM-1和CM2005.1，二者均携带B-RAF突变；以及CRMM-2，其具有N-RAS突变）注射到转基因（fli:GFP）Casper斑马鱼胚胎的卵黄囊、眼周和居维叶氏管中。拍摄荧光和共聚焦图像以评估移植细胞的表型和行为，并测试维莫非尼作为治疗CM的效果。
虽然注射到卵黄囊内的细胞死亡，注射到眼周的细胞偶尔进入循环，但注射到居维叶氏管的细胞在斑马鱼中定植：来自所有三种细胞系的细胞增殖并扩散到眼睛，在那里形成簇，还扩散到尾巴，在尾巴处我们观察到了外渗和微转移。维莫非尼是一种治疗B-RAF V600E阳性黑色素瘤的有效药物，以突变依赖的方式抑制CRMM-1和CM2005.1细胞的生长。
（fli:GFP）Casper斑马鱼胚胎可作为研究人CM细胞转移行为的有效动物模型，值得进一步测试药物疗效，以帮助CM患者的治疗。
