人原发性小肠恶性黑色素瘤裸小鼠原位移植肝转移模型的建立

目的为探讨小肠恶性黑色素瘤的发病机制和实验治疗提供理想的动物模型。方法将原发性小肠恶性黑色素瘤患者术中原发灶和肝转移灶新鲜瘤组织块分别植入裸小鼠的小肠黏膜层，观察原位移植成瘤率，移植瘤的侵袭和肝转移率。进行形态学光镜、电镜和免疫组织化学（免疫组化）]染色体核型和流式细胞分析。结果人小肠恶性黑色素瘤原发灶和肝转移灶新鲜瘤组织均获移植成功。建成一株人原发性小肠（原发灶）恶性黑色素瘤裸鼠原位移植模型（HSIM-0501）和一株人原发性小肠（肝转移灶）恶性黑色素瘤裸鼠原位移植肝转移模型（HSIM-0502）。移植瘤组织病理学为高度恶性黑色素瘤；免疫组化显示S-100蛋白；黑色素瘤单克隆抗体45阳性；电镜下瘤细胞质内可见大量黑色素颗粒及黑色素复合体。染色体众数55～59条；流式细胞DNA指数值1．49-1．61；均为异倍体。HSIM-0501和HSIM-0502分别传至25代和27代；共移植裸鼠317只；肿瘤移植成瘤率和液氮冻存复苏成活率均为100％。HSIM-0501肝转移率为46．2％，淋巴结转移率为36．7％；HSIM-0502肝转移率和淋巴结转移率均为100％。移植瘤在裸鼠小肠内自主侵袭生长，发生血液转移、淋巴结转移和腹腔内种植性转移。结论HSIM-0501和HSIM-0502是首次成功建立的人原发性小肠恶性黑色素裸鼠原位移植肝转移模型，可用于小肠恶性黑色素瘤的发病机制、侵袭和转移及抗转移实验治疗的研究。

Establishment of nude mice liver metastatic model of human primary malignant melanoma of the small intestine

OBJECTIVE: To provide ideal animal models for exploring pathogenesis and experimental therapy of primary malignant melanoma of the small intestine. METHODS: The histologically intact primary and liver metastatic fragments derived from surgical specimens of one patient with metastatic malignant melanoma of the small intestine were orthotopically implanted in the small intestinal mucous layer of nude mice. The take rate, invasion and liver metastasis were observed. Morphology (light microscopy, electron microscopy), immunophenotype analysis, flow cytometry and karyotype analysis were applied for the original human tumors and the transplanted tumors. RESULTS: The primary and liver metastatic fragments of malignant melanoma of the small intestine were successfully implanted in nude mice. After continuous passages in nude mice,an orthotopic model of human primary malignant melanoma of the small intestine(from the primary focus)in nude mice (termed HSIM-0501) and a liver metastatic model of human primary malignant melanoma of the small intestine (from the liver metastatic focus) in nude mice (termed HSIM-0502) were established. Histolocial examination of transplanted tumors revealed high-grade melanoma. S-100 protein and HMB45 were positive. Massive melanin granules and melanin complex were seen in cytoplasm of tumor cells.Chromosomal modal number was between 55 and 59. DNA index (DI) was 1.49-1.61, representing heteroploid. HSIM-0501 and HSIM-0502 were maintained for 25 and 27 passages in nude mice respectively. Three hundred and seventeen nude mice were used for transplantation. Both the take rate after transplantation and resuscitation rate of liquid nitrogen cryopreservation were 100%. HSIM-0501 exhibited 46.2% liver metastasis and 36.7% lymph node metastases. In HSIM-0502, both liver and lymph node metastases were 100%.The transplanted tumors autonomically and invasively grew in the small intestines of nude mice and hematogenous metastasis, lymph node metastasis and celiac planting metastasis occurred. CONCLUSION: Two nude mice liver metastatic models of human primary malignant melanoma of the small intestine are successfully established, which provide ideal animal models for the research of pathogenesis,metastasis biology and anti-metastatic experimental therapy of primary malignant melanoma of the small intestine.