人原发性结直肠恶性淋巴瘤裸小鼠原位移植高转移模型的建立

Establishment of high metastasis models of human primary colorectal lymphoma in nude mice by orthotopic transplantation

目的建立人结直肠恶性淋巴瘤裸小鼠原位移植高转移模型,并探讨其生物学特性。方法将人原发性结直肠恶性淋巴瘤原发灶和肝转移灶术中新鲜瘤组织块植入裸小鼠结直肠黏膜层内,观察原位移植成瘤率、移植瘤的侵袭和转移率,并进行形态学(光镜、电镜、免疫组织化学)、染色体核型和流式细胞术分析。结果依据WHO新的恶性淋巴瘤分类法,从4例结直肠淋巴瘤标本中筛选出1株人结肠(非霍奇金B细胞)恶性淋巴瘤裸鼠原位移植肝转移模型(HCBL0301)和1株人直肠(非霍奇金B细胞)恶性淋巴瘤裸鼠原位移植高转移模型(HRBL0302)。移植瘤组织病理学均为非霍奇金(大B细胞性)高度恶性淋巴瘤,免疫组织化学示CD19、CD20、CD22、CD45阳性,CD3、CD7阴性。染色体众数范围55～69条。流式细胞术示DI值1.59～1.71,均为异倍体。HCBL0301和HRBL0302分别传至31代和27代,共移植裸鼠326只;自第3代起肿瘤的移植生长率和液氮冻存复苏成活率均为100%。HCBL0301多转移肝右叶,转移率为100%,淋巴结转移率和腹腔种植转移率为67.4%;HRBL0302多转移左右肝叶,转移率为63.7%,淋巴结转移率及腹腔种植转移率为56.4%。人结直肠恶性淋巴瘤在裸鼠结直肠内自主侵袭性生长,并发生血液转移、淋巴转移和腹腔内种植性转移。结论首次成功地建立人结直肠恶性淋巴瘤裸小鼠原位移植瘤模型HCBL0301和HRBL0302,并具有自发性高转移特点,可用于结直肠恶性淋巴瘤的基础及实验治疗研究。

Objective To reproduce high metastasis models of human primary colorectal lymphoma in nude mice by orthotopic transplantation, and to investigate their biologic features. Methods Histologically fresh lymphoma tissues from primary and liver metastatic lesions of human primary colorectal lymphoma obtained during operations were transplanted into colorectal mucosa of nude mice. Tumorgenecity, invasion, metastasis, morphology （light microscopy, electron microscopy and immunohistochemistry）, karyotype analysis and DNA content of the orthotopically transplanted tumors were studied. Results According to the new WHO classification of malignant lymphoma, a strain of liver metastasis model of human primary colonic lymphoma （non- Hodgkins, B cell） in nude mice by orthotopic transplantation （HCBL-0301）, and a strain of high metastasis model of human primary rectal lymphoma （non-Hodgkin＇s, B cell） in nude mice by orthotopic transplantation （HRBL-0302） were successfully screened from four cases of human primary colorectal lymphoma. Histopathology of transplanted tumors showed high grade non-Hodgkin＇s large B cell lymphoma. The cells were positive for CD19, CD20, CD22 and CD45, but negative for CD3 and CD7. The number of chromosome was between 55 and 69. DNA index （DI） was 1.59-1.71 G. e. heteroploid）. So far, HCBL0301 and HRBL-0302 had been passaged for 31 and 27 generations in nude mice respectively, and transplantation was successful in 326 nude mice. Since the third generation, both the growth rate of transplantation and rate of resuscitation after being thawed from liquid nitrogen cryopreservation were 100%. In HCBL-0301, metastasis to the right lobe of liver was most common and metastatic rate was 100%; additionally, rates metastasis to of lymph node and peritoneal seeding were 67. 4%. In HRBL-0302. metastasis to the left and right lobes of liver was most common with metastasis rate of 63. 7%, and rates of metastasis to lymph node and peritoneal seeding were 56. 4%. Transplanted human primary colorectal lymphoma could autonomously and invasively grow in the colorectum of nude mice, with occurrence of hematogenic, lymph node and implantation metastases. Conclusions The study successfully replicated high metastasis models of human primary eolorectal lymphoma in nude mice by orthotopic transplantation HCBL-0301 and HRBL-0302 moclels can be used in the research on pathogenesis, mechanism of invasion and metastasis and experimental therapy of human primary colorectal lymphoma.