人源化Ph染色体阳性急性淋巴细胞白血病小鼠模型的建立

Establishment of Mouse Model with Humanized Philadelphia Chromosome-positive Acute Lymphoblastic Leukemia

本研究旨在建立一种新型人源化Ph染色体阳性急性淋巴细胞白血病(Ph+ALL)小鼠异种移植模型。4-6周NOD/SCID小鼠经亚致死剂量60Co全身照射后,给予抗小鼠CD122单克隆抗体腹腔注射,在预处理后24 h内经小鼠膝关节骨髓腔注射Ph+ALL患者骨髓单个核细胞。于移植后8-12周通过流式细胞术检测受鼠骨髓和脾脏中人源细胞的植入水平及其免疫表型,应用实时定量聚合酶链式反应(RQ-PCR)和荧光原位杂交(FISH)检测受鼠骨髓和脾脏中人BCR/ABL1水平,并通过苏木精-伊红染色和抗人CD19,抗人CD34免疫组化染色评价人源Ph+ALL细胞在受鼠各组织器官中的迁移浸润能力。结果表明,在接受Ph+ALL患者细胞移植的受鼠骨髓和脾脏细胞中,人源Ph+ALL(huCD45+CD19+)细胞不仅有不同程度植入,而且植入细胞具有与Ph+ALL患者相似的细胞形态学、免疫表型和细胞遗传学特征。此外,人源Ph+ALL细胞还广泛迁移浸润到受鼠的脑、肝脏和肾脏等组织器官中。结论:抗CD122抗体预处理的NOD/SCID小鼠联合骨髓腔注射能够支持Ph+ALL患者骨髓单个核细胞的有效植入,为人类Ph+ALL白血病启动细胞鉴定及临床新药筛选研究提供一种新型异种移植模型。

The purpose of this study was to extablish a novel xenotransplant mouse model with human Philadelphia chromosome-positive acute lymphoblastic leukemia （Ph＋ ALL）.The bone marrow mononuclear cells （BMMNC） were separated from newly diagnosed Ph ＋ ALL patients,and injected into 2.1 Gy of 60Co irradiated and anti-CD122-conditioned NOD/SCID mice through intrafemoral injection.Human hematopoietic chimerism in bone marrow and spleen of the recipients was detected by flow cytometry.Morphological analysis of murine marrow cells were performed using May-Giemsa staining.BCR/ABL1 level was detected by RQ-PCR and FISH assays.Furthermore,leukemia infiltration in the organs was evaluated by hematoxylin-eosin staining,immunohistochemical staining with anti-human CD19 and anti-human CD34 antibodies.The results indicated that the unsorted BMMNC from Ph ＋ ALL patients were able to repopulate human Ph ＋ ALL in vivo.The percentages of human CD45 ＋ CD19 ＋ cells in bone warrow,and spleen of the recipient mice were 87.2％ ± 10.1％ and 79.9％ ± 9.2％,respectively.Furthermore,the engrafted cells possessed same morphology,phenotypic and cytogenetic characteristics as cells from the original Ph ＋ ALL patients.Compatible with the clinical features,transplanted Ph＋ ALL cells infiltrated into the brain,liver,and kidney of the recipients.It is concluded that the human-mouse xenotransplant established model using intrafemoral injection of an anti-CD122-conditioned NOD/SCID repopulation may provide a promising system to study the biology of human Ph ＋ ALL in vivo.