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A Detailed Protocol for Characterizing the Murine C1498 Cell Line and its Associated Leukemia Mouse Model
Published on: October 14, 2016
[Establishment and characterization of a new human myeloid leukemia cell line SH-2]
Hui-Ying Qiu1, Yong-Quan Xue, Jun Zhang
1Department of Hematology, The First Affiliated Hospital of Soochow University, Jiangsu Institute of Hematology, Suzhou 215006, China.
Objective:
To establish and characterize a novel human myeloid leukemia cell line SH-2.
Methods:
Bone marrow mononuclear cells (BMMNC) isolated from a AML-M2 patient, who failed to obtain complete remission after chemotherapy and allogenic bone marrow transplantation were passed in a long term IMDM culture medium supplemented with 20% fetal calf serum. Stromal cells were retained and rh-IL-3 was added in the culture system. A new human myeloid leukemia cell line SH-2 was successfully established with a cytogenetic characteristics of a loss of Y chromosome (-Y), a derivative chromosome 16 resulting from unbalanced translocation between chromosome 16 and 17, monosome 17, trisomy 19 and p53 alteration. Various methods were employed to characterize SH-2 cell line.
Results:
SH-2 cells has been maintained without cytokine and stromal cells for more than 3 years without EB virus and mycoplasma contamination. SH-2 cells had the basically same morphological, immunophenotypic and cytogenetic features as the patient's leukemia cells did, such as myeloid morphology, an immunophenotype of CD13+, CD33+, CD56+, CD16/56+ and a hypodiploid karyotype of 45, X, -Y, der(16)t(16;17)(q24;ql2), -17, +19, which were gradually decreased and replaced by the near-tetraploid cells with a karyotype of 73-102(80), XX, -Y, -Y, del (q131)x2, der(16)t(16;17)(q24;q12)x2, -17, -17, +19, +19. FISH and multiple FISH delineated all the abnormalities and revealed a loss of one p53 allele due to monosomy 17. DNA direct sequencing detected a point mutation of CAG to CAT at codon 576 of exon 5 in another p53 allele. RT-PCR showed that SH-2 cells expressed apoptosis-related genes (bcl-2, Fas, GST-pi and p2) rather than MDR-related genes. Short tandem repeat PCR provided powerful evidence for the derivation of SH-2 cell line from the patient's leukemia cells. SH-2 cells had certain colony formation and tumorigenic capacities in nude and SCID mice.
Conclusion:
SH-2 is a new myeloid leukemia cell line with a unique biology background, and will provide a useful tool for leukemia research.
