The Journal of Practical Medicine >

2023 , Vol. 39 >Issue 19: 2456 - 2460

DOI: https://doi.org/10.3969/j.issn.1006-5725.2023.19.008

Basic Research

Construction of chronic myeloid leukemia K562 cell line with overexpression of human TCRP1 gene based on lentivirus vector and its biological function detection

  • Xiaorong LIU ,
  • Yue HE ,
  • Yan CHEN ,
  • Zefeng XIN ,
  • Xuwen DENG ,
  • Huifeng ZHONG ,
  • Yunsheng. CHEN
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  • *.Department of Clinical laboratory，Shenzhen Children′s Hospital，Shenzhen 518034，China
    *.Institute of Pediatrics，Shenzhen Children′s Hospital，Shenzhen 518034，China

Received date: 2023-06-09

  Online published: 2023-11-22

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Abstract

Objective To construct the chronic myeloid leukemia （CML） K562 cell line with overexpression of human tongue cancer drug resistance related gene （TCRP1） by lentivirus vector and to test its biological function. Methods The recombinant plasmid of TCRP1 and the packaging plasmid were co-transfected into 293T cells， and the K562 cells were infected after the titer of the virus was determined. Puromycin was used to screen TCRP1 overexpression cell line K562/TCRP1， and the expression of TCRP1 was detected by fluorescence quantitative PCR and Western blot. The proliferation of K562/TCRP1 and its control cell line was detected by continuous cell counting method and CCK-8 method respectively. The drug sensitivity of the two cell lines to different concentrations of imatinib （IM） was analyzed. Results The TCRP1 lentivirus expression vector was successfully transfected into K562 cells. The results of fluorescence quantitative PCR and Western blot showed that the expression of TCRP1 in K562/TCRP1 cell line was significantly higher than that in the control group at the level of mRNA and protein. The results of continuous cell counting and CCK-8 methods showed that the proliferation of K562/TCRP1 cells were enhanced， and the IC50 value of IM-treated K562/TCRP1 cells was significantly higher than that of the control cells （P < 0.05）. Conclusion The K562 cell line with overexpression of TCRP1 was successfully constructed using lentivirus vector， and we found that the overexpression of TCRP1 might enhance the proliferation and IM resistance of K562 cells， which provided a basis for further exploring the possible role of TCRP1 in the pathogenesis of chronic myeloid leukemia.

Key words： tongue cancer drug resistance related gene 1; overexpression; K562 cell line; chronic myelocytic leukemia

Cite this article

Xiaorong LIU , Yue HE , Yan CHEN , Zefeng XIN , Xuwen DENG , Huifeng ZHONG , Yunsheng. CHEN . Construction of chronic myeloid leukemia K562 cell line with overexpression of human TCRP1 gene based on lentivirus vector and its biological function detection[J]. The Journal of Practical Medicine, 2023 , 39(19) : 2456 -2460 . DOI: 10.3969/j.issn.1006-5725.2023.19.008

References

1 JABBOUR E， KANTARJIAN H. Chronic myeloid leukemia： 2022 update on diagnosis， therapy， and monitoring［J］. Am J Hematol， 2022， 97（9）：1236-1256.
2 MINCIACCHI V R， KUMAR R， KRAUSE D S. Chronic Myeloid Leukemia： A Model Disease of the Past， Present and Future［J］. Cells， 2021， 10（1）：117.
3 CORTES J， PAVLOVSKY C， SAUSSELE S. Chronic myeloid leukaemia［J］. Lancet， 2021， 398（10314）：1914-1926.
4 LIU H， JIA X， LUO K， et al. TCRP1 activated by mutant p53 promotes NSCLC proliferation via inhibiting FOXO3a［J］. Oncogenesis，2022， 11（1）：19.
5 WANG C， LIU H， QIU Q， et al. TCRP1 promotes NIH/3T3 cell transformation by over-activating PDK1 and AKT1［J］. Oncogenesis， 2017， 6（4）：e323.
6 FORD M， MAURO M， AFTANDILIAN C， et al. Management of Chronic Myeloid Leukemia in Children and Young Adults［J］. Curr Hematol Malig Rep， 2022， 17（5）：121-126.
7 ABDULMAWJOOD B， COSTA B， ROMA-RODRIGUES C， et al. Genetic Biomarkers in Chronic Myeloid Leukemia： What Have We Learned So Far？ ［J］ .Int J Mol Sci， 2021， 22（22）：12516.
8 SHANMUGANATHAN N， BRANFORD S. The Hidden Pathogenesis of CML： Is BCR-ABL1 the First Event？［J］. Curr Hematol Malig Rep， 2019， 14（6）：501-506.
9 LIU X， WANG C， GU Y， et al. TCRP1 contributes to cisplatin resistance by preventing Pol beta degradation in lung cancer cells［J］. Mol Cell Biochem， 2015， 398（1/2）：175-183.
10 GU Y， FAN S， LIU B， et al. TCRP1 promotes radioresistance of oral squamous cell carcinoma cells via Akt signal pathway［J］. Mol Cell Biochem， 2011， 357（1/2）：107-113.
11 刘孝荣，文飞球，马东礼，等.TCRP1在慢性髓系白血病细胞中的表达及其对细胞增殖的影响［J］. 中国医师杂志， 2017，19（5）：692-696.
12 LIN S C， HAGA K， ZENG X L， et al. Generation of CRISPR-Cas9-mediated genetic knockout human intestinal tissue-derived enteroid lines by lentivirus transduction and single-cell cloning［J］. Nat Protoc， 2022， 17（4）：1004-1027.
13 ALVES R， GONCALVES A C， RUTELLA S， et al. Resistance to Tyrosine Kinase Inhibitors in Chronic Myeloid Leukemia-From Molecular Mechanisms to Clinical Relevance［J］. Cancers （Basel）， 2021， 13（19）.4820.
14 ZHAO S， LI X， YIN L， et al. TCRP1 induces tamoxifen resistance by promoting the activation of SGK1 in MCF-7 cells［J］.Oncol Rep， 2020， 43（6）：2017-2027.
15 GU Y， ZHANG Z， YIN J， et al. Epigenetic silencing of miR-493 increases the resistance to cisplatin in lung cancer by targeting tongue cancer resistance-related protein 1（TCRP1） ［J］. J Exp Clin Cancer Res， 2017， 36（1）：114.
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