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Human primary myelofibrosis cell strain and application thereof

A technology of myelofibrosis and cell lines, which is applied in the fields of biology and oncology, can solve the problems of unestablished cell lines of PMF patients, and achieve the effect of shape stability

Active Publication Date: 2021-10-26
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, no cell lines derived from PMF patients have been established at home and abroad

Method used

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  • Human primary myelofibrosis cell strain and application thereof
  • Human primary myelofibrosis cell strain and application thereof
  • Human primary myelofibrosis cell strain and application thereof

Examples

Experimental program
Comparison scheme
Effect test

example 1ZY

[0027] Example 1 ZYXY-M2 cell line preparation

[0028] Primary cell culture: fresh high white blood cell isolation specimens obtained from the First Affiliated Hospital of Zhejiang University School of Medicine (male, 62 years old, PMF, white blood cell 178.1*10 9 / L) Immediate isolation of leukemic mononuclear cells. In a biosafety cabinet, take 6ml of the separation solution and add it dropwise to a 15ml sterile centrifuge tube previously added with 6ml of the lymphocyte separation solution, and centrifuge at 2000 rpm for 20 minutes. After centrifugation, take the mononuclear cell layer into a new 15ml sterile centrifuge tube, add 5ml sterile 1xPBS to resuspend the cells, and centrifuge at 2000 rpm for 5 minutes. After discarding the supernatant, add sterile erythrocyte lysate to lyse the cells at room temperature for 5 minutes, then centrifuge at 2000 rpm for 5 minutes. Discard the supernatant, add 5ml IMDM complete medium (IMDM 90% + fetal bovine serum 10%) to resuspend...

example 2

[0031] Biological properties and application of example 2 people's acute myeloid leukemia cell line

[0032] The invention adopts the IMDM medium containing 10% fetal bovine serum to cultivate the cell line, so that the cell line can grow stably in vitro and be passed down stably. Observed under the microscope, the cells are suspended or weakly attached to the wall, single or clustered, round or oval. Wright-Giemsa staining showed that the cells were mainly primitive erythrocytes, the cells were large, the cytoplasm was dark blue, some had a large number of vacuoles, there were light stained areas around the nucleus, the nuclear chromatin was fine and granular, and there were obvious nucleoli. The cell line did not express CD34 and CD11b by flow cytometry, but highly expressed CD71. Whole-exome sequencing of the cell line cells revealed that the cell line cells were negative for JAK2, CALR, and MPL mutations, and positive for TP53, ASXL1, and IKZF1 mutations in genes related ...

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Abstract

The invention discloses a human primary myelofibrosis cell strain as well as a construction method and application thereof. The primary myelofibrosis cell strain is named as human primary myelofibrosis cell strain ZYXY-M2, and was preserved in China Center for Type Culture Collection (Wuhan university, Wuhan, China) on January 20th, 2021 with the preservation number of CCTCC NO:C202145. The human primary myelofibrosis cell strain is obtained by extracting and separating peripheral blood of a clinical primary myelofibrosis patient to obtain mononuclear cells, and performing in-vitro culture and continuous natural passage. The leukemia cell strain JAK2, CALR and MPL is negative in mutation, ASXL1 is positive in mutation, TP53 is positive in mutation, IKZF1 is positive in mutation, IDH1 is positive in mutation, FLT3 is positive in mutation, and TET1 is positive in mutation. The human primary myelofibrosis cell strain has good in-vitro proliferation ability, can be used as a cell material for studying the development mechanism of human primary myelofibrosis and the in-vitro study of individualized treatment, and can be used for screening and evaluating human primary myelofibrosis drugs in vitro and in vivo and guiding clinical medication.

Description

technical field [0001] The invention relates to the fields of biology and oncology, in particular to a human myelofibrosis cell line and its construction method and application. Background technique [0002] Malignant tumors of the blood system originating from the myeloid system are divided into acute myeloid leukemia and its related tumors and chronic myeloid tumors according to the degree of differentiation, proportion, cell morphology, and molecular characteristics of the original cells at the time of onset. The most common chronic myeloid neoplasms are BCR-ABL fusion gene-positive chronic myeloid leukemia and BCR-ABL-negative myeloproliferative neoplasms (MPN). The most common MPNs are polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF). Studies have shown that the positive probabilities of JAK2, CALR, and MPL mutations in PV, ET, and PMF are: 95%, 0%, and 0% in PV patients, 60%, 20%, and 3% in ET patients, and 60%, 20%, and 3% in PMF...

Claims

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Application Information

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IPC IPC(8): C12N5/09C12Q1/02
CPCC12N5/0694G01N33/5011C12N2509/00C12N2503/02G01N2500/10G01N33/5047
Inventor 金洁李枫林周一乐俞文娟王敬瀚黄昕王云贵
Owner ZHEJIANG UNIV
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