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A kind of human primary myelofibrosis cell line and its application

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

Active Publication Date: 2022-03-25
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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  • A kind of human primary myelofibrosis cell line and its application
  • A kind of human primary myelofibrosis cell line and its application
  • A kind of human primary myelofibrosis cell line and its application

Examples

Experimental program
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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 line and its construction method and application. The primary myelofibrosis cell line is named human primary myelofibrosis cell line ZYXY‑M2, and was deposited in the China Center for Type Culture Collection (Wuhan University, Wuhan, China) on January 20, 2021, with the preservation number CCTCC NO :C202145. The present invention is obtained by extracting and separating mononuclear cells from the peripheral blood of patients with clinical primary myelofibrosis, and culturing them in vitro for continuous natural passage. The leukemia cell line is negative for JAK2, CALR, and MPL mutations, positive for ASXL1 mutations, positive for TP53 mutations, positive for IKZF1 mutations, positive for IDH1 mutations, positive for FLT3 mutations, and positive for TET1 mutations. With good in vitro proliferation ability, it can be used as a cell material for studying the mechanism of human primary myelofibrosis and individualized treatment in vitro. It can also be used for in vivo and in vitro research of drug screening, evaluation, and guidance for human primary myelofibrosis. 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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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/09C12Q1/02
CPCC12N5/0694G01N33/5011C12N2509/00C12N2503/02G01N2500/10
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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