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B-lymphoblastoid model induced by lactic acid to differentiate epithelioid cells and its construction and application

A technology of B lymphoblastoid cell differentiation, applied in the field of medical bioengineering, to achieve the effect of enhancing cell migration ability

Inactive Publication Date: 2018-10-12
SHANGHAI DERMATOLOGY HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although Epstein-Barr virus transformation technology is available to prepare immortalized B lymphoblasts, there is no report about a kind of immortalized epithelioid cell-differentiated B lymphoblasts

Method used

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  • B-lymphoblastoid model induced by lactic acid to differentiate epithelioid cells and its construction and application
  • B-lymphoblastoid model induced by lactic acid to differentiate epithelioid cells and its construction and application
  • B-lymphoblastoid model induced by lactic acid to differentiate epithelioid cells and its construction and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] 1. Isolation of B cells from peripheral blood:

[0037] The collected peripheral blood was diluted 1:1 with Hank’s solution / PBS solution / serum-free RPMI1640, and the lymphocyte separation solution stored at 4°C in the dark was taken out of the refrigerator. Centrifuge at 2000rpm for 20min at room temperature. Gently pass the pipette through the plasma layer to the buffy coat layer, suck out the buffy coat layer cells between the plasma layer and the separation liquid layer along the periphery of the centrifuge tube, and place them in a new centrifuge tube. Aspirate as little separation fluid as possible. Add RPMI1640, take a sample and count the cells after pipetting evenly. Peripheral blood mononuclear cells were sorted by CD20 magnetic beads to obtain B lymphocytes.

[0038] 2. Preparation of EB virus particles

[0039] Use B95.8 cells to culture Epstein-Barr virus: B95.8 cells were suspended in 10% fetal bovine serum RPMI1640 medium, and the cell concentration wa...

Embodiment 2

[0060] Example 2: Screening for the appropriate concentration of lactic acid induction

[0061] experimental method:

[0062] 1. Separation of B cells in peripheral blood, same as in Example 1

[0063] 2. The preparation of EB virus particles is the same as in Example 1

[0064] 3. Utilize Epstein-Barr virus transformation technology to transform lymphoblastoid cells, with embodiment 1

[0065] 4. Apply different lactic acid concentrations for screening, and the concentrations are respectively set to 0mM, 5mM, 10mM, 20mM, 30mM, 60mM and 80mM.

[0066] (1) MTT method was used to detect the effects of different concentrations of lactic acid on the proliferation of LCL.

[0067] Cells were seeded in 96-well plates (5×10 4 each hole), add different concentrations of lactic acid on the next day, set blank control group and zero adjustment group as the control group, discard the supernatant after incubation for 24 hours, wash twice with PBS, add 100 microliters of lactic acid-fr...

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Abstract

The invention relates to the field of medical bioengineering, and in particular relates to a B lymphocytoblast model realizing differentiation of epithelioid cells through lactic acid induction as well as a preparation method and an application of the B lymphocytoblast model. According to the B lymphocytoblast model, the normal B cells are converted into the immortalized LCL through EB viruses, then the immortalized B cells are cultured through application of lactic acid induction, and thus the cells realizing differentiation of epithelioid cells and being capable of secreting large amount of extracellular matrix are obtained; the migration capacity of the cells is enhanced, and a study tool is provided for researching the malignant evolution of B cell lymphomas.

Description

technical field [0001] The invention relates to the technical field of medical bioengineering, in particular, it is a B lymphoblastoid model differentiated from immortalized epithelioid cells, which can be used for basic research on B lymphoma, especially B lymphoma in an acidic microenvironment Mechanisms of deterioration and metastasis. Background technique [0002] According to different clinical behaviors, B-cell lymphoma is divided into indolent lymphoma and aggressive lymphoma. Indolent lymphoma usually develops slowly and can maintain stable disease and long-term survival for many years. Aggressive lymphoma has a higher degree of malignancy and usually requires more intensive treatment. Indolent lymphomas can evolve into aggressive lymphomas, but the mechanism is unclear. The lactic acid produced by tumor cells through efficient glycolysis forms an extracellular acidic microenvironment that is not suitable for the survival of normal cells, and promotes the prolifera...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/869C12N5/10C12Q1/02
Inventor 莫小辉谭飞
Owner SHANGHAI DERMATOLOGY HOSPITAL
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