Method for inducing initial B cells to differentiate into regulatory B cells in in-vitro way and culture conditions thereof

A technology for inducing differentiation and B cells, applied in cell culture active agents, biochemical equipment and methods, tissue culture, etc., to achieve the effect of improving survival rate, good application prospects, and improving the ability to induce differentiation

Active Publication Date: 2018-09-28
BEIJING CHAOYANG HOSPITAL CAPITAL MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, no studies have reported how to induce CD19 + CD24 hi CD27 + Production of regulatory B cells

Method used

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  • Method for inducing initial B cells to differentiate into regulatory B cells in in-vitro way and culture conditions thereof
  • Method for inducing initial B cells to differentiate into regulatory B cells in in-vitro way and culture conditions thereof

Examples

Experimental program
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Embodiment 1

[0048] Example 1, a method for inducing naive B cells to differentiate into CD19 in vitro + CD24 hi CD27 + Medium for Regulatory B Cells

[0049] The method provided by the present invention for inducing naive B cells to differentiate into CD19 in vitro + CD24 hi CD27 + The medium for regulatory B cells is RPMI-1640 medium, fetal bovine serum, lipopolysaccharide (LPS), CD40 ligand (CD40L), interleukin 1β (IL-1β), interleukin 6 (IL-6 ), interleukin 4 (IL-4), interleukin 21 (IL-21) and interleukin 23 (IL-23) were mixed and obtained, wherein the volume fraction of fetal bovine serum in the culture medium was 10 %, the concentration of LPS in the medium is 10 μg / mL, the concentration of CD40L in the medium is 1 μg / mL, the concentration of IL-1β in the medium is 20 ng / mL, and the concentration of IL-6 in the medium is 20 ng / mL mL, the concentration of IL-4 in the medium is 20ng / mL, the concentration of IL-21 in the medium is 50ng / mL, and the concentration of IL-23 in the medi...

Embodiment 2

[0050] Example 2. Inducing naive B cells to differentiate into CD19 + CD24 hi CD27 + Regulatory B Cell Approach

[0051] 1. Induce naive B cells to differentiate into CD19 + CD24 hi CD27 + Regulatory B Cell Approach

[0052] 1. Patients diagnosed with pleural effusion shall collect 500 mL of pleural effusion through standard thoracentesis within 24 hours of admission, and add heparin anticoagulant to the pleural effusion sample. All samples were immediately transported to the laboratory in ice boxes. All patients with pleural effusion were from Beijing Chaoyang Hospital Affiliated to Capital Medical University, with informed consent.

[0053] 2. Centrifuge the pleural effusion sample collected in step 1 at 1500 rpm at 4°C for 10 minutes to obtain cell pellets.

[0054] 3. Add Ficoll-Hypaque gradient centrifugation (Pharmacia, Uppsala, Sweden) to the cell pellet obtained in step 2, and use gradient centrifugation to separate mononuclear cells. For specific steps, refer ...

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Abstract

The invention discloses a method for inducing initial B cells to differentiate into regulatory B cells in an in-vitro way and culture conditions thereof. The method comprises the following step of performing induced culture on the initial B cells in an inducing culture medium, wherein the inducing culture medium consists of an RPMI-1640 culture medium, fetal calf serum, lipopolysaccharide, a CD40ligand, interleukin 1 beta, interleukin 6, interleukin 4, interleukin 21 and interleukin 23. The method has the advantages that the initial B cells are cultured by the inducing culture medium, so thatthe enough number of regulatory B cells (CD19<+>CD24<hi>CD27<+>) is obtained; the ability of inducing the initial B cells to differentiate into the regulatory B cells (CD19<+>CD24<hi>CD27<+>) is obviously improved, the survival rate of the regulatory B cells (CD19<+>CD24<hi>CD27<+>) after inducing differentiating is increased, and the good application prospect is obtained.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a method for inducing naive B cells to differentiate into regulatory B cells in vitro and culture conditions thereof. Background technique [0002] Pleural effusion is a common clinical symptom characterized by pathological fluid accumulation in the pleural cavity. The pleural cavity is the potential gap between the visceral and parietal pleura. In normal people, there is 5-15 mL of fluid in the pleural cavity, which acts as a lubricant during respiratory movement. Pleural effusion occurs when fluid in the pleural space increases or decreases for any reason. There are a large number of immune cells in pleural effusion, such as lymphocytes, macrophages, granulocytes, and pleural mesothelial cells, among which B cells play an important immune role in pleural effusion. In the 1970s, scientists first described an immunosuppressive B cell, and many years later, the f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0781
CPCC12N5/0635C12N2501/2301C12N2501/2304C12N2501/2306C12N2501/2321C12N2501/2323
Inventor 施焕中王雯翟侃伊凤双吴秀芝
Owner BEIJING CHAOYANG HOSPITAL CAPITAL MEDICAL UNIV
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