Preparation for inhibiting immunity and treating graft-versus-host diseases (GVHD) and preparation method of preparation

A graft-versus-host and preparation technology, which can be used in drug combinations, animal cells, pharmaceutical formulations, etc., can solve problems such as enhanced inhibition

Active Publication Date: 2012-08-22
嘉兴莱普晟医疗科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, the clinical application attempts of MSC are mainly focused on direct autologous or allogeneic cell transplantation. It has been reported that MSC has a dose-dependent regulatory effect on immune cells, especially the inhibitory effect on T cell proliferation. , the inhibitory effect is strengthened

Method used

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  • Preparation for inhibiting immunity and treating graft-versus-host diseases (GVHD) and preparation method of preparation
  • Preparation for inhibiting immunity and treating graft-versus-host diseases (GVHD) and preparation method of preparation
  • Preparation for inhibiting immunity and treating graft-versus-host diseases (GVHD) and preparation method of preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Example 1. Preparation of monocytes.

[0032] Add density gradient agent Histopaque-1077 (Sigma) to the extracted bone marrow or peripheral blood leukocyte suspension purchased directly from the blood bank, and add 30mL bone marrow extract or peripheral blood leukocyte suspension to every 15mL Histopaque. The bone marrow extract or peripheral blood leukocyte suspension was centrifuged at room temperature for 30 minutes at a speed of 400G in the presence of a gradient agent. After the layers are separated, the opaque layer in the middle is aspirated with a sterile disposable needle, which is the suspension rich in myeloid mononuclear cells.

Embodiment 2

[0033] Example 2. Acquisition of MSC cells.

[0034] The method for obtaining type I MSC cells: the bone marrow or peripheral blood mononuclear cells were added to DMEM culture medium with 10% human serum albumin or autologous serum at a rate of 1×10 per square centimeter. 6 The density of cells was cultured for 2 days, the suspended cells were removed, and the adherent cells were cultured for 21 days. The obtained type I MSCs are spindle-shaped and have typical MSC characteristics, that is, they express a large number of cell surface receptors CD90, CD105 and CD73, and do not contain CD14, CD34, CD45 and other hematopoietic stem cell and monocyte receptors.

[0035] The method for obtaining type II MSC cells: the bone marrow or peripheral blood mononuclear cells were added with 10% human serum albumin or autologous serum in DMEM culture medium at a rate of 1×10 per square centimeter. 6 The density of cells was cultured for 2 days, and magnetic bead sorting was carried out by...

Embodiment 3

[0037] Example 3. Obtaining preparations for suppressing immunity and treating graft-versus-host disease (GVHD).

[0038] The various types of MSCs obtained in Example 2 were placed in a culture medium without any growth factors, and were cultured in 0.9% medical saline for 2 days in an environment with an oxygen concentration of 0.5% (2 × 10 per square centimeter). 5 cells), and 1% medical human serum albumin or autologous serum can be added. After the culture was completed, the cell-free medium rich in cell growth factors was collected, and the adherent MSC cells were discarded. The collected cell-free medium was passed through a filter with a pore size of 0.2 microns to remove cell impurities and debris, and then stored in a -80°C freezer for later use. As the case may be, every 1x10 6 Each MSC cell can prepare 2-5 ml of the preparation for suppressing immunity and treating graft-versus-host disease (GVHD).

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Abstract

The invention relates to a preparation for inhibiting the immunity and treating the graft-versus-host diseases (GVHD) and a preparation method of the preparation. The preparation method of the preparation comprises the following steps that: mesenchymal stem cells (MSCs) are obtained through inducing monocytes, further, the MSCs are cultured under the low serum condition, finally, the MSCs are separated to obtain a cell-free culture medium, in addition, the cell-free culture medium is subjected to corresponding post treatment, and the preparation can be obtained. In vitro and in vivo experiments show that the preparation for inhibiting the immunity and treating the GVHD can obviously inhibit the immune reaction of receptors so that the GVHD symptoms are effectively relieved.

Description

technical field [0001] The invention relates to a preparation for suppressing immunity and treating graft-versus-host disease (GVHD) and a preparation method thereof. Background technique [0002] As early as the 19th century, in the field of wound healing research, it was first suggested that there might be non-hematopoietic stem cells in the bone marrow. However, it was not until 1976 that Mesenchymal stem cells (MSCs) were reported for the first time, and people began to pay attention to them gradually. MSC is a kind of stem cell with multi-directional differentiation potential existing in the bone marrow. When allogeneic or even heterogeneous mesenchymal stem cells are transplanted into the host, they can differentiate into mesoderm, ectoderm or mesoderm under certain induction conditions. Cells that characterize the endoderm and participate in host organization. This has made a huge impact on people's understanding of allogeneic transplantation. Even in some studies, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/14A61K35/28A61K35/12C12N5/0775A61P37/06A61K35/35
Inventor 顾丽娅杨子江顾茂健
Owner 嘉兴莱普晟医疗科技有限公司
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