Recombinant mesenchymal stem cell and application thereof

A mesenchymal stem cell and cell technology, applied in the field of recombinant mesenchymal stem cells and its application, can solve the problem of lack of effective killing of tumor cells

Inactive Publication Date: 2019-04-09
河北生命原点生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Aiming at the fact that existing MSCs cells can directional migrate to the tumor occurrence site of the body but lack the ability to effectively kill tumor cells, the present invention provides a recombinant mesenchymal stem cell and its application

Method used

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  • Recombinant mesenchymal stem cell and application thereof
  • Recombinant mesenchymal stem cell and application thereof
  • Recombinant mesenchymal stem cell and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] A recombinant mesenchymal stem cell that integrates the cDNA expressing interleukin-21 into the genome of the mesenchymal stem cell. Specifically:

[0033] 1) Obtain cDNA of IL-21: Collect 400ml of adult male peripheral blood, use NK cell separation medium (produced by Tianjin Haoyang Company, NK2011H) to obtain NK cells by density gradient centrifugation, add Trizol reagent to lyse, then add chloroform to extract RNA, centrifuge Obtain the supernatant of total RNA containing natural killer cells, add isopropanol to centrifuge the RNA, remove the supernatant, obtain the total RNA of natural killer cells, and reverse transcribe to obtain cDNA of natural killer cells. IL-21 is secreted by natural killer cells, so the cDNA obtained by reverse transcription of natural killer cell RNA contains a large amount of IL-21 cDNA. The reverse transcription system is shown in the table below:

[0034]

[0035] Prepare the reverse transcription system on ice according to the abov...

Embodiment 2

[0056] The hUC-MSCs, Vector / hUC-MSCs, and IL-21 / hUC-MSCs obtained in Example 1 were used to conduct tumor killing experiments in vitro. Among them, Vector / hUC-MSCs are hUC-MSCs infected with empty lentivirus; IL-21 / hUC-MSCs are hUC-MSCs infected with lentivirus carrying IL-21 expression sequence.

[0057] The test method is as follows: using human SMMC-7721 liver cancer cells as target cells, three kinds of tumor-killing cells, hUC-MSCs, Vector / hUC-MSCs, and IL-21 / hUC-MSCs, were mixed with 5x10 4 SMMC-7721 liver cancer cells were co-cultured, and the number of hUC-MSCs, Vector / hUC-MSCs, and IL-21 / hUC-MSCs was 2x10 4 After 72 hours, the proliferation of liver cancer cells in each group was detected by CCK8 method.

[0058] Test results such as Figure 5 As shown, the ordinate represents the OD value of liver cancer cells. Compared with the blank control group without tumor killer cells, hUC-MSCs, Vector / hUC-MSCs, IL-21 / hUC-MSCs can significantly inhibit SMMC-7721 liver cancer...

Embodiment 3

[0060] The hUC-MSCs, Vector / hUC-MSCs, and IL-21 / hUC-MSCs obtained in Example 1 were used to carry out tumor killing experiments in vivo.

[0061] The test method is as follows: construct a nude mouse model with SMMC-7721 subcutaneous tumor transplantation, and inject 200 μl of normal saline (Saline), hUC-MSCs, Vector / hUC-MSCs, and Vector / hUC- MSCs, IL-21 / hUC-MSCs, anti-tumor experiments, infusion twice a week, continuous treatment for 3 weeks, in which three kinds of cells hUC-MSCs, Vector / hUC-MSCs and IL-21 / hUC-MSCs were resuspended In 200μl of normal saline, inject into the mouse body, the amount of each injection is 1×10 6 indivual.

[0062] Antitumor results such as Figure 6 Shown: Compared with the normal saline (Saline) group, the tumor volume growth of the hUC-MSCs, Vector / hUC-MSCs and IL-21 / hUC-MSCs groups was significantly slowed down, while the IL-21 / hUC-MSCs group was significantly slower than the hUC-MSCs group. The growth rate of tumor in MSCs and Vector / hUC-M...

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Abstract

The invention relates to the technical field of tumor treatment, specifically to recombinant mesenchymal stem cell and application thereof. The recombinant mesenchymal stem cell belongs to mesenchymalstem cell used for killing tumor formed by integrating cDNA for expressing interleukin-21 to gene of mesenchymal stem cell, which can perform effective positioning the position of a tumor, can perform high-level expression of interleukin-21, can rapidly activate immune cell through secreted interleukin-2 after accurately positioning the position of the tumor, rapidly and effectively inhibit and kill tumor cell, and has very important research significance in the treatment field of tumor and cancer.

Description

technical field [0001] The invention relates to the technical field of tumor treatment, in particular to a recombinant mesenchymal stem cell and its application. Background technique [0002] Tumor is a disease that seriously threatens human health today. In recent years, it ranks first in the global statistics. The incidence of tumor is also showing a rising trend year by year. After tumor deterioration, it can develop into cancer. Globally, more than 500,000 patients die from liver cancer every year. [0003] Up to now, surgery is still the most effective means of treating tumors, but this is only for patients with benign tumors. In the actual clinical diagnosis and treatment process, some tumor patients have developed into malignant tumors after being confirmed. For example, the diagnosed patients with liver cancer have often advanced to the middle and late stage, and only 10%-30% of the patients can undergo radical resection at this time, but the five-year survival rate...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12N15/867C12N15/66A61K35/28A61P35/00
CPCA61K35/28A61P35/00C07K14/54C12N15/66C12N15/86C12N2740/15043
Inventor 李陶姚辉袁洁
Owner 河北生命原点生物科技有限公司
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