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Stem cell preparation for treating primary liver cancer and preparation method thereof

A technology of stem cell preparation and primary liver cancer, which is applied to medical preparations containing active ingredients, medical raw materials derived from mammals, and pharmaceutical formulas, etc., can solve the problems of immunogenicity and limited quantity easily infected by viruses, and achieve Effects of normal karyotype and telomerase activity, high activity, and strong proliferative ability

Active Publication Date: 2014-06-18
奥思达干细胞有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, mesenchymal stem cells mainly come from bone marrow and fat, but the number of mesenchymal stem cells in bone marrow and fat is limited, which limits its clinical application because of its susceptibility to virus infection and strong immunogenicity.

Method used

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  • Stem cell preparation for treating primary liver cancer and preparation method thereof
  • Stem cell preparation for treating primary liver cancer and preparation method thereof
  • Stem cell preparation for treating primary liver cancer and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1: Preparation of human umbilical cord mesenchymal stem cell preparation

[0037]The umbilical cord that passed the test was fully washed with D-hank's balanced salt solution, the residual blood in the umbilical vein and umbilical artery was washed away, the adventitial tissue and vascular tissue of the umbilical cord were separated and removed, and cut into pieces of about 1 cubic millimeter , transferred to collagenase type II with a mass fraction of 0.1%, digested at 37°C for 20 hours, passed through a 100-mesh sieve, collected the filtrate containing cells, centrifuged at 1200 rpm for 10 minutes at room temperature, discarded the supernatant, and retained the precipitate; The cells were inoculated in culture flasks containing 10% fetal bovine serum and DMEM / F12 medium containing 100u / mL double-antibody (penicillin, streptomycin), and the culture flasks were placed at 37°C with a volume fraction of 5% carbon dioxide. Cultivate in a saturated humidity envir...

Embodiment 2

[0047] Embodiment 2: the preparation of PRP lysate

[0048] The whole process of making PRP lysate is strictly aseptic. Take 10ml of venous blood from the patient, centrifuge it at 200g for 20min, and collect the supernatant, which is platelet-rich plasma. Platelet Rich Plasma Pass-80 0 C refrigerator and 37 0 C water bath quick freezing - thawing, the supernatant after centrifugation is the platelet-rich plasma lysate.

Embodiment 3

[0049] Example 3: Detection of hepatocyte growth factor and fibroblast growth factor-4 to promote the differentiation of umbilical cord mesenchymal stem cells into hepatocyte-like cells

[0050] 1. Cell culture: Inoculate umbilical cord mesenchymal stem cells in a culture dish containing 10% fetal bovine serum and 100u / mL double-antibody (penicillin, streptomycin) in DMEM / F12 culture medium, change the medium every other day, and wait for the cells to fuse Realize when the speed reaches 50%;

[0051] 2. Experimental grouping: Group A (negative control group): no induction factor; Group B: HGF (20 ng / ml); Group C: FGF-4 (10 ng / m1); Group D: HGF (20 ng / ml). m1)+FGF-4 (10 ng / m1); E group (positive control group): L-02 human liver cell line;

[0052] 3. RT-PCR detection: RT-PCR detection of AFP and ALb mRNA was carried out in each group on the 14th and 21st day of induction, and groups A and E were respectively negative and positive control groups. Expression of AFP: AFP Sense 5...

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Abstract

The invention discloses a stem cell preparation for treating primary liver cancer and a preparation method thereof. The preparation contains (60-100)*10<6> pieces / ml of mesenchymal stem cells, 20ng / ml of HGF and 10ng / ml of FGF-4, and takes autologous platelet-rich plasma lysate as a solvent. The preparation method comprises the following steps of: (1) collecting an umbilical cord, and carrying out primary culture to obtain cells; (2) detecting by using a flow cytometry, wherein such cells express CD29, CD73, CD90 and CD105; (3) collecting cells, namely dissolving (60-100)*10<6> mesenchymal stem cells of the human umbilical cord in 1 ml of autologous platelet-rich plasma lysate, and adding 20ng of HGF and 10ng of FGF-4 to be prepared into 1 ml of a stem cell preparation. According to the stem cell preparation disclosed by the invention, the source of raw materials is rich, the raw materials are easy to obtain, the umbilical cord mesenchymal stem cells are weakly immunogenic, and platelet-rich plasma is taken from the patients, so no immunogenicity exists.

Description

technical field [0001] The invention relates to a cell preparation for treating primary liver cancer and a preparation method thereof. The cells in this cell preparation are derived from mesenchymal stem cells in the umbilical cord, supplemented with hepatocyte growth factor and fibroblast growth factor-4, and the cells are suspended in 1 ml of autologous platelet-rich plasma lysate for the treatment of primary liver cancer caused liver cell damage. Background technique [0002] Primary liver cancer (hepatocellular carcinoma, HCC) ranks fifth in the incidence of malignant tumors, and the global annual death toll is about 1 million. Currently, most patients with primary liver cancer lack effective treatments, and the overall survival rate is only 3% to 5%. The occurrence of primary liver cancer is the result of long-term effects of multiple factors, multiple pathways, and multiple steps, including external environmental carcinogenic factors and self-genetic factors. It has...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/18A61K35/44A61P35/00A61P1/16A61K35/51
Inventor 周萱
Owner 奥思达干细胞有限公司
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