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Hepatic stem cell preserving solution and applications of hepatic stem cell preserving solution

A technology of hepatic stem cells and human hepatic stem cells, applied in the field of hepatic stem cell preservation solution, can solve the problems of unclear mechanism of transdifferentiation of hepatic stem cells, and achieve the effect of non-toxic side effects, high safety, and repairing chronic liver damage.

Active Publication Date: 2014-09-03
北京清美联创干细胞科技有限公司
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  • Abstract
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  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although hepatic stem cells can form insulin-secreting cells through various pathways, the mechanism of transdifferentiation of hepatic stem cells into pancreatic cells is still unclear

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Embodiment 1, preparation of hepatic stem cell preservation solution

[0019] Human serum albumin injection was purchased from Harbin Shiheng Bioengineering Pharmaceutical Co., Ltd., the product catalog number is (92) Weiyao Zhunzi (Ha Shiheng) S-01, and each liter contains 20 g of effective dose of human serum albumin protein.

[0020] Heparin calcium injection was purchased from Sanofi Winthrop Industrie Company, the product catalog number is J20040119.

[0021] The preparation method of liver stem cell preservation solution I: 5mL human serum albumin injection (containing 0.1g effective dose of human serum albumin), 0.4mL heparin calcium injection, 2.60g sodium chloride, 2.48g sodium gluconate, 1.82 g sodium acetate, 0.18g potassium chloride and 0.15g magnesium chloride were dilute to 100mL with water.

[0022] The preparation method of liver stem cell preservation solution II: 10mL human serum albumin injection (containing 0.2g effective dose of human serum albumi...

Embodiment 2

[0025] Example 2, Cell Expansion of Human Hepatic Stem Cells

[0026] D-hank'S balanced salt buffer solution: the solvent is water, containing the following solutes: 0.8% NaCl, 0.04% KCl, 0.006% Na 2 HPO 4 ·H 2 O and 0.006% KH 2 PO 4 ; % are mass percentages.

[0027] 1. Wash 1 bottle of human liver stem cells twice with 10ml D-hank’S balanced salt buffer.

[0028] 2. Collect the cells and digest them with trypsin at 37°C for 6 minutes (5-7 minutes are acceptable); observe the cells under an inverted microscope during the digestion process, and gently pat the culture bottle with the palm of your hand to ensure that the human hepatic stem cells are completely digested.

[0029] 3. Stop the digestion, then add 10ml of D-hank’S balanced salt buffer solution to each bottle, and repeatedly pipette to suspend the cells to obtain a cell suspension.

[0030] 4. Count the viable cells of the cell suspension, the method is as follows: mix equal volumes of the cell suspension and 0...

Embodiment 3

[0032] Embodiment 3, the application of the hepatic stem cell preservation solution prepared in embodiment 1

[0033] The human hepatic stem cells in Example 2 were resuspended in the four kinds of hepatic stem cell preservation solutions prepared in Example 1, and the proportions were all 6.8×10 5 Human liver stem cell live cells: 1ml of liver stem cell preservation solution, stored at 4°C for 12 hours, then sampled (cell suspension) to measure cell survival rate.

[0034] Cell viability assay method: Mix 0.5ml of cell suspension with 0.5ml of 0.2% trypan blue solution to obtain a mixed solution; draw 10 microliters of the mixed solution and drop it on the edge of the cover slip to make the mixed solution fill the cover slip and counting plate Between; stand still for 1 minute; randomly select several fields of view, count 300 cells in total (count the number of colored cells, that is, the number of dead cells), cell survival rate=(300-number of dead cells) / 300×100%; repeat cou...

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Abstract

The invention discloses hepatic stem cell preserving solution and applications of the hepatic stem cell preserving solution. The hepatic stem cell preserving solution provided by the invention is prepared by fixing the volume of injection solution including 0.1 to 1g of human albumin, 2.60 to 4.97g of sodium chloride, 2.48 to 4.74g of sodium gluconate, 1.82 to 3.40g of sodium acetate, 0.18 to 0.35g of potassium chloride, 0.15 to 0.28g of magnesium chloride, and 0.4 to 0.7mL of heparin calcium based on effective dose by water to be reach to 100mL. The hepatic stem cell preserving solution provided by the invention is used for storing hepatic stem cells, and the survival rate of the hepatic stem cell is more than 85% and even more than 95% within 12 hours. The cell suspension which is obtained by floating the hepatic stem cells on the hepatic stem cell preserving solution can be used as the medicine for treating diabetes mellitus, and has the characteristics of being high in stability, good in curative effect, high in safety, being without toxic or side effect, convenient to store and transport, applicable to massive clinical use, and broad in application prospect. The hepatic stem cell preserving solution brings good news for diabetic patients, and provides a new way for clinical stem cell use.

Description

technical field [0001] The invention relates to a hepatic stem cell preservation solution and its application. Background technique [0002] "Diabetes" is a disease in which glucose in the blood tends to accumulate excessively. The alias given to it abroad is "Silent Killer", especially "adult type diabetes". , that is, there is one "diabetic" patient in ten people. Once suffering from "diabetes", life expectancy will be reduced by as much as ten years, and possible complications can spread throughout the body. "Diabetes" itself is also very painful. People often feel dry mouth and want to drink water, wake up many times in the middle of the night due to polyuria, feel hungry despite eating a lot of food, lose weight, lethargy, etc. Wait, it always makes people feel that there is something wrong with the whole body. By the time an obvious situation can be felt somewhere, the condition of "diabetes" has developed to a certain extent. And the terrible complications are al...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01N1/02
Inventor 田杰
Owner 北京清美联创干细胞科技有限公司
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