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Method for preparing porcine hepatocyte and mesenchymal stem cell co-microencapsulated internal bio-artificial liver

A bone marrow mesenchymal, bioartificial technology, applied in animal cells, vertebrate cells, artificial cell constructs, etc., can solve the problem that the built-in cells cannot survive for a long time, affect the graft's support for the host's liver function, and have limited proliferation ability. To maintain biological function, improve treatment effect, and prolong survival time

Inactive Publication Date: 2011-10-19
THE AFFILIATED DRUM TOWER HOSPITAL MEDICAL SCHOOL OF NANJING UNIV
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Problems solved by technology

Especially after entering the liver failure patients with homeostasis disorder, the built-in cells usually cannot survive for a long time, and their proliferation ability is extremely limited, which seriously affects the support of the graft to the host liver function

Method used

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  • Method for preparing porcine hepatocyte and mesenchymal stem cell co-microencapsulated internal bio-artificial liver
  • Method for preparing porcine hepatocyte and mesenchymal stem cell co-microencapsulated internal bio-artificial liver

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Experimental program
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Embodiment Construction

[0022] Establishment of porcine bone marrow mesenchymal stem cells in vitro culture and expansion system:

[0023] Extract about 5ml of bone marrow tissue from the ilium of miniature pigs used in adult health experiments, transfer it into a centrifuge tube, blow and mix it, and then slowly add it into the centrifuge tube containing 5ml of lymphocyte separation solution along the tube wall according to the volume ratio of 1:1, and use a centrifuge After centrifugation, carefully draw the middle cloudy cell layer, wash with phosphate buffer, add DMEM medium containing 10% fetal bovine serum, and then place it at 37°C, 5% CO 2 cultured in an incubator. The non-adherent cells were discarded by the monolayer cell adherent culture method, digested with trypsin and subcultured, and the growth of the cells was observed.

[0024] Isolation and purification of primary porcine hepatocytes:

[0025] Porcine hepatocytes were isolated by two-step collagenase in situ perfusion. After the ...

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Abstract

The invention relates to a method for preparing a porcine hepatocyte and mesenchymal stem cell co-microencapsulated internal bio-artificial liver. The invention comprises the following steps: establishing an in-vitro culture amplification system for porcine mesenchymal stem cells, performing in-situ perfusion separation of porcine primary hepatocytes by a two-step collagenase method, and preparing alginate-poly-L-lysine microencapsulated hepatocytes and mesenchymal stem cells. The invention overcomes the defects that internal cells usually can not survive for a long time; the proliferation ability is quite limited; and the function of the injected hepatocytes for generating various stimulating factors for promoting liver regeneration is inhibited. The invention introduces mesenchymal stem cells into an internal bio-artificial liver system, fully simulates the in-vivo microenvironment of hepatocytes by heterogeneous intercellular interaction, effectively prolongs the survival time of hepatocytes, promotes the proliferation and differentiation of hepatocytes, maintains the specific biological functions of hepatocytes, and thus significantly improves the efficacy of the internal bio-artificial liver on treating hepatic failure.

Description

technical field [0001] The invention relates to a built-in bioartificial liver support system, in particular to a method for co-microencapsulating pig liver cells and bone marrow mesenchymal stem cells into a built-in bioartificial liver. Background technique [0002] Bioartificial livers mainly include external and internal bioartificial livers. The basic principle is to place the hepatocytes cultured and proliferated in vitro in a circulation device or directly in the body, and the patient's blood / body fluid exchange substances with the cultured hepatocytes through the biofiber semipermeable membrane, so as to achieve the ideal artificial liver support function. Hepatocytes are the core material of the bioartificial liver, and the supporting effect of the bioartificial liver on patients with liver failure depends on the function of the hepatocytes used. At present, primary porcine hepatocytes are widely used in bioartificial liver research because of their anatomical stru...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61L27/38C12N5/071C12N5/0775A61M1/34
Inventor 施晓雷丁义涛顾劲扬
Owner THE AFFILIATED DRUM TOWER HOSPITAL MEDICAL SCHOOL OF NANJING UNIV
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