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KR-102963000-B1 - Scaffold loaded with adipose-derived stem cells overexpressing MFG-E8 for preventing or treating of liver fibrosis

KR102963000B1KR 102963000 B1KR102963000 B1KR 102963000B1KR-102963000-B1

Abstract

The present invention relates to a scaffold for the prevention or treatment of liver fibrosis, and more specifically, to a scaffold for the prevention or treatment of liver fibrosis loaded with adipose-derived stem cells overexpressing MFG-E8, prepared by transforming adipose-derived stem cells with a vector containing a gene encoding MFG-E8. Since the scaffold according to the present invention exhibits an excellent therapeutic effect on liver fibrosis by suppressing the expression of fibrosis-inducing factors related to liver fibrosis, it can be developed as an active ingredient in a composition for the prevention, treatment, or improvement of liver fibrosis.

Inventors

  • 김세준
  • 한재현
  • 김옥희

Assignees

  • 가톨릭대학교 산학협력단

Dates

Publication Date
20260511
Application Date
20211223

Claims (15)

  1. A scaffold for the prevention or treatment of liver fibrosis loaded with stem cells transformed by a vector containing a gene encoding MFG-E8 (milk fat globule-epidermal growth factor-factor VIII), The above stem cells are adipose-derived stem cells (ASCs), and The above scaffold is a scaffold for the prevention or treatment of liver fibrosis, characterized in that it is for epidermal grafting.
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  3. In paragraph 1, The above scaffold is a scaffold for the prevention or treatment of liver fibrosis, characterized by being composed of serum-derived components.
  4. In paragraph 1, A scaffold for the prevention or treatment of liver fibrosis, characterized in that the liver fibrosis is induced by liver toxins.
  5. In paragraph 4, A scaffold for preventing or treating liver fibrosis, characterized in that the above-mentioned liver toxin is a substance that causes damage or disease to liver tissue and is one or more selected from the group consisting of thioacetamide (TAA), carbon tetrachloride (CCl4), tBHP (tert-butyl hydroperoxide), acetaminophen, tacrine, rubratoxin B, and hydrogen peroxide ( H₂O₂ ).
  6. A pharmaceutical composition for the prevention or treatment of liver fibrosis comprising, as an active ingredient, a scaffold loaded with stem cells transformed with a vector containing a gene encoding MFG-E8, The above stem cells are adipose-derived stem cells (ASCs), and A pharmaceutical composition for the prevention or treatment of liver fibrosis, characterized in that the scaffold above is for epidermal grafting.
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  8. In paragraph 6, A pharmaceutical composition for the prevention or treatment of liver fibrosis, characterized in that the above scaffold is composed of serum-derived components.
  9. In paragraph 6, A pharmaceutical composition for the prevention or treatment of liver fibrosis, characterized in that the liver fibrosis is induced by liver toxins.
  10. In Paragraph 9, A pharmaceutical composition for preventing or treating liver fibrosis, characterized in that the above-mentioned liver toxin is a substance that causes damage or disease to liver tissue and is one or more selected from the group consisting of thioacetamide (TAA), carbon tetrachloride (CCl4), tBHP (tert-butyl hydroperoxide), acetaminophen, tacrine, rubratoxin B, and hydrogen peroxide ( H₂O₂ ).
  11. A step of transforming isolated stem cells with a vector containing a gene encoding MFG-E8; and A step of loading the above-mentioned transformed stem cells onto a scaffold; A method for manufacturing a scaffold for the prevention or treatment of liver fibrosis comprising, The above stem cells are adipose-derived stem cells (ASCs), and A method for manufacturing a scaffold for the prevention or treatment of liver fibrosis, characterized in that the above scaffold is for epidermal grafting.
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  13. In Paragraph 11, A method for manufacturing a scaffold for the prevention or treatment of liver fibrosis, characterized in that the above scaffold is composed of serum-derived components.
  14. In Paragraph 11, A method for manufacturing a scaffold for the prevention or treatment of liver fibrosis, characterized in that the liver fibrosis is induced by liver toxins.
  15. In Paragraph 14, A method for manufacturing a scaffold for preventing or treating liver fibrosis, characterized in that the above-mentioned liver toxin is a substance that causes damage or disease to liver tissue and is one or more selected from the group consisting of thioacetamide (TAA), carbon tetrachloride (CCl4), tBHP (tert-butyl hydroperoxide), acetaminophen, tacrine, rubratoxin B, and hydrogen peroxide ( H₂O₂ ).

Description

Scaffold loaded with adipose-derived stem cells overexpressing MFG-E8 for preventing or treating liver fibrosis The present invention relates to a scaffold for the prevention or treatment of liver fibrosis, and more specifically, to a scaffold for the prevention or treatment of liver fibrosis loaded with adipose-derived stem cells overexpressing MFG-E8, prepared by transforming adipose-derived stem cells with a vector containing a gene encoding MFG-E8. Liver fibrosis is characterized by the formation of thick fibrous septa in the liver due to the activation of hepatic stellate cells (HSCs) caused by repeated chronic liver disease, damage to liver tissue, and infiltration of inflammatory cells, which secrete collagen, the main substance of fibrosis, by the HSCs. Liver cirrhosis is a condition in which liver function declines as liver fibrosis progresses further. Causes of liver cirrhosis include HBV, HCV, alcoholic liver disease, and toxic liver disease, and if liver cirrhosis persists, it eventually leads to liver cancer. 30-40% of hepatitis B and 20-30% of hepatitis C cases progress to liver cirrhosis, and a significant number of these cases progress to liver cancer. Liver cirrhosis causes complications such as ascites, hepatitis, hepatic coma, and variceal bleeding, which drastically reduce the quality of life. Liver fibrosis/cirrhosis is known to be an irreversible reaction, but there have been recent reports suggesting that it may be reversible. Adipose-derived stem cells (ASCs) are one of the most useful mesenchymal stem cells (MSCs) and have many advantages, such as being easier to harvest than bone marrow, existing in large quantities, being widely available through various surgeries, having a yield rate about 1,000 times higher than bone marrow (2% vs 0.002%), and having a rapid proliferation rate. MFG-E8 (milk fat globule-epidermal growth factor-factor VIII), also known as lactaherin or SED1, is a protein secreted from milk fat globules that attaches to integrin-expressing cells and is known to 1) promote phagocytosis of apoptosis cells, 2) promote survival, proliferation, and angiogenesis of endothelial cells induced by VEGF, 3) promote migration of epithelial cells, and 4) inhibit fibrosis by eliminating inflammation and apoptosis cells. Three-dimensional cell culture technology is steadily advancing due to its superior biocompatibility compared to two-dimensional technology, and there are consistent reports indicating that two-dimensional and three-dimensional cell culture models exhibit different responsiveness to drugs. In particular, because cell-to-cell connections are established in all aspects within three-dimensional models, intercellular interaction is possible; these conditions are similar to physiological states, and the cells exhibit high differentiation potential due to their natural morphology, such as ellipsoids or polarized states. In the present invention, MFG-E8 was introduced into adipose-derived stem cells to transform them into adipose-derived stem cells overexpressing MFG-E8, and then loaded onto a scaffold very similar to the extracellular matrix (ECM) of liver tissue to study the therapeutic effect on liver fibrosis in a mouse model of liver fibrosis. As a result, it was confirmed that they possess excellent anti-fibrotic ability, thereby completing the present invention. Figure 1 shows the effect of adipose-derived stem cell culture medium transformed with MFG-E8 on cell viability in LX2 hepatic stellate cells (HSCs) activated with thioacetamide (TAA). (Ct, control; MCM, adipose-derived stem cell culture medium transformed with MFG-E8) Figure 2 shows the effect of MFG-E8-transformed adipose-derived stem cell culture medium on the expression of fibrosis-inducing factors in TAA-treated LX2 HSCs, confirmed by Realtime PCR. (Ct, control; NCM, adipose-derived stem cell culture medium; MCM, adipose-derived stem cell culture medium transformed with MFG-E8) Figure 3 shows the effect of MFG-E8-transformed adipose-derived stem cell culture medium on the expression of fibrosis-inducing factors in TAA-treated LX2 HSCs, confirmed by Western blot. (Ct, control; NCM, adipose-derived stem cell culture medium; MCM, adipose-derived stem cell culture medium transformed with MFG-E8) Figure 4 shows the changes in the expression of fibrosis-inducing factors at the mRNA level in mouse liver tissue obtained after implanting a scaffold loaded with adipose-derived stem cells transformed with MFG-E8 into a liver fibrosis model constructed by TAA treatment. (Ct, control; NCM, group loaded with adipose-derived stem cells on the scaffold; MCM, group loaded with adipose-derived stem cells transformed with MFG-E8 on the scaffold) Figure 5 shows the changes in the expression of fibrosis-inducing factors at the protein level, confirmed by Western blot, in mouse liver tissue obtained after implanting a scaffold loaded with adipose-derived stem cells transformed with MFG-E8 into a liver fibrosis model const