Decellularized nerve graft and method of manufacturing the same
The present invention relates to a decellularized nerve graft using allogeneic and heterologous nervous tissues and a method of manufacturing the same. In the present invention, by using a low-concentration basic solution and a surfactant as a decellularization solution, cell and tissue toxicity caused by a solvent or surfactant remaining in the tissue may be minimized by minimizing the use of a basic solution and an anionic surfactant in the entire manufacturing process. In addition, a peristaltic pump may be used to maintain the tissue structure and effectively remove lipid and cells.
1 . A method of manufacturing a decellularized nerve graft, comprising:
providing a nervous tissue;
a) removing a lipid component of the nervous tissue, thereby providing lipid component- removed nervous tissue; and
b) removing cells from the lipid component-removed nervous tissue, wherein:
b) is performed by treating the lipid component-removed nervous tissue with a basic solution that comprises one or more bases and is free of an anionic surfactant, and then with a separate anionic surfactant solution that comprises one or more anionic surfactants and is free of the one or more bases;
in b), the time for treatment with the basic solution is 30 minutes to 9 hours, and the time for treatment with the anionic surfactant solution is 30 minutes to 9 hours; and
a) and b) are performed using a peristaltic pump system operated at 50 to 300 rpm (revolutions per minute);
the one or more bases is selected from the group consisting of sodium hydroxide, potassium hydroxide, ammonium hydroxide, calcium carbonate, magnesium hydroxide, calcium hydroxide, and ammonia;
the one or more anionic surfactants is selected from the group consisting of sodium deoxycholate (SDC), sodium dodecyl sulfate (SDS), alkylbenzene sulfonate (ALS), alcohol ether sulfates (AES), sodium lauryl sulfate (SLS), and polyethylene glycol (PEG); and
the concentration of the one or more bases in the basic solution is 0.1 to 8N (normal concentration).
2 . The method of claim 1 , wherein, a) is performed using a delipidation solution,
wherein the delipidation solution includes a polar solvent, a non-polar solvent, or a mixed solvent thereof.
3 . The method of claim 1 , wherein the one or more bases is sodium hydroxide, and
the one or more anionic surfactants is sodium deoxycholate (SDC).
4 . The method of claim 1 , wherein the concentration of the one or more bases in the basic solution is 0.1 to 1 N (normal concentration).
5 . The method of claim 1 , wherein the peristaltic pump system comprises
a peristaltic pump;
a chamber connected with the peristaltic pump that contains the nervous tissue;
a reservoir connected with the peristaltic pump that stores a treatment liquid for treating the nervous tissue; and
a waste water storage bin connected with the chamber where the treatment liquid that has been used for treating the nervous tissue is discharged.
6 . The method of claim 5 , wherein the treatment liquid used for treating the nervous tissue is recycled and reused.
7 . The method of claim 1 , wherein the method further comprises:
washing the decellularized nerve graft, wherein
the washing is performed in the peristaltic pump system.
8 . The method of claim 1 , wherein the method further comprises:
lyophilizing and sterilizing the decellularized nerve graft.
9 . A decellularized nerve graft manufactured by the preparation method of claim 1 , wherein the decellularized nerve graft has a uniform pore size of 10 to 20 μm.