Method of preparing an adipose tissue derived matrix
An acellular soft tissue-derived matrix includes a collagenous tissue that has been delipidated and decellularized. Adipose tissue is among the soft tissues suitable for manufacturing an acellular soft tissue-derived matrix. Exogenous tissuegenic cells and other biologically-active factors may be added to the acellular matrix. The acellular matrix may be provided as particles, a slurry, a paste, a gel, or in some other form. The acellular matrix may be provided as a three-dimensional scaffold that has been reconstituted from particles of the three-dimensional tissue. The three-dimensional scaffold may have the shape of an anatomical feature and serve as a template for tissue repair or replacement. A method of making an acellular soft tissue-derived matrix includes steps of removing lipid from the soft tissue by solvent extraction and chemical decellularization of the soft tissue.
1. A method of preparing an adipose tissue-derived matrix, comprising the steps of:
(a) obtaining an adipose tissue;
(b) mechanically reducing the size of the adipose tissue;
(c) delipidating the adipose tissue by contacting the adipose tissue with a polar organic solvent such that lipids are transferred from the adipose tissue to the polar organic solvent, thereby producing a delipidated adipose tissue from which substantially all of the lipids native to the adipose tissue have been removed;
(d) decellularizing the delipidated adipose tissue after step (c), thereby producing a delipidated, decellularized adipose tissue which is essentially free of native cells and cellular components.
2. The method claim 1 , comprising the further step of separating the adipose tissue from lipids released from the adipose tissue during the mechanically reducing step (b).
3. The method of claim 1 , wherein said step of mechanically reducing the size of the adipose tissue includes grinding the adipose tissue with a coarse grinding plate, then grinding the ground adipose tissue with a fine grinding plate.
4. The method of claim 1 , wherein said delipidating step includes washing the adipose tissue with a polar organic solvent, wherein the polar solvent includes a substituted hydrocarbon having a number of carbon atoms in the range of two carbon atoms to six carbon atoms, and said substituted hydrocarbon is selected from the group consisting of a chlorinated hydrocarbon, a fluorinated hydrocarbon, an alcohol, an ether, a ketone, an aldehyde, an ester, an organic acid, and combinations thereof.
5. The method of claim 1 , wherein said delipidating step includes blending the adipose tissue with a polar organic solvent, Wherein the polar solvent includes a substituted hydrocarbon having a number of carbon atoms in the range of two carbon atoms to six carbon atoms, and said substituted hydrocarbon is selected from the group consisting of a chlorinated hydrocarbon, a fluorinated hydrocarbon, an alcohol, an ether, a ketone, an aldehyde, an ester, an organic acid, and combinations thereof.
6. The method of claim 1 , wherein said delipidating step includes homogenizing the adipose tissue with a polar organic solvent, wherein the polar solvent includes a substituted hydrocarbon having a number of carbon atoms in the range of two carbon atoms to six carbon atoms, and said substituted hydrocarbon is selected from the group consisting of a chlorinated hydrocarbon, a fluorinated hydrocarbon, an alcohol, an ether, a ketone, an aldehyde, an ester, an organic acid, and combinations thereof.
7. The method of claim 1 , wherein said decellularizing step includes contacting the delipidated adipose tissue with a detergent solution, wherein said detergent is present in said solution at a concentration in a range of from about 0.1% to about 5.0% (w/v).
8. The method of claim 7 , wherein the detergent solution includes a deoxycholate salt.
9. The method of claim 7 , wherein the detergent solution includes a mixture of water and alcohol.
10. The method of claim 9 , wherein the alcohol is present in the detergent solution in an amount in a range of from about 20% to about 40% by volume.
11. The method of claim 1 , comprising the further step of soaking the delipidated, decellularized adipose tissue in a disinfectant solution.
12. The method of claim 11 , comprising the further step of blending the delipidated, decellularized adipose tissue with the disinfectant solution.
13. The method of claim 11 , wherein the disinfectant solution is an acidic solution that includes peracetic acid.
14. The method of claim 11 , wherein the disinfectant solution includes an alcohol and a glycol.
15. The method of claim 11 , wherein substantially all substances that pose a significant risk of causing an immunogenic response in a patient receiving said adipose tissue-derived matrix are removed from the adipose tissue.
16. The method of claim 11 , wherein substantially all native lipids and native nucleic acids are removed from the adipose tissue.
17. The method of claim 1 , comprising the further step of contacting the delipidated, decellularized adipose tissue with a solution having an acidic pH so as to form a gel.
18. The method of claim 1 , comprising the further step of blending the delipidated, decellularized adipose tissue in a polar liquid, whereby the delipidated, decellularized adipose tissue and the polar liquid form a flowable gel.
19. The method of claim 1 , comprising the further steps of drying the delipidated, decellularized adipose tissue, and reducing the dried delipidated, decellularized adipose tissue to a dried particulate form.
20. The method of claim 8 , wherein the detergent solution includes a mixture of water and alcohol.
21. The method of claim 12 , wherein the disinfectant solution is an acidic solution that includes peracetic acid.
22. The method of claim 12 wherein the disinfectant solution includes an alcohol and a glycol.
23. The method of claim 12 , wherein substantially all substances that pose a significant risk of causing an immunogenic response in a patient receiving said adipose tissue-derived matrix are removed from the adipose tissue.
24. The method of claim 12 , wherein substantially all native lipids and native nucleic adds are removed from the adipose tissue.