Extracellular matrix material
View Patent ↗An extracellular matrix material is described. The material has a cross-linked scaffold comprising fibrin or fibrinogen, and a bulking agent. Deposited on the scaffold is a calcium phosphate mineral phase. Also described are methods for forming such materials.
1. An extracellular matrix material, comprising:
a cross-linked scaffold comprising fibrin or fibrinogen, and a bulking agent, wherein the fibrin or fibrinogen and the bulking agent are covalently cross-linked, and the bulking agent is covalently cross-linked to the fibrin or fibrinogen; and
a ceramic deposited on the scaffold.
2. An extracellular matrix according to claim 1 , wherein the ceramic is, or comprises, a calcium phosphate mineral phase.
3. An extracellular matrix material according to claim 1 , wherein the bulking agent is an alginate, a glycosaminoglycan, hydroxyethylstarch, ethyl cellulose, Xanthan gum, or agarose.
4. A process for preparing an extracellular matrix material, comprising:
depositing a ceramic on a cross-linked scaffold comprising fibrin or fibrinogen, and a bulking agent, wherein the fibrin or fibrinogen and the bulking agent are covalently cross-linked, and the bulking agent is covalently cross-linked to the fibrin or fibrinogen.
5. A process according to claim 4 , wherein the ceramic is a calcium phosphate mineral phase.
6. A process according to claim 4 , wherein the calcium phosphate mineral phase is deposited on the scaffold by biomimetic deposition.
7. A process according to claim 4 , wherein the calcium phosphate mineral phase is deposited on to the scaffold by immersing the scaffold in a fluid comprising
i) Ca 2+ and HPO 4 2− ;
ii) Ca 2+ , HPO 4 2 , Na + , K + , Mg 2+ , Ca 2+ , Cl − , HCO 3 − , HPO 4 2− , HPO 4 2− and SO 4 2− ;
iii) Ca 2+ , HPO 4 2 , Na + , K + , Mg 2+ , Ca 2+ , Cl − , HCO 3 − , and HPO 4 2− ; or
iv) Ca 2+ , HPO 4 2 , Na + , and K + .
8. A process as defined in claim 5 , wherein the calcium phosphate mineral phase is deposited on to the scaffold by contacting the scaffold with, or immersing the scaffold in, a fluid comprising one or more of:
a) 1 to 30 mM Ca 2+ ;
b) 0.5 to 10 mM HPO 4 2− ;
c) 100 to 800 mM Na + ;
d) 1 to 50 mM K + ;
e) 0.5 to 20 mM Mg 2+ ;
f) 100 to 800 mM Cl − ;
g) 1 to 100 mM HCO 3 − ; and
h) 1 to 10 mM SO 4 2− .
9. A process according to claim 7 , wherein the scaffold is contacted with, or immersed in, the fluid for at least 12 hours, at least 24 hours, at least 2 days, at least 5 days, at least 6 days, or at least 9 days.
10. A process according to claim 7 , wherein the scaffold is contacted with, or immersed in the fluid for up to 20 days.
11. A process according to claim 4 , comprising generating the scaffold by:
(a) mixing an aqueous solution of fibrinogen with a coagulating agent and a bulking agent;
(b) incubating the mixture obtained in step (a) with a cross-linking agent; and
(c) washing the cross-linked composition obtained in step (b) to remove the cross-linking agent.
12. A process according to claim 4 , comprising forming the scaffold by a process comprising the following steps:
(a) mixing an aqueous solution of fibrinogen with a coagulating agent and a bulking agent;
(b) incubating the mixture obtained in step (a) with a cross-linking agent; and
(c) washing the cross-linked composition obtained in step (b) to remove the cross-linking agent.
13. A process as defined in claim 11 , wherein the coagulating agent comprises an enzymatic or non-enzymatic coagulating agent.
14. A process as defined in claim 11 , wherein step (a) comprises mixing with a foaming agent.
15. A process according to claim 14 , wherein the foaming agent comprises a non-ionic detergent, a thermosensitive gelling surfactant, a diphosphatydylglycerol type phospholipid, a mixture of an immiscible phase with the aqueous fibrinogen solution phase, a surfactant, or a sugar-acyl surfactant.
16. A process according to claim 15 , wherein the foaming agent sugar-acyl surfactants are selected from the class of pyranoside, maltoside, and acyl-sucrose surfactants.
17. A process according to claim 15 , wherein the sugar-acyl surfactants are selected from the group consisting of OGP, ODM, DGP and DdGP, TGP, HGP, DMP, decyl sucrose (nDS), dodecylsucrose (nDdS), OGM, and DMP and DdGP.
18. A process according to claim 11 , wherein the cross-linking agent used in step (b) is selected from: carbodiimide coupling agents, azide coupling agents; diisocyanate cross-linking agents; epoxide cross-linking agents; and aldehyde cross-linking agents.
19. A process according to claim 11 for preparing an extracellular matrix composition having a predetermined shape, wherein either (i) the mixture of step (a) is cast in a mould of a predetermined shape, frozen and optionally lyophilised prior to the incubation step (c), or; (ii) the product of step (d) is produced in a mould of a predetermined shape, and the product is then frozen and optionally lyophilised.
20. A method of bone regeneration, the method comprising application of an extracellular matrix material as defined in claim 1 to a bone defect.
21. The method of claim 20 , wherein the bone is in vitro, ex vivo or in vivo.
22. An extracellular matrix material according to claim 1 , which is lyophilized.