Patent EA-004455-B1 (EA)
Title: Bio-reactor device and method of cultivating live cells
Inventor: VAN DER MERWE SCHALK
IPC: C12N5/071
Filed: 2001-08-27
Abstract: 1. A bio-reactor device (12), which comprises a holder (14), containing a three-dimensional matrix of open-cell foam material (18), the foam material (18) having a porous interior and containing, in its porous interior and anchored thereby, live cells, the bio-reactor device (12) also comprising a liquid contained in the holder (14), which liquid is an oxygen carrier and which saturates and immerses the matrix (18), the liquid comprising, at least in part, a synthetic or semi-synthetic oxygen-carrying constituent, the bio-reactor device (12) being characterized in that the matrix (18) is a unitary or integral continuous mass of the foam material (18), the foam material (18) being an open-cell foam material (18) and the device (12) farming part of an oxygen carrier liquid circulation circuit arranged for circulation of oxygen carrier liquid into the holder (14), for circulation of said liquid by perfusion through the matrix (18) in which the live cells are anchored, and for circulation thereof out of the holder (14). 2. A device as claimed in Claim 1, in which the open-cell foam material (18) is biocompatible and bio-stable, being a polymeric foam material (18). 3. A device as claimed in Claim 2, in which the foam material (18) is a foam of a polymer selected from the group consisting of polyurethane polymers, polyvinyl chloride polymers, polyethylene polymers, polypropylene polymers, polystyrene polymers, copolymers of the aforegoing, styrene-butadiene copolymers, styreneacrylonitrile copolymers, acrylonitrile-butadiene-styrene copolymers, poly (methylmethacrylate) polymers, polyamide polymers, poly (hexamethylene adipamide) polymers, poly (hexamethylene sebacamide) polymers, polycapralactone polymers, poly (ethylene terephthalate) polymers, poly (butylene terephthalate) polymers, polycarbonate polymers, polyacetal polymers, polyvinyl alcohol polymers, urea formaldehyde polymers, fluorine-containing polymers, silicone-containing polymers, and mixtures of the aforegoing polymers and copolymers. 4. A device as claimed in any one of the preceding claims, in which the live cells are mammalian cells, the oxygen carrier liquid forming a blood substitute which also acts as a carbon dioxide carrier. 5. A device as claimed in any one of the preceding claims, in which the oxygen carrier liquid comprises an aqueous liquid, the oxygen carrier liquid having, as the synthetic or semi-synthetic oxygen-carrying constituent, at least one member selected from the group consisting of cell-free haemoglobins, cross-linked haemoglobins, liposome-encapsulated haemoglobins and perfluorocarbons. 6. A device as claimed in Claim 5, in which the synthetic or semi-synthetic oxygen-carrying constituent is a synthetic perflourocarbon, the oxygen carrier liquid being an emulsion of an organic phase and an aqueous phase, the organic phase comprising the perfluorocarbon and being the disperse phase of the emulsion, and the aqueous phase being the continuous phase of the emulsion and comprising the aqueous liquid. 7. A device as claimed in Claim 6, in which the synthetic or semi synthetic constituent is a cell-free haemoglobin, the cell-free haemoglobin being dissolved in the aqueous liquid, such as blood plasma. 8. A device as claimed in Claims 6, in which the synthetic or semi-synthetic constituent is a liposome-encapsulated haemoglobin, the liposome-encapsulated haemoglobin being suspended as a suspension thereof in the aqueous liquid, such as blood plasma. 9. A device as claimed in any one of the preceding claims, in which the holder (14) is a reaction vessel or container (14) having an oxygen carrier liquid inlet and an oxygen carrier liquid outlet, the oxygen carrier liquid outlet being at a lower level than the 6 oxygen carrier liquid inlet and forming an oxygen carrier liquid drain, the holder (14) being provided with a temperature regulation device (16) for regulating the temperature of the matrix (18) and of the oxygen carrier liquid. 10. A device as claimed in any one of the preceding claims, which forms part of a temperature regulating liquid circuit for circulating a temperature regulating liquid to and away from the holder (14), the holder (14) being provided with a temperature regulation device (16) in the form of a hollow temperature regulating jacket (16) for the holder (14), the jacket (16) having an inlet (20) and an outlet (22) for a temperature regulating liquid, the temperature regulating circuit comprising a liquid circulation device for 570 circulating the temperature regulating liquid along and around the temperature regulating circuit, a thermostat, and a heat transfer device remote from the holder (14) and operative in response to signals from the thermostat to control the temperature of the temperature regulating liquid and hence to regulate the temperature of the matrix (18) and of the oxygen carrier liquid. 11. A device as claimed in any one of the preceding claims, in which the oxygen carrier liquid circulation circuit includes at least one device which is selected from the group consisting of liquid circulation devices (24) for circulating the oxygen carrier liquid along and around the circuit, biomass removal devices (30) for removing biomass from oxygen carrier liquid issuing from the holder (14), separation devices (38) for separating the synthetic or semi-synthetic constituent of the oxygen carrier liquid from other constituents thereof, carbon dioxide removal devices (44) for removing carbon dioxide from the oxygen carrier liquid, oxygenation devices (52) for oxygenating the oxygen carrier liquid, and de-aeration devices (64) for de-aerating the oxygen carrier liquid. 12. A device as claimed in Claim 11, in which the circulation circuit includes a plurality of the devices of the group, the devices being arranged, starting with the holder (14), in a sequence which comprises a single biomass removal device (30), followed in turn by a single separation device (38), a single carbon dioxide removal device (44), a single oxygenation device (52), a single de-aeration device (64) and a single liquid circulation device (24), the holder (14) following the liquid circulation device so that the circuit is an endless or closed circuit, and the liquid circulation device (24) being arranged to circulate the oxygen carrier liquid along and around the circuit in the direction in which the devices of the group follow one another in said sequence. 13. A device as claimed in any one of Claims 1-12, in which the oxygen carrier liquid circulation circuit is connected to a blood circulation circuit for circulating blood into contact with oxygen carrier liquid in the oxygen carrier liquid circulation circuit, for delivering cell-free blood constituents to the oxygen carrier liquid circuit, and for receiving or withdrawing cell-free aqueous oxygen carrier liquid constituents from the oxygen carrier liquid circuit, the blood circulation circuit including a blood circulation device (76) for circulating blood along and around the blood circulation circuit, a plasma separation device (68) for separating blood circulating along and around the blood circulation circuit into a cell-free blood plasma fraction and a blood cell-containing fraction containing blood cells and some blood plasma, and a de-aeration device (48) for de-aerating blood circulating along and around the blood circulation circuit. 14. A device as claimed in Claim 13, in which the devices of the blood circulation circuit are arranged in a sequence, in which sequence the blood circulation device (76) is followed in turn by the plasma separation device (68) and the de-aeration device (48), the blood circulation device (76) being arranged to circulate blood along and around the blood circulation circuit in the direction in which the devices of the blood circulation circuit follow one another in said sequence. 15. A device as claimed in Claim 13 or Claim 14, in which the connection between the oxygen liquid circulation circuit and the blood circulation circuit is a connection between the oxygen carrier liquid circulation circuit and the plasma separation device (68) of the blood circulation circuit. 16. A method of cultivating live cells, the method comprising circulating a liquid which is an oxygen carrier and which comprises, at least in part, a semisynthetic or synthetic oxygen-carrying constituent, through a matrix (18) of foam material having a porous interior in which live cells are anchored, the matrix forming part of an oxygen carrier liquid circulation circuit arranged for circulation of oxygen carrier liquid into a holder (14) containing the three-dimensional matrix (18), through the matrix (18), and out of the holder (14), the cells absorbing oxygen and nutrients from the liquid and the liquid absorbing carbon dioxide and other metabolic products from the cells, the method being characterized in that the matrix (18) is a unitary or integral continuous mass of open-cell foam material, and the circulating of the oxygen carrier liquid through the matrix (18) is by perfusing the oxygen carrier liquid through the matrix (18). 17. A method as claimed in Claim 16, characterized in that the oxygen carrier liquid comprises an aqueous constituent in the form of blood plasma, perfusing the liquid through the matrix (18) being by circulating the liquid along and around an oxygen carrier liquid circulation circuit and through the matrix (18), and the method including the step of regulating the temperature of the matrix (18) and oxygen carrier liquid, to keep the temperature of the matrix (18) and oxygen carrier squid within a desired range of values. 18. A method as claimed in Claim 17, characterized in that it includes the step of removing biomass from the oxygen carrier liquid circulating along and around the oxygen carrier liquid circuit and discharging the removed biomass to waste. 19. A method as claimed in Claim 17 or Claim 18, characterized in that it includes separatin