CN204779609U - Extracorporal circulatory system perfusion founds bioreactor of organizational project liver - Google Patents
Extracorporal circulatory system perfusion founds bioreactor of organizational project liver Download PDFInfo
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- CN204779609U CN204779609U CN201520113093.9U CN201520113093U CN204779609U CN 204779609 U CN204779609 U CN 204779609U CN 201520113093 U CN201520113093 U CN 201520113093U CN 204779609 U CN204779609 U CN 204779609U
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Abstract
The utility model relates to a three -dimensional apparatus and method for of cultivateing of tissue. This bioreactor includes external perfusion circulation system, pressure burden feedback regulation system and irritates flow control module, liver tissue culture module. External perfusion circulation system includes that confined pipe -line system, tissue pour into more new installation of generator and perfusion liquid suspendible. Pressure burden feedback regulation system includes that the interior pressure monitoring system of pipeline, computer control module and system receive the perfusion device that computer control adjusted. Liver tissue culture module includes the structure of sterile culture room and the three -dimensional support of tissue culture. The utility model discloses internal environment can be simulated, for in vitro culture organizational project organ provides a device, an instrument is provided for three -dimensional condition inferior trunk cell fifferentiation and cell growth's research.
Description
Technical field
The utility model belongs to organizational project and biological manufacture field, relates to the bio-reactor that (but being not limited to) liver organization dimensional culture builds organizational project liver.Can realize cell seeding, the perfusion in tissue culture stage and extracorporeal circulation in organizational project liver Process of in vitro, the blood circulation under simulation physiological condition, significant parameter can regulate.
Background technology
Liver is the removing toxic substances wanted of body weight for humans and digestion organs, and the End-stage liver disease such as hepatitis, liver cirrhosis, severe liver failure have had a strong impact on the life quality of people, become one of the highest disease of whole world mortality ratio.At present, liver transplantation is generally acknowledged unique effective methods for the treatment of.Only, about there are 300,000 End-stage liver disease patient wait orthotopic liver transplantations in China, and the patient that can accept orthotopic liver transplantation treatment only accounts for 45%.In global range, for liver shortage, the situation is tense, and Most patients is dead because by the time supplying liver.External structure organizational project liver becomes solution and supplies one of approach of liver shortage.
Organizational engineering closes ultimate principle, basic theories, basic fundamental and the method for using biology and engineering science, and how research is built with bioactive histoorgan, under in vitro conditions in order to the subject of the histoorgan of alternative disease damage.Organizational project liver is one of the study hotspot in this field.Build desirable engineered artificial liver, need research three critical problems: desirable seed cell source, timbering material and bio-reactor close to physiological environment can be provided for cytoskeleton mixture.
In the evolution of hepatic tissue bio-reactor, successively plate bio-reactor, rotation bottle-type bio-reactor, hollow fiber bioreactor, perfusion bedrest type bio-reactor (BasakE.Uygun are there is, GavriellePrice, NimaSaeidi, Maria-LouisaIzamis, TimBerendsen, MartinYarmush, KorkutUygun.DecellularizationandRecellularizationofWhole Livers.J.Vis.Exp.2011, (48): e2394), liver cell parcel/suspension type biological reactor.Because above-mentioned bio-reactor does not all solve the problem that oxygen supply and cell grow in three-dimensional rack, their use is restricted.At present these bio-reactors can Some substitute exhaustion liver in the short period of time, but it can not substitute liver function completely, real organizational project liver (YuCB can not be turned out, PanXP, LiLJ.2009.Progressinbioreactorsofbioartificiallivers.Hep atobiliaryPancreatDisInt8:134-140.).
Therefore, when building tissue engineering artificial liver bio-reactor in vitro, should be taken into account: perfusion effect enough ensures the planting depth of cell in three-dimensional rack, provides the continous perfusion of condition in analogue body, and provide the biology in analogue body, mechanical environment and enough nutrition to cell to exchange.
The utility model provides the tissue culture environment under simulation physiological condition, and implements adjustable hydrostaticpressure to cell.
Utility model content
The purpose of this utility model is that design can build the bio-reactor of organizational project liver by dimensional culture liver organization, for the in vitro study of differentiation of stem cells, the external structure of organizational project liver, substitute the perfect condition of human body for liver to realize bioartificial liver.
The bio-reactor that the utility model provides, comprising:
Nutrient solution storage tank, is provided with perfusion nutrient solution in described nutrient solution storage tank;
Tubing system, described tubing system comprises inlet tube and outlet pipe;
Sterile culture room, described sterile culture indoor are provided with three-dimensional rack; Described three-dimensional rack comprises support influx and support spout, described support influx is connected with one end of inlet tube, described support spout is connected with one end of outlet pipe, and the other end of described inlet tube is all connected with described nutrient solution storage tank with the other end of described outlet pipe;
Perfusion device, described perfusion device is arranged on described inlet tube and described outlet pipe.
Wherein, described perfusion device comprises the first peristaltic pump and the second peristaltic pump.
Wherein, this bio-reactor comprises pressure negative-feedback regu-lation system, and described pressure negative-feedback regu-lation system comprises manifold pressure Monitoring systems, computer central control system and the perfusion device by computer central control system regulating and controlling.
Wherein, described manifold pressure Monitoring systems is pressure transmitter.
Wherein, described pressure transmitter imports the pressure signal in tubing system into computer central control system in real time, and computer central control system regulates described first peristaltic pump and the second peristaltic pump automatically according to described pressure signal, thus controls perfusion rate and perfusion flow.
Wherein, also comprise perfusate suspendible updating device, described perfusate suspendible updating device comprises suspendible shaking table, Y-tube.
Wherein, described Sterile culture room is made by aseptic biocompatible materials.
Wherein, Growth of Cells is inner for the described three-dimensional rack cultivating hepatic tissue, and this three-dimensional rack has the vascular system of simulation liver topography.
Wherein, described three-dimensional rack is resilient material.
Wherein, described nutrient solution storage tank and described Sterile culture room are positioned in CO2gas incubator.
This bio-reactor comprises: extracorporeal circulation perfusion system, containing extracorporeal circulation pipeline system, tissue perfusion producer, the perfusate suspendible updating device closed; Manifold pressure negative-feedback regu-lation system, comprises manifold pressure Monitoring systems, conputer controlled module, perfusion device (peristaltic pump) by computer regulating and controlling containing pressure negative-feedback regu-lation system; Liver organization culture system, containing the Sterile culture room of hepatic tissue cultivation, for the three-dimensional rack cultivating hepatic tissue.
The tubing system of nutrient solution in nutrient solution storage tank by closing, enter liver through the first peristaltic pump and cultivate three-dimensional rack, for three-dimensional rack inner cell provides nutritive substance, the cell of three-dimensional rack and inside thereof is cultivated in Sterile culture room.Nutrient solution, after three-dimensional rack internal recycle, enters closed tubing system through support spout, enters nutrient solution storage tank under the control of the second peristaltic pump.There is pressure monitor system can Real-Time Monitoring pipeline system internal pressure in the tubing system closed, and import pressure information into computer central control system, computer central control system is according to this pressure information and the pressure demand preset, regulate the speed of two peristaltic pumps, thus maintain the hydrostaticpressure in whole tubing system.
Nutrient solution storage tank separately has aseptic fluid inlet, gas feed (gas such as oxygen, carbonic acid gas can be entered in nutrient solution storage tank by gas feed) and gas-filtering device, to meet nutrient solution oxygenation, add carbonic acid gas, change nutrient solution and to eliminate the demand of bubble in tubing system.
In such scheme, whole tubing system is closed system, and tubing system comprises inflow pipe and outlet pipe, and inflow pipe and outlet pipe all adopt aseptic biocompatible materials.Sterile culture room also adopts sterile, biocompatible material.
Another technical characteristic of the utility model has real-time pressure sensors in tubing system, and the pump speed flow of peristaltic pump is regulated by the monitoring of computer central control system all in real time.
The utility model has the following advantages compared with already present liver bio-reactor:
The first, circulated completely in whole three-dimensional rack by nutrient solution, cell suspension can be taken to each corner of whole three-dimensional rack, thus improve the planting density of cell on three-dimensional rack, the distribution of Reality simulation liver inner cell.
The second, circulated completely in whole three-dimensional rack by nutrient solution, can be all cells in three-dimensional rack and effective nutrition supply, gaseous interchange are provided, the survival rate of internal stent cell is provided.
3rd, realizing pulsation by peristaltic pump stimulates, blood flow in analogue body.
4th, by manifold pressure Monitoring systems, computer central control system, peristaltic pump negative-feedback regu-lation, hydrostaticpressure in liver under effective realization simulation physiological condition.And the adjustment of hydrostaticpressure, be beneficial to the Mechanism Study of Vitro hepatic regeneration.
5th, computer central control system, by software adjustment, adapts to different Research Requirements and tissue construction demand.
Accompanying drawing explanation
Fig. 1 is the theory structure sketch that extracorporeal circulation perfusion builds the bio-reactor of organizational project liver.
Reference numeral:
1-nutrient solution storage tank; 2-gas feed;
3-tubing system; 5-first peristaltic pump;
6-second peristaltic pump; 7-pressure monitor system;
8-computer central control system; 9-Sterile culture room;
10-three-dimensional rack 11-support influx;
12-support spout.
Embodiment
The utility model is cellularised and two stages of tissue culture from timbering material, for external structure organizational project liver provides technical scheme.
The bio-reactor that the utility model provides, comprising:
Nutrient solution storage tank 1, is provided with perfusion nutrient solution in described nutrient solution storage tank 1;
Tubing system 3, described tubing system 3 comprises inlet tube and outlet pipe;
Sterile culture room 9, is provided with three-dimensional rack 10 in described Sterile culture room 9; Described three-dimensional rack 10 comprises support influx 11 and support spout 12, described support influx 11 is connected with one end of inlet tube, described support spout 12 is connected with one end of outlet pipe, and the other end of described inlet tube is all connected with described nutrient solution storage tank 1 with the other end of described outlet pipe
Perfusion device, described perfusion device is arranged on described inlet tube and described outlet pipe.
Wherein, described perfusion device comprises the first peristaltic pump 5 and the second peristaltic pump 6.
Wherein, this bio-reactor comprises pressure negative-feedback regu-lation system, and described pressure negative-feedback regu-lation system comprises manifold pressure Monitoring systems 7, computer central control system 8 and the perfusion device by computer central control system 8 regulating and controlling.
Wherein, described manifold pressure Monitoring systems 7 is pressure transmitter.
Wherein, described pressure transmitter imports the pressure signal in tubing system 3 into computer central control system 8 in real time, and computer central control system 8 regulates described first peristaltic pump 5 and the second peristaltic pump 6 automatically according to described pressure signal, thus controls perfusion rate and perfusion flow.
Wherein, also comprise perfusate suspendible updating device, described perfusate suspendible updating device comprises suspendible shaking table, Y-tube.
Wherein, described Sterile culture room 9 is made by aseptic biocompatible materials.
Wherein, Growth of Cells is inner for the described three-dimensional rack 10 cultivating hepatic tissue, and this three-dimensional rack 10 has the vascular system of simulation liver topography.
Wherein, described three-dimensional rack 10 is resilient materials.
Wherein, described nutrient solution storage tank 1 and described Sterile culture room 9 are positioned in CO2gas incubator.
Nutrient solution storage tank 1 separately has aseptic fluid inlet, gas feed 2 (gas such as oxygen, carbonic acid gas can be entered in nutrient solution storage tank 1 by gas feed) and gas-filtering device, to meet nutrient solution oxygenation, add carbonic acid gas, change nutrient solution and to eliminate the demand of bubble in tubing system 3.
The stage that timbering material is cellularised, i.e. cell seeding stage.Nutrient solution storage tank 1 and Sterile culture room 9 are placed in CO2gas incubator, cell suspension in nutrient solution storage tank 1 passes through continous perfusion, or two pipeline alternating currents are to input, make cell be distributed to each position of three-dimensional rack 10, improve the adhesive rate of cell on three-dimensional rack 10 material.Cell infusion time and concentration can depend on the needs.After quiescent culture for some time, tissue perfusion cultivation can be carried out.
In the tissue culture stage, as shown in Figure 1, nutrient solution storage tank 1 and Sterile culture room 9 are also placed in CO2gas incubator.The tubing system 3 of nutrient solution in cell culture fluid storage tank 1 by closing, enter liver through the first peristaltic pump 5 and cultivate three-dimensional rack 10, for three-dimensional rack 10 inner cell provides nutritive substance, the cell of three-dimensional rack 10 and inside thereof is cultivated in Sterile culture room 9.Nutrient solution, after three-dimensional rack 10 internal recycle, enters closed tubing system 3 through support spout 12, under the control of the second peristaltic pump 6, enter nutrient solution storage tank 1.There is pressure monitor system 7 can Real-Time Monitoring pipeline system 3 internal pressure in the tubing system 3 closed, and import pressure information into computer central control system 8, computer central control system 8 is according to pressure and the pressure demand preset, regulate the speed of two peristaltic pumps, thus maintain the hydrostaticpressure in whole tubing system 3.
Embodiment 1: stem cell takes off cytoskeleton full liver and cultivates
To under aseptic condition, after the full liver of rat takes off cell, obtain the biological three-dimensional rack with liver network structure and vascular system.Support influx 11 is hepatic vein, and support spout 12 is hepatic vein, support influx 11 and support spout 12 is connected to inlet tube and the outlet pipe of bio-reactor.The inlet tube of bio-reactor is connected with nutrient solution storage tank 1 with the other end of outlet pipe, in nutrient solution storage tank 1 aseptically preset 10
6/ ml stem cell suspension.Nutrient solution storage tank 1, nutrient solution suspendible device and Sterile culture room 9 are placed in CO2gas incubator, by peristaltic pump, cell suspension are slowly pumped into full liver and take off in cytoskeleton, pour into 1 circulation.Leave standstill after one day, liquid in nutrient solution storage tank 1 is replaced by tissue culture medium, carries out continous perfusion cultivation.After one week, observe culture effect.
Embodiment 2: nutrient solution is on the impact of liver growth
Rats'liver is excised under aseptic condition, after red corpuscle in surface and pipeline is rinsed well, measuring liver volume size, be placed in Sterile culture room 9, hepatic vein is connected the inlet tube of bio-reactor, hepatic vein connection outlet pipe, is connected the inlet tube of bio-reactor with nutrient solution storage tank 1 with the other end of outlet pipe.Nutrient solution storage tank 1, nutrient solution suspendible device and liver Sterile culture room 9 are placed in CO2gas incubator, by peristaltic pump, liver nutrient solution continous perfusion is entered in rats'liver, maintaining hepatic vein pressure at inlet is 10cm water column (being about physiological status pressure), support and measure liver size after two weeks, observe nutrient solution to the impact of liver growth.
Embodiment 3: different hydrostaticpressure is on the impact of liver regeneration
Rats'liver is excised under aseptic condition, after red corpuscle in surface and pipeline is rinsed well, measuring liver volume size, be placed in Sterile culture room 9, hepatic vein is connected the inlet tube of bio-reactor, hepatic vein connection outlet pipe, is connected the inlet tube of bio-reactor with nutrient solution storage tank 1 with the other end of outlet pipe.Nutrient solution storage tank 1, nutrient solution suspendible device and Sterile culture room 9 are placed in CO2gas incubator, by peristaltic pump, liver nutrient solution continous perfusion is entered in rats'liver, maintaining hepatic vein pressure at inlet is that 15cm water column cultivates (higher than physiological status), measure liver size after cultivating two weeks, observe high hydrostatic pressure to the impact of liver growth.
Continous perfusion circulation is the key factor of regulating cell growth and function.The feature of the utility model co-culture device can be the extracorporeal circulation that cultured cells in three-dimensional rack 10 provides lasting simulation physiological condition, realize the effect that cell growth provides nutrition, gaseous interchange, for co-culture of cells provides a kind of adjustable growing environment, provide a kind of method for exploring tissue culture technique.
The foregoing is only the utility model preferred embodiment, be not limited to the utility model, for a person skilled in the art, the utility model can have various modifications and variations.All within spirit of the present utility model and principle, any amendment done, equivalent replacement, improvement etc., all should be included within protection domain of the present utility model.
Claims (10)
1. extracorporeal circulation perfusion builds a bio-reactor for organizational project liver, it is characterized in that, comprising:
Nutrient solution storage tank, is provided with perfusion nutrient solution in described nutrient solution storage tank;
Tubing system, described tubing system comprises inlet tube and outlet pipe;
Sterile culture room, described sterile culture indoor are provided with three-dimensional rack; Described three-dimensional rack comprises support influx and support spout, described support influx is connected with one end of inlet tube, described support spout is connected with one end of outlet pipe, and the other end of described inlet tube is all connected with described nutrient solution storage tank with the other end of described outlet pipe;
Perfusion device, described perfusion device is arranged on described inlet tube and described outlet pipe.
2. extracorporeal circulation perfusion as claimed in claim 1 builds the bio-reactor of organizational project liver, and it is characterized in that, described perfusion device comprises the first peristaltic pump and the second peristaltic pump.
3. extracorporeal circulation perfusion as claimed in claim 2 builds the bio-reactor of organizational project liver, it is characterized in that, this bio-reactor comprises pressure negative-feedback regu-lation system, and described pressure negative-feedback regu-lation system comprises manifold pressure Monitoring systems, computer central control system and the perfusion device by computer central control system regulating and controlling.
4. extracorporeal circulation perfusion as claimed in claim 3 builds the bio-reactor of organizational project liver, and it is characterized in that, described manifold pressure Monitoring systems is pressure transmitter.
5. extracorporeal circulation perfusion as claimed in claim 4 builds the bio-reactor of organizational project liver, it is characterized in that, described pressure transmitter imports the pressure signal in tubing system into computer central control system in real time, computer central control system regulates described first peristaltic pump and the second peristaltic pump automatically according to described pressure signal, thus controls perfusion rate and perfusion flow.
6. the extracorporeal circulation perfusion according to any one of claim 1-5 builds the bio-reactor of organizational project liver, and it is characterized in that, also comprise perfusate suspendible updating device, described perfusate suspendible updating device comprises suspendible shaking table, Y-tube.
7. the extracorporeal circulation perfusion according to any one of claim 1-5 builds the bio-reactor of organizational project liver, and it is characterized in that, described Sterile culture room is made by aseptic biocompatible materials.
8. the extracorporeal circulation perfusion according to any one of claim 1-5 builds the bio-reactor of organizational project liver, it is characterized in that, Growth of Cells is inner for the described three-dimensional rack cultivating hepatic tissue, and this three-dimensional rack has the vascular system of simulation liver topography.
9. the extracorporeal circulation perfusion according to any one of claim 1-5 builds the bio-reactor of organizational project liver, and it is characterized in that, described three-dimensional rack is resilient material.
10. the extracorporeal circulation perfusion according to any one of claim 1-5 builds the bio-reactor of organizational project liver, and it is characterized in that, described nutrient solution storage tank and described Sterile culture room are positioned in CO2gas incubator.
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Cited By (6)
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| CN105586249A (en) * | 2016-03-07 | 2016-05-18 | 浙江大学 | Circulating perfusion biological reactor device capable of achieving circulating perfusion of three-dimensional support |
| CN108465126A (en) * | 2018-05-25 | 2018-08-31 | 中国人民解放军总医院 | Simulate the preparation facilities of the bladder acellular matrix of physiology course |
| CN109055216A (en) * | 2018-09-21 | 2018-12-21 | 中国科学院苏州生物医学工程技术研究所 | High-throughput 3D cell, class loading and organoid dynamic cultivation system |
| CN109312289A (en) * | 2016-05-19 | 2019-02-05 | 齐藤光次 | Culture apparatus, cultural method and the culture organ produced using the cultural method |
| CN110636815A (en) * | 2017-05-02 | 2019-12-31 | 塞拉Ip股份公司 | Intracorporeal perfusion system |
| CN111304078A (en) * | 2018-12-11 | 2020-06-19 | 上海市第五人民医院 | Device and method for constructing tissue engineering bone tissue by simulating cerebrospinal fluid biomechanical environment |
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2015
- 2015-02-16 CN CN201520113093.9U patent/CN204779609U/en not_active Expired - Fee Related
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105586249A (en) * | 2016-03-07 | 2016-05-18 | 浙江大学 | Circulating perfusion biological reactor device capable of achieving circulating perfusion of three-dimensional support |
| CN109312289A (en) * | 2016-05-19 | 2019-02-05 | 齐藤光次 | Culture apparatus, cultural method and the culture organ produced using the cultural method |
| US11306281B2 (en) | 2016-05-19 | 2022-04-19 | Koji Saito | Culture device, culture method and cultured organ produced by the culture method |
| CN110636815A (en) * | 2017-05-02 | 2019-12-31 | 塞拉Ip股份公司 | Intracorporeal perfusion system |
| CN110636815B (en) * | 2017-05-02 | 2022-03-08 | 塞拉Ip股份公司 | Intracorporeal perfusion system |
| US11529463B2 (en) | 2017-05-02 | 2022-12-20 | Seraip Ag | In-body perfusion system |
| CN108465126A (en) * | 2018-05-25 | 2018-08-31 | 中国人民解放军总医院 | Simulate the preparation facilities of the bladder acellular matrix of physiology course |
| CN109055216A (en) * | 2018-09-21 | 2018-12-21 | 中国科学院苏州生物医学工程技术研究所 | High-throughput 3D cell, class loading and organoid dynamic cultivation system |
| CN109055216B (en) * | 2018-09-21 | 2024-03-22 | 中国科学院苏州生物医学工程技术研究所 | High-throughput 3D cell, tissue-like and organ-like dynamic culture system |
| CN111304078A (en) * | 2018-12-11 | 2020-06-19 | 上海市第五人民医院 | Device and method for constructing tissue engineering bone tissue by simulating cerebrospinal fluid biomechanical environment |
| CN111304078B (en) * | 2018-12-11 | 2023-12-08 | 上海市第五人民医院 | Device and method for constructing tissue engineering bone tissue by simulating cerebrospinal fluid biomechanical environment |
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