CN104207859A - Method and special equipment utilizing spin accumulation method to prepare tissues and organs - Google Patents
Method and special equipment utilizing spin accumulation method to prepare tissues and organs Download PDFInfo
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- CN104207859A CN104207859A CN201410473055.4A CN201410473055A CN104207859A CN 104207859 A CN104207859 A CN 104207859A CN 201410473055 A CN201410473055 A CN 201410473055A CN 104207859 A CN104207859 A CN 104207859A
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
Abstract
The invention relates to a method and special equipment utilizing a spin accumulation method to prepare tissues and organs, and belongs to the technical field of tissue engineering. Based on a core dissolving technology and a cell assembly technology, various cell growth factors, inhibitors or medicines are compounded into a cross-linkable/polymerized hydrogel material, filamentous hydrogel extruded from nozzles is crosslinked/polymerized and compositely synthesized into a polymer solution, is washed by a culture solution and then is wound and accumulated on a rotary inner core, the hydrogel containing various types of seed cells and the hydrogel to which endothelial cells and nerve cells are adhered respectively form a main body part and a blood vessel and nerve system, the inner core is dissolved after completion of forming and the tissue and organ with spiral blood vessels and nerves are obtained after culturing for a period of time. The method and the special equipment provided by the invention are simple in process and easy in forming of the blood vessel and nerve system, and is especially suitable for forming the tissue and organ with a lumen difficultly formed by the traditional tissue engineering, and the prepared tissue and organ can be applied to a damaged tissue and organ or directly transplanted in vivo to replace the damaged tissue and organ.
Description
Technical field
The present invention relates to the method and the special equipment that utilize and rotate method of piling and prepare histoorgan, belong to field of tissue engineering technology.
Background technology
The reparation of human body disease damage tissue and organ and to substitute be the science frontier attracted people's attention this century is also a sciences problems needing solution badly.The intrinsic drawback such as to have a big risk because biological tissue organ transplantation exists donor shortage and immunologic rejection, be difficult to become the effective way solving this problem.Accordingly, arise at the historic moment by the organizational project (Tissue Engineering) improved for the purpose of this type of illness treatment level, organizational project is intended to the three-dimensional structure of external structure cell and material, by developing into the transplantation substitute had with natural tissues similar structures and function after suitable cultivation and training.In recent years, organizational project is the focus of research both at home and abroad always, develops rapidly and achieves many achievements, but with regard to its product applications, mainly concentrates on relatively simple a few field such as bone tissue engineer and skin tissue engineering of structure at present.Trace it to its cause, the very important point is exactly, the vitro construction method (as Electrospun, die casting etc.) of existing organizational project structure is difficult to build the blood vessel network structure with the similar complexity of natural tissues, cause the cell survival rate of construction inner low, be therefore difficult to the structure realizing bulk tissue.
Traditional organizational project follows the Constructed wetlands of " from top to bottom " of " cell-scaffold compound ", i.e. first shaped bracket, then cell seeding on support.But when stent size and complexity are brought up to a certain degree, the difficulty of stent forming and cell compound will increase greatly.Cell three-dimensional controlled group packing technique (Wang X H, Yan Y N, Zhang R J.Rapid prototyping as a tool for manufacturing bio-artificial livers.Trends in Biotechnology, 2007,25 (11): 505-513.), i.e. cell 3D printing technique, follows the thinking that " support-cell " integration builds.In cell printing process, cell (or cell aggregation) and colloidal sol (presoma of hydrogel) are placed in simultaneously or celliferous culture fluid is placed in separately the shower nozzle of printer, the deposition position containing cell drop is controlled by computer, print in the position pointwise of specifying, on the basis having printed one deck continue print another layer, be layering formed three-dimensional Duo Xi Bao ?gel rubber system.The feature of cell printing technology is the uniformity that can ensure cell density and distribution, and can realize common assembling and the fixed point arrangement of various kinds of cell.But effective transportation problem of oxygen and nutrient substance, namely " vascularization problem " remains the bottleneck problem that the method builds transplantation complicated tissue organ.
The existing histoorgan by cell 3D printing-forming is all pile up according to plane in layer, and because of the inherent characteristics that it is shaped, the shaping of microchannel (blood vessel) depends on the character of forming accuracy and material.And for having the shaping of inner-cavity structure histoorgan, the cell printing technology relying on plane to pile up inevitably needs increase support or rotate forming table or shower nozzle orientation in its forming process, the difficulty considerably increasing design and be shaped, sometimes even also helpless.
Chinese patent literature CN1654028 proposes a kind of method utilizing molten core legal system tubing network, the material of water solublity, inanimate object toxicity is first utilized to manufacture the inner core of support, then biocompatible materials is coated on support inner core, water-soluble except support inner core with distillation after air-dry.The method can only the better simply grid shape stent of shaped structure, and the shaping for the histoorgan with complicated blood vessel is helpless.
Summary of the invention
The object of the invention is to overcome in traditional organizational project the cavity type histoorgan being difficult to build and there is complicated blood vessel and neuromechanism, a kind of method and the special equipment of preparing histoorgan based on rotation method of piling are provided.The method is on the basis of cell controlled group packing technique and soluble core technique, object is the preparation method providing a kind of new histoorgan, be convenient to prepare and there is spiral net-shaped blood vessel and neural histoorgan, be particularly useful for making the histoorgan with inner-cavity structure.
Technical scheme of the present invention is as follows:
Utilize the process rotating method of piling and prepare histoorgan, it is characterized in that special equipment of the present invention comprises bracing or strutting arrangement, many nozzle components, motion, forming device and control system; Described bracing or strutting arrangement comprises supporting leg and base plate; Described motion adopt Dao Gui ?slide block mechanism, comprise X direction guiding rail, X to slide block, Y-direction guide rail, the first Y-direction slide block, the second Y-direction slide block, Z-direction guide rail, Z-direction slide block, described X direction guiding rail is located on base plate, and Z-direction guide rail is fixed in X on slide block, and Y-direction guide rail is fixed on Z-direction slide block; Described many nozzle components are at least containing six shower nozzles, comprise at least two squash type shower nozzles and four spray-type shower nozzles, the nozzle inside diameter of two squash type shower nozzles is respectively 200 μm ~ 1mm and 50 μm ~ 500 μm, the mesh diameter of spray-type spray nozzle is 50 μm ~ 100 μm, and shower nozzle divides two groups to be arranged on the first Y-direction slide block and the second Y-direction slide block respectively; Described forming device comprises forming containers, support and motor, and forming containers to be arranged on base plate and to be in the bottom of nozzle component, and support installing is in forming containers and be connected with motor output shaft.
Special equipment of the present invention is characterized in that: the structure of described support to be two ends be cylindric, middle be prism-shaped, material is rustless steel or titanium alloy.
Special equipment of the present invention is characterized in that: described first Y-direction slide block and the second Y-direction slide block have and first of Y-direction guide rail parallel the little guide rail and the second little guide rail, the first small slide block installed respectively by first little guide rail and the second little guide rail) and the second small slide block, the first small slide block and the second small slide block have can the hole of stationary nozzle.
Utilize the method rotating method of piling and prepare histoorgan, it is characterized in that the method comprises the following steps:
1) material preparation:
Preparation 0.2 ~ 20% (w/v) hydrogel solution, 1 ~ 10% (w/v) cross-linking agent/polymer fluid, 0.1 ~ 10% (w/v) synthesize macromolecular solution, 0.2 ~ 10% (w/v) gelatin solution and cell culture fluid; Be separated or induction seed cell, seed cell density is 1 × 10
3~ 1 × 10
6individual/mL, is mixed and made into the hydrogel solution containing seed cell by seed cell and hydrogel solution by 1 ~ 9:9 ~ 1 volume ratio; Be separated or inducing nerve cell and endotheliocyte, and make cell suspension, neurocyte suspension and endotheliocyte suspension density are 1 × 10
3~ 1 × 10
8individual/mL;
2) inner core preparation
When preparation has the histoorgan of inner chamber, according to the cavity shape will preparing histoorgan, with its threedimensional model of three-dimensional drawing software design, make the mould with this inner-cavity structure by quick shaping method, when the histoorgan of forming solid, mould is the cylinder of the thick 8 ~ 10mm of diameter; Support is put into mould central authorities, reducing temperature to 8 ~ 10 DEG C add gelatin solution in mould after makes gelatin solidify, open the histoorgan inner core that mould obtains center belt supporting frame, apply hydrogel outward at the histoorgan inner core of belt supporting frame, and carry out being cross-linked/being polymerized with cross-linking agent/polymer fluid;
3) be shaped
The hydrogel solution and not celliferous hydrogel solution that contain seed cell are respectively charged in the first squash type shower nozzle and the second squash type shower nozzle, cross-linking agent/polymer fluid, neurocyte suspension, endotheliocyte suspension and synthesis macromolecular solution are respectively charged into the 4th spray-type shower nozzle, second spray-type shower nozzle, in first spray-type shower nozzle and the 3rd spray-type shower nozzle, cell culture fluid is loaded in forming containers, support installing to be connected with motor output shaft in forming containers, after the model of histoorgan to be formed is imported control system, start former, start stack shaping, in forming process, support rotates around Y negative sense, shower nozzle does translational motion, when the main part of shaping histoorgan, first squash type shower nozzle moves to the hydrogel that assigned address extrudes containing seed cell and is wrapped on inner core, and the dead astern that the 4th spray-type shower nozzle that cross-linking agent/polymer fluid is housed simultaneously moves to the first squash type shower nozzle sprays cross-linking agent/polymer fluid and is cross-linked it/is polymerized, when shaping blood vessel, second squash type shower nozzle moves to assigned address and extrudes not celliferous hydrogel and be wrapped on inner core, 4th spray-type shower nozzle moves to the second squash type shower nozzle dead astern simultaneously, first spray-type shower nozzle and the 3rd spray-type shower nozzle move to the second squash type shower nozzle dead ahead respectively, under the inner core rotated drives, carry out being cross-linked/being polymerized under the hydrogel of new winding moves to the 4th spray-type shower nozzle successively, wash through culture fluid, to the first spray-type shower nozzle, sprinkling endotheliocyte and composite synthesis macromolecular solution is carried out under 3rd spray-type shower nozzle, as body and the spirit through time, second squash type shower nozzle moves to assigned address and extrudes not celliferous hydrogel and be wrapped on inner core, 4th spray-type shower nozzle moves to the second squash type shower nozzle dead astern simultaneously, second spray-type shower nozzle and the 3rd spray-type shower nozzle move to the second squash type shower nozzle dead ahead respectively, under the inner core rotated drives, carry out being cross-linked/being polymerized under the hydrogel of new winding moves to the 4th spray-type shower nozzle successively, wash through culture fluid, to the second spray-type shower nozzle, the 3rd spray-type shower nozzle, carry out sprinkling neurocyte and composite synthesis macromolecule, so repeatedly be wound around and pile up until form the histoorgan precursor containing inner core,
4) subsequent treatment
Cultivating being placed in calorstat containing the histoorgan precursor of inner core, moltenly taking out support except after inner core, after cultivating, form rotational-like histoorgan, neural and blood vessel spirally runs through wherein.
The method preparing histoorgan of the present invention, is characterized in that: described seed cell is have the stem cell of differentiation capability or have the adult cell of physiologically active; Described stem cell is fat stem cell, embryonic stem cell, blood stem cell, bone marrow stem cell, induction type versatile stem cell, and described adult cell is adipose cell, hepatocyte, bladder cells, lymphocyte, myocardial cell or nephrocyte; Described hydrogel is the sodium alginate or the fibrinogen solution that to be compounded with in gelatin, collagen, matrigel, carrageenan, chitosan, agar, hyaluronic acid, matrigel, elastin laminin and laminin one or more; In described hydrogel can the various cell cryopreservation agent of compound, cell growth factor, medicine and anticoagulant; Cell cryopreservation agent is one or more complex in dimethyl sulfoxide (DMSO), glycerol and dextrose; Described cell growth factor is one or more complex in VEGF (VEGF), basic fibroblast growth factor (b-FGF), hepatocyte growth factor (HGF), human blood platelets derived growth factor (PDGF-BB) and transforminggrowthfactor-β1 (TGF-β 1); Described medicine is antitumor drug, this antitumor drug is astragalus polysaccharides, cis diamino dichloro network platinum (DDP), mitomycin (MMC), one or more complex in 5-fluorouracil (5-FU), Claritin, antibiotic, viral vaccine; Described anticoagulin is one or more complex in heparin, paclitaxel; The solute of described synthesis macromolecular solution is one or more in polyurethane, polycaprolactone, Merlon, Polyethylene Glycol, Poly(D,L-lactide-co-glycolide, polyester and polyhydroxy acid ester, and solvent is TEG or Isosorbide-5-Nitrae-dioxane.
The present invention compared with prior art, has the technique effect of following advantage and salience:
1. molten core method and 3D Method of printing combine by the present invention, and traditional 3D Method of printing that can be shaped is difficult to the histoorgan with inner chamber be shaped;
2. the use of many shower nozzles can realize the shaping of various kinds of cell heterogeneous body and surface compound treatment, meets the requirement of natural tissues organ to various kinds of cell;
3. spray blood vessel (nerve) generation type of endothelium (nerve) cell in hydrogel surface to be shaped the blood vessel (nerve) with labyrinth, the thin film that synthesis macromolecular solution is formed improves endothelium (nerve) cell number that hydrogel surface adheres to, and effectively promotes the formation of blood vessel (nerve);
4. in forming process, cell culture fluid can be that cells with nutrient material again can organic solvent in the cross-linking agent/polymer fluid on detergent gel surface and synthesis macromolecule, reduces its damage to cell.
Accompanying drawing explanation
Fig. 1 is special equipment three dimensional structure sketch of the present invention.
Fig. 2 is the three dimensional structure sketch of the first Y-direction slide block.
Fig. 3 is histoorgan forming process schematic diagram of the present invention.
Fig. 4 is histoorgan forming technology flow chart of the present invention.
Fig. 5 a is not celliferous hydrogel; Fig. 5 b is for spraying the hydrogel after endothelium (nerve) cell; Fig. 5 c is the hydrogel after spraying synthesis macromolecular solution.
Fig. 6 is the structural representation of histoorgan of the present invention.
In figure: 101 ?base plate; 102 ?forming containers; 103 ?support; 104 ?the first spray-type shower nozzle; 105 ?the second spray-type shower nozzle; 106 ?Z-direction slide block; 107 ?Y-direction guide rail; 108 ?the 3rd spray-type shower nozzle; 109 ?the first small slide block; 110 ?the first little guide rail; 111 ?the first Y-direction slide block; 112 ?the second Y-direction slide block; 113 ?the first squash type shower nozzle; 114 ?the second squash type shower nozzle; 115 ?the 4th spray-type shower nozzle; 116 ?the second small slide block; 117 ?the second little guide rail; 118 ?Z-direction guide rail; 119 ?electric rotating machine; 120 ?X to slide block; 121 ?X direction guiding rail; 401 ?inner core; 402 ?nozzle component; 403 ?material; 404 ?water-setting collodion silks; 405 ?culture fluid; 601 ?main part; 602 ?blood vessel (nerve); 603 ?inner chamber.
Detailed description of the invention
Below in conjunction with drawings and Examples, the present invention is further described.
Fig. 1 is the special equipment utilizing rotation method of piling to prepare histoorgan provided by the invention, and this equipment comprises many nozzle components, motion, forming device and control system (picture).Described motion is realized by slide block-guide rail (groove), comprises X direction guiding rail 121 and X to slide block 120, Y-direction guide rail 107 and the first Y-direction slide block 111, second Y-direction slide block 112, Z-direction guide rail 118 and Z-direction slide block 106 and the first small slide block 109, second small slide block 116 and the little guide rail 117 of the first little guide rail 110, second, described X direction guiding rail 121 is located on base plate 101, described Z-direction guide rail 118 is fixed in X on slide block 120, described Y-direction guide rail 107 is fixed on Z-direction slide block 106, described first little guide rail 110 and the second little guide rail 117 are arranged on the first Y-direction slide block 111 and the second Y-direction slide block 112 respectively, described many nozzle components are at least containing six shower nozzles, comprise at least two squash type shower nozzles and four spray-type shower nozzles, points two groups are separately fixed on two parallel Y-direction slide blocks, first spray-type shower nozzle 104, second spray-type shower nozzle 105 and the 3rd spray-type shower nozzle 108 are arranged on the first Y-direction slide block 111, wherein the first spray-type shower nozzle 104 and the second spray-type shower nozzle 105 are arranged in the shower nozzle holddown groove on the first Y-direction slide block 111, and the 3rd spray-type shower nozzle 108 is arranged on the shower nozzle holddown groove of the first small slide block 109, 4th spray-type shower nozzle 115 and at least two squash type shower nozzles, first squash type shower nozzle 113 and the second squash type shower nozzle 114, be arranged on the second Y-direction slide block 112, wherein the first squash type shower nozzle 113 and the second squash type shower nozzle 114 are fixed on the shower nozzle holddown groove of the second Y-direction slide block 112, 4th spray-type shower nozzle 115 is arranged on the shower nozzle holddown groove of the second small slide block 116, the nozzle inside diameter of the first squash type shower nozzle 113 and the second squash type shower nozzle 114 be respectively 200 μ m ?1mm and 50 μ m ?500 μm, the mesh diameter of four spray-type spray nozzles be 50 μ m ?100 μm, described forming device comprises forming containers 102, support 103 and motor 119, forming containers 102 is arranged on immediately below base plate 101 upper nozzle assembly, removable support 103 to be arranged in forming containers 102 and to be connected with the output shaft of motor 119, described support 103 for having the solid material to cytotoxic of certain mechanical strength, as rustless steel or titanium alloy, described control system is responsible for controlling the D translation motion of nozzle component and the rotary motion of support.
Fig. 2 is the three dimensional structure sketch of the first Y-direction slide block, described first Y-direction slide block has two holes being used for fixing shower nozzle and one and can allow the cephalomotor macropore of spray, macropore both sides are provided with the first little guide rail 110, first small slide block 109 can along little guide rail movement, drive the 3rd spray-type shower nozzle 108 on it to be that the first spray-type shower nozzle 104 second spray-type shower nozzle 105 works in turn, the second Y-direction slide block structure and the first Y-direction slide block similar.
Fig. 3 is the flow chart utilizing rotation method of piling formation tissue organ of the present invention, mainly comprises the three large key steps such as the preparation before shaping, histoorgan forming precursor and subsequent operation.
Preparation before shaping comprises the preparation of material and the making of inner core, and preparation 0.2 ~ 20% (w/v) hydrogel solution, 1 ~ 10% (w/v) cross-linking agent/polymer fluid, 0.1 ~ 10% (w/v) synthesize macromolecular solution, the gelatin solution of 0.2 ~ 10% (w/v) and cell culture fluid; According to the type of the histoorgan that will be shaped, the autologous stem cells with differentiation capability is separated from patient, as fat stem cell, embryonic stem cell, blood stem cell, bone marrow stem cell, induction type versatile stem cell, or there is physiologically active adult cell, if adipose cell, hepatocyte, bladder cells, lymphocyte, myocardial cell or nephrocyte are as seed cell, and make cell suspension, seed cell density is 1 × 10
3~ 1 × 10
8individual/mL, is 1 ~ 9:9 ~ 1 Homogeneous phase mixing by volume by the hydrogel solution after seed cell suspension and sterilizing, is prepared into the aqueous gel mixture containing seed cell; Described hydrogel be can composite gelatin, collagen, matrigel, carrageenan, chitosan, agar, hyaluronic acid, matrigel, elastin laminin, one or more sodium alginate or fibrinogen solution in laminin, in described hydrogel can the various cell cryopreservation agent of compound, cell growth factor, medicine, anticoagulant.Cell cryopreservation agent is one or more complex in dimethyl sulfoxide (DMSO), glycerol, dextrose.Described cell growth factor is that endothelial cell growth factor (ECGF) is (as one or more complex in VEGF (VEGF), basic fibroblast growth factor (b-FGF), hepatocyte growth factor (HGF), human blood platelets derived growth factor (PDGF-BB), transforminggrowthfactor-β1 (TGF-β 1).Described medicine be antitumor drug (as astragalus polysaccharides, cis diamino dichloro network platinum (DDP), mitomycin (MMC), one or more complex in 5-fluorouracil (5-FU), Claritin, antibiotic (as penicillin, gibberellins, tetracycline), viral vaccine.Described anticoagulin is one or more complex in heparin, paclitaxel; The solute of described synthesis macromolecular solution be specially polyurethane (PU), polycaprolactone (PCL), Merlon, Polyethylene Glycol, Ju Ru Suan ?one or more in co-glycolic acid (PLGA), polyester and polyhydroxy acid ester, solvent is TEG or 1,4 ?dioxane; Take out the support 103 on forming containers 102 and carry out sterilization treatment, when there is the histoorgan of inner chamber when being shaped, according to the construction features of the histoorgan inner chamber that will be shaped, with its threedimensional model of three-dimensional drawing software design, make the mould with this inner-cavity structure by quick shaping method, support 103 is stood in mould, in mould, adds gelatin solution, reducing temperature to 10 ~ 12 DEG C makes gelatin solidify, and obtains the histoorgan inner core of center belt supporting frame; As shaping solid tissue organ, inner core is the cylindrical sleeves of thick 5 ~ 6mm; Then apply one deck hydrogel outward at the histoorgan inner core of belt supporting frame, and carry out being cross-linked/being polymerized with cross-linking agent/polymer fluid.
Fig. 4 is rotation stack shaping process schematic of the present invention, to be respectively charged in equipment in the first squash type shower nozzle 113 and the second squash type shower nozzle 114 containing the hydrogel of seed cell and not celliferous hydrogel, cross-linking agent/polymer fluid, neurocyte suspension and endotheliocyte suspension are respectively charged in the 4th spray-type shower nozzle 115, second spray-type shower nozzle 105 and the first spray-type shower nozzle 104, synthesis macromolecular solution loads in the 3rd spray-type shower nozzle shower nozzle 108, support 103 to be arranged in forming containers 102 and forming containers 102 in loading cell culture fluid 405, after the model of histoorgan to be formed is imported control system, start former, start stack shaping, in forming process, support 103 rotates around Y negative sense, shower nozzle does translational motion, when shaping histoorgan main part, first squash type shower nozzle 113 moves to the hydrogel that assigned address extrudes containing seed cell and is wrapped on inner core, and the dead astern that the 4th spray-type shower nozzle 115 that cross-linking agent/polymer fluid is housed simultaneously moves to the first squash type shower nozzle 113 sprays cross-linking agent/polymer fluid and is cross-linked it/is polymerized, when shaping blood vessel, second squash type shower nozzle 114 moves to assigned address and extrudes not celliferous hydrogel and be wrapped on inner core, 4th spray-type shower nozzle 115 moves to the second squash type shower nozzle 114 dead astern simultaneously, first spray-type shower nozzle 104 and the 3rd spray-type shower nozzle 108 move to the second squash type shower nozzle 114 dead ahead respectively, under the inner core rotated drives, the hydrogel of new winding moves to the 4th spray-type shower nozzle successively to carry out for 115 times being cross-linked/being polymerized, wash through culture fluid, through the first spray-type shower nozzle 104, 3rd spray-type shower nozzle 108 times carries out sprinkling endotheliocyte and composite synthesis macromolecule, during as shape nerve fibre bundle, second squash type shower nozzle 114 moves to assigned address and extrudes not celliferous hydrogel and be wrapped on inner core, 4th spray-type shower nozzle 115 moves to the first squash type shower nozzle 113 dead astern simultaneously, second spray-type shower nozzle 105 and the 3rd spray-type shower nozzle 108 move to the second squash type shower nozzle 114 dead ahead respectively, under the inner core rotated drives, the hydrogel of new winding moves to the 4th spray-type shower nozzle successively to carry out for 115 times being cross-linked/being polymerized, wash through culture fluid, through the second spray-type shower nozzle 105, 3rd spray-type shower nozzle 108 times carries out sprinkling neurocyte and composite synthesis macromolecule, so repeatedly be wound around accumulation, until the histoorgan needed for being shaped.
Fig. 5 a is not celliferous hydrogel; Fig. 5 b is for spraying the hydrogel after endothelium (nerve) cell; Fig. 5 c is the hydrogel after spraying synthesis macromolecular solution; Time shaping blood vessel (nerve), the not celliferous water-setting collodion silk sprayed from shower nozzle is after crosslinked/polymerization and washing, be sprayed one deck endothelium (nerve) cell and synthesis macromolecular solution successively, formed Ning Jiao ?Xi Bao ?membrane structure, in follow-up incubation, hydrogel is hydrolyzed gradually, and endotheliocyte or nerve growth also connect each other, finally forms blood vessel and neuromechanism.
Fig. 6 is the structural representation of histoorgan provided by the invention, and described histoorgan is revolving body or nearly revolving body, is made up of inner chamber 603 and peripheral entity; Described inner chamber 603 is closed surface shape that is cylindric, spherical or that surrounded by other curved surfaces; Described peripheral entity by containing or not celliferous water-setting collodion silk be wound around accumulation form; Water-setting collodion silk 602 containing seed cell is wound agent structure, not celliferous water-setting collodion silk 603 surface-coated endotheliocyte and synthesis macromolecular solution form vascular system, and not celliferous hydrogel surface coating neurocyte and synthesis macromolecular solution form nervous system.
Enumerate several specific embodiment below, to understand the present invention further:
The preparation in embodiment 1 atrium
(1) material preparation
Extract fat stem cell (ADSCs), endotheliocyte (ECs), neurocyte, sternzellen and myocardial cell (CMCs), subculture, obtains cell material, and endotheliocyte and neurocyte concentration are 8 × 10
7individual/mL, sternzellen and myocardial cell density are 2 × 10
6individual/mL; Get 2g sodium alginate and 7g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, in this mixed liquor, add the cells frozen storing liquid DMSO of mass body volume concentrations 3%, sterilization treatment obtain Ming Jiao ?Sodium Alginate Hydrogel Films material; Get 4g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtains gelatin solution; The calcium chloride water of preparation 0.05g/ml is as its cross-linking agent/polymer fluid; Be dissolved in by degradable polycarbonate in TEG and obtain the synthesis macromolecular solution that mass body volume concentrations is 2%, sterilization treatment is stand-by; Prepare cell culture fluid DMEM and 1% collagen solution.By the mixed liquor of sternzellen and myocardial cell and hydrogel with volume ratio be 1:1 be mixed to get containing Xin Ji ?the hydrogel solution of sternzellen.
(2) inner core preparation
According to the shape and structure feature of atrium inner chamber, with the three-dimensional fusiform model of three-dimensional drawing software design, the mould with this inner-cavity structure is made by quick shaping method, support after sterilizing is stood in mould, in mould, add gelatin solution, reduce temperature to 10 ~ 12 DEG C and gelatin is solidified, form the atrium inner core of center belt supporting frame, and apply one deck collagen solution outward at the atrium inner core of belt supporting frame, and collagen is made to solidify to form the protecting film of the water insoluble solution of one deck;
(3) be shaped
By containing Xin Ji ?the hydrogel of sternzellen and DMSO and not celliferous hydrogel be respectively charged into the first squash type shower nozzle 113 and the first squash type shower nozzle 114, endotheliocyte suspension and neurocyte suspension are respectively charged into the first spray-type shower nozzle 104, second spray-type shower nozzle 105, calcium chloride water loads the 4th spray-type shower nozzle 115, synthesis macromolecular solution loads the 3rd spray-type shower nozzle 108, to be with the support installing of gelatin inner core in forming containers 102 and be connected with motor output shaft, the cell culture fluid of 100mL is loaded in forming containers 102, whole device is placed in the environment that temperature is 8 ~ 10 DEG C, forming device is started after atrium model is imported control system, start to be shaped.
(4) molten core
After shaping, the medium-term and long-term preservation of liquid nitrogen will be placed in containing the atrium precursor of gelatin inner core or calorstat is cultivated, melt except inner core, take out support, namely obtain the atrium precursor being with inner chamber.
(5) later stage cultivates
Atrium precursor is cultivated as in pulsation containers for culturing organisms, in incubation, under the multiple action of stress field, iuntercellular is impelled to connect and induced growth, endothelium (nerve) cell induction around not celliferous gelatine silk is differentiated to form the blood vessel (nerve) that internal diameter is about 1mm, after gel degradation, final formation has the atrium containing blood vessel and nerve of spontaneous contractions ability.This atrium can directly be attached on disease damage heart, reaches the object of reparative regeneration.
The preparation of embodiment 2 liver precursor
(1) material preparation
Extract human bone marrow stem cell (BSCs), fat stem cell (ADSCs), endotheliocyte (ECs), neurocyte and hepatocyte, subculture, obtains seed cell; Get 1 Fibrinogen and 7g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, wherein add the antitumor drug astragalus polysaccharides of 0.1%, sterilization treatment obtain Xian fibrillarin Yuan ?Ming Jiao ?astragalus polysaccharides hydrogel material; Get 4g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtains gelatin solution; The thrombin PBS polymer fluid of preparation 0.05g/ml; Be dissolved in by PU in TEG and obtain the synthesis macromolecular solution that mass body volume concentrations is 5%, sterilization treatment is stand-by;
The bone marrow of above-mentioned preparation is done thin born of the same parents ?hepatocyte and the former ?of fine fibrillarin bright glue ?astragalus polysaccharides hydrogel material is mixed to get fat stem cell and hepatocellular density is 1 × 10
6individual/mL containing Gan Xi Bao ?the hydrogel of fat stem cell.Endotheliocyte, fat stem cell and 0.1% heparin phosphate buffer (PBS) are mixed to get concentration and are 5 × 10
7the Nei Pi of individual/mL ?fat stem cell suspension.Neurocyte and fat stem cell are mixed to get concentration and are 5 × 10
7the Shen Jing of individual/mL ?fat stem cell suspension.
(2) inner core preparation
Rack surface is after sterilization coated with the gelatin that a layer thickness is about 5mm, reduces temperature to 10 ~ 12 DEG C and solidify to form inner core, apply hydrogel outward and cross-linked polymeric forms the protecting film that one deck is insoluble to culture fluid at the liver precursor inner core of belt supporting frame.
(3) be shaped
By containing Gan Xi Bao ?bone marrow stem cell Xian fibrillarin Yuan ?Ming Jiao ?astragalus polysaccharides hydrogel and not celliferous hydrogel be respectively charged into the first squash type shower nozzle 113 and the first squash type shower nozzle 114 Nei Pi ?Zhi fat Gan Xi Bao ?heparin suspension and Shen Jing ?fat stem cell suspension be respectively charged into the first spray-type shower nozzle 104 and the second spray-type shower nozzle 105, thrombin polymer fluid loads the 4th spray-type shower nozzle 115, and synthesis macromolecule loads the 3rd spray-type shower nozzle 108.To be with the support installing of gelatin inner core in forming containers 102 and be connected with motor output shaft, and in forming containers, load the cell culture fluid of 100mL.Whole device is placed in the environment that temperature is 10 DEG C, starts forming device and start to be shaped before importing liver after body Model.
(4) molten core
After shaping, the liver precursor containing gelatin inner core is placed in the isoperibol of about 37 DEG C, makes gelatin inner core " thawing ", take out support, namely obtain liver precursor.
(5) later stage cultivates
Liver precursor is placed in incubator cultivate, in incubation, iuntercellular connects and proliferate, endothelium (nerve) cell around not celliferous gelatine silk and the differentiation-inducing formation internal diameter of fat stem cell are about the blood vessel (nerve) of 1mm, after gel degradation, finally form liver.
The preparation of embodiment 3 bladder precursor
(1) material preparation
Extract body fat stem cell (ADSC), endotheliocyte (EC), neurocyte and bladder cells, subculture, obtains cell material; Get 1g sodium alginate, 1g chitosan and 1g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtain Hai Zao Suan Na ?Ke Ju Tang ?gelatin hydrogel material; The calcium chloride water of preparation 0.05g/ml is as its cross-linking agent; Get 2g glucose and 2g dextrose to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtain Pu Tao Tang ?dextrose solution; Be dissolved in by PLGA in TEG and obtain the solution that volumetric concentration is 5%, sterilization treatment is stand-by;
By the Zhi fat Gan Xi Bao of above-mentioned preparation ?bladder cells and Hai Zao Suan Na ?Ke Ju Tang ?the gelatin hydrogel material mixing density that obtains fat stem cell and bladder cells be 1 × 10
6individual/mL containing bladder Xi Bao ?the hydrogel of fat stem cell; Endotheliocyte and fat stem cell are mixed to get concentration and are 5 × 10
7the Nei Pi of individual/mL ?fat stem cell suspension; Neurocyte and fat stem cell are mixed to get concentration and are 5 × 10
7the Shen of individual/mL through Xi Bao ?fat stem cell suspension.
(2) inner core preparation
According to the shape and structure feature of bladder inner chamber, by the ellipse chondritic of three-dimensional drawing software design, make the mould with this inner-cavity structure by quick shaping method.Support after sterilizing is stood in mould; appropriate gelatin solution is added in mould; reducing temperature to 10 ~ 12 DEG C makes gelatin solidify; the histoorgan inner core of formation center belt supporting frame, applies hydrogel outward and cross-linked polymeric forms the protecting film that one deck is insoluble to culture fluid at the liver precursor inner core of belt supporting frame.
(3) be shaped
By containing bladder Xi Bao ?fat stem cell Hai Zao Suan Na ?Ke Ju Tang ?gelatin hydrogel and not celliferous hydrogel be respectively charged into the first squash type shower nozzle 113 and the first squash type shower nozzle 114, Nei Pi Xi Bao ?fat stem cell suspension and Shen through Xi Bao ?fat stem cell suspension be respectively charged into the first spray-type shower nozzle 104 and the second spray-type shower nozzle 105, calcium chloride cross-linking agent loads the 4th spray-type shower nozzle 115, and synthesis macromolecule loads the 3rd spray-type shower nozzle 108.To be with the support installing of gelatin inner core in forming containers 102 and be connected with motor output shaft, and in forming containers, load the cell culture fluid of 100mL.Whole device is placed in the environment that temperature is 8 ~ 10 DEG C, starts forming device after importing bladder model in a computer, starts to be shaped.
(4) molten core
After shaping, the bladder precursor containing gelatin inner core is placed in calorstat and cultivates, remove inner core, take out support, namely obtain the bladder precursor being with inner chamber.
(5) later stage cultivates
Bladder precursor is placed in calorstat cultivate, in incubation, iuntercellular connects and proliferate, endothelium (nerve) cell around not celliferous gelatine silk and the differentiation-inducing formation blood vessel (nerve) of fat stem cell, after gel degradation, final formation has bladder that is neural and blood vessel.
The preparation of embodiment 4 kidney precursor
(1) material preparation
Extract blood of human body stem cell, endotheliocyte (EC), neurocyte and nephrocyte, subculture, obtains cell material; Get 1g sodium alginate and 1g hyaluronic acid to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtain Hai Zao Suan Na ?hyaluronic acid gel material; The calcium chloride water of preparation 0.05g/ml is as its cross-linking agent; Get 4g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtains gelatin solution; Be dissolved in TEG by synthesis macromolecule PU and obtain the synthesis macromolecular solution that mass body volume concentrations is 5%, sterilization treatment is stand-by;
By the blood Gan Xi Bao of above-mentioned preparation ?nephrocyte and Hai Zao Suan Na ?the hyaluronic acid gel material mixing density that obtains blood stem cell and nephrocyte be 1 × 10
6individual/mL containing Shen Xi Bao ?the hydrogel of blood stem cell.Endotheliocyte and blood stem cell are mixed to get concentration and are 4 × 10
7the endotheliocyte blood stem cell suspension of individual/mL.Neurocyte and blood stem cell are mixed to get concentration and are 4 × 10
7the Shen of individual/mL through Xi Bao ?blood stem cell suspension.
(2) inner core preparation
Be coated with thick layer at rack surface and be about the gelatin of 5mm, reduce temperature to 10 ~ 12 DEG C and solidify to form inner core, apply hydrogel outward and cross-linked polymeric forms the protecting film that one deck is insoluble to culture fluid at the liver precursor inner core of belt supporting frame.
(3) be shaped
By containing Shen Xi Bao ?the hydrogel of blood stem cell and not celliferous hydrogel be respectively charged into the first squash type shower nozzle 113 and the second squash type shower nozzle 114, Nei Pi Xi Bao ?blood stem cell solution and Shen through Xi Bao ?blood stem cell solution be respectively charged into the first spray-type shower nozzle 104 and the second spray-type shower nozzle 105, calcium chloride cross-linking agent/polymer fluid loads the 4th spray-type shower nozzle 115, and synthesis macromolecule loads the 3rd spray-type shower nozzle 108.To be with the support installing of gelatin inner core in forming containers 102 and be connected with motor output shaft, and in forming containers, load the cell culture fluid of 100mL.Whole device is placed in the environment that temperature is 8 ~ 10 DEG C, starts forming device, start to rotate stack shaping after importing kidney model.
(4) molten core
After shaping, the kidney precursor containing gelatin film inner core is placed in calorstat and cultivates, make gelatin film inner core " thawing ", take out support, namely obtain kidney precursor.
(5) later stage cultivates
Kidney precursor is placed in incubator cultivate, in incubation, iuntercellular connects and proliferate, endothelium (nerve) cell around not celliferous gelatine silk and the differentiation-inducing formation internal diameter of blood stem cell are about the blood vessel (nerve) of 1mm, after gel degradation, final formation has blood vessel and neural kidney.
The preparation of embodiment 5 breast prosthesis
(1) material preparation
Extract body fat stem cell (ADSC), endotheliocyte (EC), neurocyte and lymphocyte, subculture, obtains cell material; Get 1g sodium alginate and 2g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtain Hai Zao Suan Na ?gelatin hydrogel material; The calcium chloride water of preparation 0.05g/ml is as its cross-linking agent/polymerization; Get 2g gelatin to be placed in 100mL culture fluid and to make it dissolve completely, sterilization treatment obtains gelatin solution; Be dissolved in TEG by synthesis macromolecule PU and obtain the synthesis macromolecular solution that mass body volume concentrations is 5%, sterilization treatment is stand-by;
By the fat stem cell of above-mentioned preparation and Hai Zao Suan Na ?gelatin hydrogel material mixing obtain fat stem cells density and be 1 × 10
6the hydrogel of the fatty stem cell of individual/mL.Wherein add 1 μM of insulin, 1 μM of dexamethasone, 50u/mL aprotinin and 0.5mmol/L isobutyl methyl xanthine derivative.Neurocyte and fat stem cell are mixed to get concentration and are 3 × 10
8the Shen of individual/mL through Xi Bao ?fat stem cell suspension.Add in fat stem cell suspension 50ng/mL endothelial cell growth factor (ECGF) (VEGF), 3ng/mL TGF ?β 1 somatomedin, 10ng/mL fibroblast growth factor (b ?FGF).
(2) inner core preparation
Be coated with thick layer at rack surface and be about the gelatin of 5mm, reduce temperature to 10 ~ 12 DEG C and solidify to form inner core, apply hydrogel outward and cross-linked polymeric forms the protecting film that one deck is insoluble to culture fluid at the liver precursor inner core of belt supporting frame.
(3) be shaped
By the Hai Zao Suan Na of fatty stem cell, insulin, dexamethasone, aprotinin and isobutyl methyl xanthine derivative ?gelatin hydrogel and not celliferous hydrogel be respectively charged into the first squash type shower nozzle 113 and the second squash type shower nozzle 114, Nei Pi Xi Bao ?fat stem cell suspension and Shen through Xi Bao ?fat stem cell suspension be respectively charged into the first spray-type shower nozzle 104 and the second spray-type shower nozzle 105, calcium chloride cross-linking agent loads the 4th spray-type shower nozzle 115, and synthesis macromolecule loads the 3rd spray-type shower nozzle 108.To be with the support installing of gelatin inner core in forming containers 102 and be connected with motor output shaft, and in forming containers, load the cell culture fluid of 100mL.Whole device is placed in the environment that temperature is 10 ~ 12 DEG C, starts forming device after importing breast model, starts to be shaped.
(4) molten core
After shaping, the breast precursor containing gelatin film inner core is placed in calorstat and cultivates, make gelatin inner core " thawing ", take out support, namely obtain breast precursor.
(5) later stage cultivates
In incubation, iuntercellular connects and proliferate, endothelium (nerve) cell around not celliferous gelatine silk and the differentiation-inducing formation internal diameter of fat stem cell are about the blood vessel (nerve) of 1mm, after gel degradation, final formation has the breast of certain function.
Claims (7)
1. utilize the special equipment rotating method of piling and prepare histoorgan, comprise bracing or strutting arrangement, many nozzle components, motion, forming device and control system; Described bracing or strutting arrangement comprises supporting leg and base plate (101); Described motion adopt Dao Gui ?slide block mechanism, comprise X direction guiding rail (121), X to slide block (120), Y-direction guide rail (107), the first Y-direction slide block (111), the second Y-direction slide block (112), Z-direction guide rail (118), Z-direction slide block (106); Described X direction guiding rail (121) is located on base plate (101), and Z-direction guide rail (118) is fixed in X on slide block (120), and Y-direction guide rail (107) is fixed on Z-direction slide block (106); It is characterized in that: described many nozzle components, at least containing six shower nozzles, divide two groups and are arranged on the first Y-direction slide block and the second Y-direction slide block respectively; Described forming device comprises forming containers (102), support (103) and motor (119), forming containers (102) is arranged on base plate (101) and goes up and be in the below of nozzle component, and support (103) to be arranged in forming containers (102) and to be connected with motor (119) output shaft.
2. as claimed in claim 1 utilize the special equipment rotating method of piling and prepare histoorgan, it is characterized in that: described support is two ends be cylindric, middle is prismatic structure.
3. the special equipment utilizing rotation method of piling to prepare histoorgan as claimed in claim 1, it is characterized in that, the material of described support is rustless steel or titanium alloy.
4. the special equipment utilizing rotation method of piling to prepare histoorgan as claimed in claim 1, it is characterized in that: described first Y-direction slide block (111) and the second Y-direction slide block (112) have and first of Y-direction guide rail parallel the little guide rail (110) and the second little guide rail (117), first little guide rail and the second little guide rail install the first small slide block (109) and the second small slide block (116) respectively, the first small slide block and the second small slide block has the hole of stationary nozzle.
5. the special equipment utilizing rotation method of piling to prepare histoorgan as claimed in claim 1, it is characterized in that: described many nozzle components comprise at least two squash type shower nozzles and four spray-type shower nozzles, the nozzle inside diameter of two squash type shower nozzles is respectively 200 μm ~ 1mm and 50 μm ~ 500 μm, and the mesh diameter of spray-type spray nozzle is 50 μm ~ 100 μm.
6. utilize the method rotating method of piling and prepare histoorgan, it is characterized in that the method comprises the following steps:
1) material preparation:
Preparation 0.2 ~ 20% (w/v) hydrogel solution, 1 ~ 10% (w/v) cross-linking agent/polymer fluid, 0.1 ~ 10% (w/v) synthesize macromolecular solution, 0.2 ~ 10% (w/v) gelatin solution and cell culture fluid; Be separated or induction seed cell, seed cell density is 1 × 10
3~ 1 × 10
6individual/mL, is mixed and made into the hydrogel solution containing seed cell by seed cell and hydrogel solution by 1 ~ 9:9 ~ 1 volume ratio; Be separated or inducing endothelial cell and neurocyte make cell suspension, endotheliocyte suspension and neurocyte suspension density are 1 × 10
5~ 1 × 10
8individual/mL;
2) inner core preparation
When preparation has the histoorgan of inner chamber, according to the cavity shape will preparing histoorgan, with its threedimensional model of three-dimensional drawing software design, make the mould with this inner-cavity structure by quick shaping method, when the histoorgan of forming solid, mould is the cylinder of the thick 8 ~ 10mm of diameter; Support (103) is put into mould central authorities, reducing temperature to 8 ~ 10 DEG C add gelatin solution in mould after makes gelatin solidify, open the histoorgan inner core (401) that mould obtains center belt supporting frame (103), apply one deck hydrogel solution outward at the histoorgan inner core (401) of belt supporting frame, and be cross-linked/polymerization with cross-linking agent/polymer fluid;
3) be shaped
The hydrogel solution and not celliferous hydrogel solution that contain seed cell are respectively charged in the first squash type shower nozzle (113) and the second squash type shower nozzle (114), cross-linking agent/polymer fluid, neurocyte suspension, endotheliocyte suspension and synthesis macromolecular solution are respectively charged into the 4th spray-type shower nozzle (115), second spray-type shower nozzle (105), in first spray-type shower nozzle (104) and the 3rd spray-type shower nozzle (108), cell culture fluid (405) is loaded in forming containers, support (103) to be arranged in forming containers (102) and to be connected with motor (119) output shaft, after the model of histoorgan to be formed is imported control system, start former, start stack shaping, in forming process, support (103) rotates around Y negative sense, shower nozzle does translational motion, when the main part of shaping histoorgan, first squash type shower nozzle (113) moves to the hydrogel that assigned address extrudes containing seed cell and is wrapped on inner core, and the dead astern that the 4th spray-type shower nozzle (115) that cross-linking agent/polymer fluid is housed simultaneously moves to the first squash type shower nozzle (113) sprays cross-linking agent/polymer fluid and is cross-linked it/is polymerized, when shaping blood vessel, second squash type shower nozzle (114) moves to assigned address and extrudes not celliferous hydrogel and be wrapped on inner core, 4th spray-type shower nozzle (115) moves to the second squash type shower nozzle (114) dead astern simultaneously, first spray-type shower nozzle (104) and the 3rd spray-type shower nozzle (108) move to the second squash type shower nozzle (114) dead ahead respectively, under the inner core rotated drives, carry out being cross-linked/being polymerized under the hydrogel of new winding moves to the 4th spray-type shower nozzle (115) successively, wash through culture fluid, to the first spray-type shower nozzle (104), sprinkling endotheliocyte and composite synthesis macromolecular solution is carried out under 3rd spray-type shower nozzle (108), as body and the spirit through time, second squash type shower nozzle (114) moves to assigned address and extrudes not celliferous hydrogel and be wrapped on inner core, 4th spray-type shower nozzle (115) moves to the second squash type shower nozzle (114) dead astern simultaneously, second spray-type shower nozzle (105) and the 3rd spray-type shower nozzle (108) move to the second squash type shower nozzle (114) dead ahead respectively, under the inner core rotated drives, carry out being cross-linked/being polymerized under the hydrogel of new winding moves to the 4th spray-type shower nozzle (115) successively, wash through culture fluid, to the second spray-type shower nozzle (105), sprinkling neurocyte and composite synthesis macromolecule is carried out under 3rd spray-type shower nozzle (108), so repeatedly be wound around and pile up until form the histoorgan precursor containing inner core,
4) subsequent treatment
Cultivating being placed in calorstat containing the histoorgan precursor of inner core, moltenly taking out support except after inner core, form revolution shape histoorgan after continuing to cultivate a period of time, neural and blood vessel (502) spirally runs through wherein.
7. a kind of method utilizing rotation method of piling to prepare histoorgan as claimed in claim 6, it is characterized in that: described seed cell is have the stem cell of differentiation capability or have the adult cell of physiologically active, described stem cell is fat stem cell, embryonic stem cell, blood stem cell, bone marrow stem cell or induction type versatile stem cell, and described adult cell is adipose cell, hepatocyte, bladder cells, lymphocyte, myocardial cell or nephrocyte; Described hydrogel is the sodium alginate or the fibrinogen solution that to be compounded with in gelatin, collagen, matrigel, carrageenan, chitosan, agar, hyaluronic acid, matrigel, elastin laminin in laminin one or more; The various cell cryopreservation agent of compound, cell growth factor, medicine, anticoagulant in described hydrogel; Cell cryopreservation agent is one or more complex in dimethyl sulfoxide, glycerol, dextrose; Described cell growth factor is one or more complex in VEGF, basic fibroblast growth factor, hepatocyte growth factor, human blood platelets derived growth factor, transforminggrowthfactor-β1; Described medicine is antitumor drug, and this antitumor drug is one or more complex in astragalus polysaccharides, cis diamino dichloro network platinum, mitomycin, 5-fluorouracil, Claritin, antibiotic and viral vaccine; Described anticoagulin is one or both complex in heparin and paclitaxel; The solute of described synthesis macromolecular solution is one or more in polyurethane, polycaprolactone, Merlon, Polyethylene Glycol, Poly(D,L-lactide-co-glycolide, polyester and polyhydroxy acid ester, and solvent is TEG or Isosorbide-5-Nitrae-dioxane.
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| CN201410473055.4A CN104207859B (en) | 2014-09-16 | 2014-09-16 | Rotation method of piling is utilized to prepare method and the special equipment of histoorgan |
| PCT/CN2014/089769 WO2016041238A1 (en) | 2014-09-16 | 2014-10-29 | Method and dedicated device for preparing tissue and organ by using spin accumulation method |
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| CN201410473055.4A CN104207859B (en) | 2014-09-16 | 2014-09-16 | Rotation method of piling is utilized to prepare method and the special equipment of histoorgan |
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| CN104658395A (en) * | 2015-02-15 | 2015-05-27 | 清华大学 | Heart simulation structure as well as forming method and special mold thereof |
| CN105012050A (en) * | 2015-07-16 | 2015-11-04 | 清华大学 | Method and special mould for preparing tissue and organ precursor with multi-branch channels |
| CN106085851A (en) * | 2016-06-29 | 2016-11-09 | 浙江大学 | The cell co-culture model printed based on 3D and manufacture method |
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Also Published As
| Publication number | Publication date |
|---|---|
| CN104207859B (en) | 2016-09-28 |
| WO2016041238A1 (en) | 2016-03-24 |
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