CN103756955A - Personalized bionic composite structure as well as preparation method of composite structure and method for drug screening by using composite structure - Google Patents

Abstract

Translated fromChinese

一种个性化仿生复合结构及其制备和用于药物筛选的方法，涉及一种基于复合多喷头三维打印技术的仿生复合结构及其制备。该仿生复合结构包括高分子外壳、分支血管支架、功能组织区、血管组织区和高分子隔离层。功能组织区为含组织细胞的水凝胶层；血管组织区为含血管种子细胞的水凝胶层；功能组织区和血管组织区在空间交替排列；高分子隔离层间隔排布在功能组织区和血管组织区内，并将功能组织区和血管组织区分成多个小区域；分支血管支架包括动脉血管和静脉血管两部分。本发明通过复合多喷头3D打印技术制备带分支血管的组织器官仿生结构；水凝胶结构不必交联或聚合；最大限度的模拟了体内血管化组织和血管的状态，为血管化的器官组织提供了借鉴。

A personalized bionic composite structure and its preparation and drug screening method relate to a bionic composite structure based on composite multi-nozzle three-dimensional printing technology and its preparation. The biomimetic composite structure includes a polymer shell, a branch blood vessel stent, a functional tissue area, a blood vessel tissue area and a polymer isolation layer. The functional tissue area is a hydrogel layer containing tissue cells; the vascular tissue area is a hydrogel layer containing vascular seed cells; the functional tissue area and the vascular tissue area are arranged alternately in space; the polymer isolation layer is arranged at intervals in the functional tissue area and the vascular tissue area, and divide the functional tissue area and the vascular tissue area into multiple small areas; the branch vascular stent includes arterial blood vessels and venous blood vessels. The invention prepares bionic structures of tissues and organs with branched blood vessels through composite multi-nozzle 3D printing technology; the hydrogel structure does not need to be cross-linked or polymerized; it simulates the state of vascularized tissues and blood vessels in the body to the greatest extent, and provides for reference.

Description

The bionical composite structure of a kind of personalization and preparation thereof and for the method for drug screening

Technical field

The invention belongs to bioengineering field, specific to biology or enzyme, its compositions field, relate to the bionical composite structure of a kind of personalization and preparation thereof and for the method for drug screening.

Background technology

In recent years, along with interdisciplinary interchange contact is day by day tight, cell arrangement technology is arisen at the historic moment, for engineering foundation has been established in organizational project, organ manufacture and drug screening.Outstanding three-dimensional printing technology wherein; also be Quick-forming (Rapid Prototyping, RP) technology, utilize principle work material [the Fedorovich NE of Computer Aided Modeling and discrete-accumulation; et al.Trends in biotechnology, 2011; 29 (12): 601-606].This technology can accurately be located biomaterial, in the assembled in vitro field of complex organ, is paid much attention to.Tissue or the organ of external shaping are of many uses, can be used for Ji Li [Griffith LG, et al.Nature Reviews Molecular Cell Biology, 2006 that research maintains organ normal morphology; 7 (3): 211-224], cancer cells is evaded the treatment measures such as chemotherapy [Yamada KM, et al.Cell, 2007; 130 (4): 601-610], transplanting [Griffith LG, et al.Science Signaling, 2002 of diseased organ; 295 (5557): 1009] and drug screening.External many scientific research groups have completed the transformation of multiple RP technology, to realize assembling or the printing containing cell three-dimensional structure based on numerical control.Organ manufacturing center of department of mechanical engineering of domestic Tsing-Hua University (Center of Organ Manufacturing) is developing a series of rapid forming equipments; and utilize these equipment successfully to prepare [the Wang XH such as simple vasoganglion, liver organization and bone renovating material; et al.Trends in Biotechnology, 2007; 25:505; Wang XH, et al.Tissue Engineering Part B, 2010; 16:189; Wang XH.Artificial organs, 2012; 36:591].Cell/substrate material three-dimensional structure after assembling generally needs stability crosslinked or polymerization natural macromolecular material maintenance three-dimensional structure.

Since the nineties in last century, the exploitation of new drug mainly completes by the bioinformation detection platform based on molecule and cell levels, the single cell of planting that mainly utilizes external planar to cultivate.Yet two dimensional surface is cultivated the real-life three-dimensional environment that can not simulate the corresponding many cells of human body (at least three kinds) tissue completely.Adopting two-dimentional culturing cell is that the medicine overwhelming majority that target cell screens does not have activity [Kang XH, et al.Tissue Engineering, 2005 in animal body; 11:456-458].In recent years along with the microscale of biological study, the indices that biologists are increasing finds cells survival all has vital impact to the growth conditions of cell as oxygen concentration, existence temperature, and this also makes the in-vitro simulated of tissue and cultivates especially challenging.The development of combinatorial chemistry, genomics makes can be used for compound or the medicament categories sharp increase of screening; the development that forces the flux of drug screening to grow with each passing hour; confirmatory method in the past becomes no longer applicable; high-flux medicaments sifting (High throughput Screening; HTS) be therefore suggested and promote rapidly [Rodrigues AD.Pharmaceutical research, 1997; 4 (11): 1504-1510].But HTS technology adopts single two dimension of planting cell to cultivate, and structure function of cell is imperfect, the definite lead compound of this technology is the follow-up expection pharmacologically active of tool not.Adopt that a kind of new the high medicaments sifting model of the success ratio that more meets human structurology---High content screening (High Content Screening, HCS) technology determines that lead compound becomes the common recognition in drug research field.Cell need to be provided by the information that provides from various materials such as DNA, RNA, protein, meta-bolites and ions, and shows individual features; And the interaction on time and space of ion, meta-bolites, macromole and organoid is the basis that organism is realized complicated vital functions.Therefore; want to understand more clearly target spot institute's role in cell function of picking out; just need an instrument that can automatically extract the information that target spot expresses in cell; and this instrument; HCS technology [Giuliano KA; et al.Assay and drug development technologies, 2003; 1 (4): 565-577].With respect to HTS technology, HCS technology can realize many target spots and select.HCS is used the means such as fluorescently-labeled antibody, DNA probe, bio-ligand or bio-reactor to collect target spot in the intracellular impact on links such as cellular form variation, growth, differentiation, apoptosis, can in single test, obtain the bulk information about each components such as cell DNA, protein.With respect to take the HCS technology that orifice plate is unit, HTS the selection result is more accurate, has more biological significance.

Patent (application number 200910079726.8) relates to a kind of Vascularized fat depot and construction process thereof based on subregion, fat granule is wrapped in micro-capsule, the three-dimensional fatty tissue structure that obtains having communicating structure, structure is mainly fatty district and angiosomes.But the method needs a large amount of linking agents (as CaCl₂), can damage cytoactive; The drug main of the method will discharge by micro-capsule, and drug effect is difficult for being applied to total body.Patent (application number 200910155794.8) relates to a kind of High content screening method of Remedies for diabetes, by Rapid Prototyping technique, obtains celliferous three-dimensional structure, and the three-dimensional structure containing islet cells is carried out to drug screening.But the method is not containing vessel branch system, and vascular endothelial cell is only distributed in hydrogel surface.The method also needs a large amount of linking agents to be stable into body.

Existing organizational project cell/support complex structure body can be expressed the characteristic of tissue or organ in certain dimension in vitro, but can not simulate the cellulous real structure containing vascular system completely.Hydrogel material used in the present invention can be cross-linked, because polymeric isolation layer and shell support for structure provides mechanics; Sealing coat and shell, can be grand (micro-) sight vesicular structure, have guaranteed the exchange of nutritive substance; The three-dimensional very big degree of blood vessel tissue has been simulated internal milieu; Branch vessel structure can be connected with the inside and outside recycle system, is convenient to HCS drug screening.These factors impel us to utilize compound many shower nozzles three-dimensional printing technology preparation simulate the state of intracorporeal organ and use it for drug screening containing the bionical composite structure of personalization of branch vessel.

Summary of the invention

The object of the invention is the bionical composite structure of a kind of personalization and preparation thereof and for the method for drug screening, make arteries and vein blood vessel state in its better analogue body, structure does not need to be cross-linked, cell density in structure can be very high, the present invention can be used as the state that bionical composite structure comes blood vessel tissue in analogue body, and then for drug screening.

The bionical composite structure of personalization, is characterized in that: the bionical composite structure of described personalization comprises polymer shell, branch vessel support, functional organization district, vascular tissue district and polymeric isolation layer; Described functional organization district is for containing histiocytic hydrogel layer; Described vascular tissue district is the hydrogel layer containing blood vessel seed cell; Arrange at space-alternating in described functional organization district and vascular tissue district; Described polymeric isolation layer is intervally arranged in functional organization district and vascular tissue district, and Bing Jiang functional organization district and vascular tissue are distinguished into a plurality of zonules; Described branch vessel support comprises arteries and vein blood vessel two portions, and arteries is containing at least one entrance and several branches, and vein blood vessel is containing at least one outlet and several branches; Described branch vessel support adopts synthesized polymer material, and branch vessel support is through between polymer shell, functional organization district, vascular tissue district and polymeric isolation layer; Described polymer shell adopts natural polymer or synthesized polymer material.

In technique scheme, the internal diameter of described branch vessel support is 10 μ m～10mm, and wall thickness is 100 μ m～2mm; The bed thickness of described polymeric isolation layer and polymer shell is 0.1 μ m～2mm.

The hydrogel layer containing blood vessel seed cell containing histiocytic hydrogel layer and vascular tissue district in functional organization of the present invention district is natural polymer hydrogel, and this natural polymer is at least one in gelatin, sodium alginate, Fibrinogen, collagen, matrigel, carrageenin, chitosan, agar, hyaluronic acid, matrigel, elastin and laminin; In described natural polymer hydrogel, be compounded with at least one in cell growth factor, heparin, leptin, taxol and Radix Astragali anti-cancer powder.

The mass body volume concentrations of natural polymer hydrogel of the present invention is 0.1～40%; Described blood vessel seed cell and histiocytic concentration are 1 * 10³～1 * 10⁸individual/mL; The histocyte in described functional organization district is a kind of in adult tissue's cell, adult stem cell and cancer cells; The blood vessel seed cell in described vascular tissue district is at least one in vascular endothelial cell, fat stem cell, mesenchymal stem cells MSCs, navel blood stem cell, bone marrow stem cell, embryonic stem cell and induced multi-potent stem cells.

The preparation method of the bionical composite structure of a kind of personalization provided by the invention, is characterized in that the method comprises the following steps:

1) design branch vessel support three-dimensional model, divides two groups and imitates respectively arteries and vein blood vessel;

2) by synthesized polymer material, prepare containing histiocytic hydrogel, being loaded in the different shower nozzles of compound many shower nozzles 3 D-printing equipment containing the hydrogel of blood vessel seed cell and the synthetic macromolecular solution preparing of preparing;

3) prepare branch vessel support: according to step 1) three-dimensional model, utilize compound many shower nozzles 3 D-printing equipment to extrude synthetic macromolecular solution, and freeze-drying removes organic solvent, or melt extrudes synthesized polymer material, obtain branch vessel support;

4) prepare functional organization district: utilize compound many shower nozzles 3 D-printing equipment successively to print to prepare containing histiocytic hydrogel, printing path is along branch vessel support, print at interval, obtains preliminary three-dimensional structure;

5) prepare vascular tissue district: utilize compound many shower nozzles 3 D-printing equipment successively to print, the hydrogel containing blood vessel seed cell preparing is filled in the space of preliminary three-dimensional structure to three-dimensional structure in the middle of obtaining;

6) prepare polymeric isolation layer and polymer shell: in step 4) and step 5) successively in print procedure, after printing one deck or some layers, utilize the synthetic macromolecular solution of compound many shower nozzles 3 D-printing equipment spraying and extract organic solvent, or melt extrude synthesized polymer material, form polymeric isolation layer, obtain the alternating structure body of celliferous hydrogel layer and polymeric isolation layer; In the peripheral spraying of alternating structure body, synthesize macromolecular solution and extract organic solvent, or melt extruding synthesized polymer material, forming polymer shell, obtaining personalized bionical composite structure precursor;

7) to the bionical composite structure precursor of above-mentioned personalization, provide somatomedin, realize the vascularization in vascular tissue district, make functional organization's district's formative tissue, the cell in Bing Shi vascular tissue district and functional organization district produces contact, finally obtains the described bionical composite structure of personalization.

In preparation method of the present invention, described compound many shower nozzles 3 D-printing equipment employing melt extrudes forming technique or low temperature depositing forming technique is prepared branch vessel support, polymeric isolation layer and polymer shell; When employing melt extrudes forming technique and prepares branch vessel support, polymeric isolation layer and polymer shell, described synthesized polymer material is at least one in acrylonitrile-butadiene-styrene (ABS), urethane, tetrafluoroethylene and carbohydrate; When adopting low temperature depositing forming technique to prepare branch vessel support, polymeric isolation layer and polymer shell, the solute of described synthetic macromolecular solution is at least one in urethane, polycarbonate, polyureas ester, PGA, poly-succinic ester, polyether ester, polyoxyethylene glycol, poly(lactic acid), polycaprolactone, Poly(D,L-lactide-co-glycolide, polyester and polyhydroxy acid ester, the solvent of described synthetic macromolecular solution is TEG or Isosorbide-5-Nitrae-dioxane.The mass body volume concentrations of described synthetic macromolecular solution is 1%～30%; In described synthetic macromolecular solution, be compounded with anticoagulin, this anticoagulin is heparin or taxol.

The bionical composite structure of a kind of personalization provided by the invention, for the method for drug screening, is characterized in that the method comprises the steps:

1) if the histocyte in functional organization district adopts adult tissue's cell or adult stem cell, the bionical composite structure of personalization is supplied with to the nutrient solution containing pathogenic medicine, make the bionical composite structure of above-mentioned personalization produce pathological symptom; If the histocyte in functional organization district adopts cancer cells, do not do pathogenic processing;

2) the bionical composite structure of personalization is supplied with to the nutrient solution containing different sorts and various dose curative drug, real-time or follow-up collection nutrient solution;

3) analyze the above-mentioned nutrient solution of having collected, the kind of obtaining medical treatment property medicine, the effect of dosage to described personalized composite structure, by detecting, contrast biological indicator, the effect of judgement medicine.

In the method for drug screening of the present invention, described biological indicator is at least one in carbohydrate lipid metabolism index, liver function index, renal function index, cancer cells index and vascularization index.Described pathological symptom is diabetes, organ inflammation, organ failure or tumour.

The present invention compared with prior art, has the technique effect of following advantage and high-lighting: 1. polymeric isolation layer of the present invention is divided into some communities by hydrogel layer, forms the structure that hydrogel layer and polymeric isolation layer are alternately arranged; Polymeric isolation layer supports and biochemical provide protection for hydrogel structure provides mechanics, can guarantee the shaping of low concentration of water gel; Hydrogel needn't be cross-linked, and cell density can be enough large.2. the formation cell communication in vascular tissue of the present invention district and functional organization district, alternately arrange in interval, and the state of simulation Liao Tinei blood vessel tissue and blood vessel, for the organ-tissue of vascularization provides reference.3. polymer shell of the present invention supports and biochemical provide protection for internal structure provides mechanics, and vivo and vitro favourable and described personalized biomimetic features is cultivated.4. branch vessel support outlet of the present invention can be connected with vascular system in external pulsation culture systems or body with entrance.5. the method for drug screening of the present invention can be analyzed the effect of medicine to the bionical composite structure of personalization in time, is beneficial to the drug screening work to metabolic trouble, organ disease or cancer.

Accompanying drawing explanation

Fig. 1 is the structural representation of personalized bionical composite structure.

Fig. 2 is the schematic diagram of branch vessel support.

Fig. 3 a and Fig. 3 b are respectively the schematic diagram in functional organization district and the vascular tissue district of printing.

Fig. 4 a and Fig. 4 b are respectively alternating structure and the explosive view thereof in polymeric isolation layer and functional organization district and vascular tissue district.

In figure: 101 – arterial blood entrances; 102 – branch vessel supports; 103– functional organization district; 104– vascular tissue district; 105 – venous blood outlets; 106 – polymer shells; 401 – polymeric isolation layers.

Embodiment

Below in conjunction with drawings and Examples, the present invention is further described.

Fig. 1 is the structural representation of personalized bionical composite structure, and the bionical composite structure of described personalization comprisespolymer shell 106,branch vessel support 102,functional organization district 103,vascular tissue district 104 andpolymeric isolation layer 401; Describedfunctional organization district 103 is for containing histiocytic hydrogel layer; Describedvascular tissue district 104 is the hydrogel layer containing blood vessel seed cell; Arrange at space-alternating in 103Hevascular tissue district 104, described functional organization district; Describedpolymeric isolation layer 401 is intervally arranged in 103He vascular tissue district,functional organization district 104, and 103He vascular tissue district, Bing Jiangfunctional organization district 104 is divided into a plurality of zonules; Describedbranch vessel support 102 comprises arteries and vein blood vessel two portions, and arteries is containing at least one entrance and several branches, and vein blood vessel is containing at least one outlet and several branches; Describedbranch vessel support 102 adopts synthesized polymer material, andbranch vessel support 102 is through betweenpolymer shell 106,functional organization district 103,vascular tissue district 104 andpolymeric isolation layer 401; Describedpolymer shell 106 adopts natural polymer or synthesized polymer material.

The internal diameter of described branch vessel support is 10 μ m～10mm, and wall thickness is 100 μ m～2mm; The bed thickness of described polymeric isolation layer and polymer shell is 0.1 μ m～2mm.Fig. 2 is branch vessel support schematic diagram, the hydrogel layer containing blood vessel seed cell containing histiocytic hydrogel layer and vascular tissue district in described functional organization district is natural polymer hydrogel, and this natural polymer is at least one in gelatin, sodium alginate, Fibrinogen, collagen, matrigel, carrageenin, chitosan, agar, hyaluronic acid, matrigel, elastin and laminin; In described natural polymer hydrogel, be compounded with at least one in cell growth factor, heparin, leptin, taxol and Radix Astragali anti-cancer powder.The mass body volume concentrations of described natural polymer hydrogel is 0.1～40%; Described blood vessel seed cell and histiocytic concentration are 1 * 10³～1 * 10⁸individual/mL; The histocyte in described functional organization district is a kind of in adult tissue's cell, adult stem cell and cancer cells; The blood vessel seed cell in described vascular tissue district is at least one in vascular endothelial cell, fat stem cell, mesenchymal stem cells MSCs, navel blood stem cell, bone marrow stem cell, embryonic stem cell and induced multi-potent stem cells.

The preparation method of the bionical composite structure of a kind of personalization provided by the invention comprises the following steps: 1) design branch vessel support three-dimensional model, divides two groups and imitates respectively arteries and vein blood vessel; 2) by synthesized polymer material, prepare containing histiocytic hydrogel, being loaded in the different shower nozzles of compound many shower nozzles 3 D-printing equipment containing the hydrogel of blood vessel seed cell and the synthetic macromolecular solution preparing of preparing; 3) prepare branch vessel support: according to step 1) three-dimensional model, utilize compound many shower nozzles 3 D-printing equipment to extrude synthetic macromolecular solution, and freeze-drying removes organic solvent, or melt extrudes synthesized polymer material, obtain branch vessel support; 4) prepare functional organization district: utilize compound many shower nozzles 3 D-printing equipment successively to print to prepare containing histiocytic hydrogel, printing path is along branch vessel support, as shown in Figure 3 a, print at interval, obtains preliminary three-dimensional structure; 5) prepare vascular tissue district: utilize compound many shower nozzles 3 D-printing equipment successively to print, as shown in Figure 3 b, the hydrogel containing blood vessel seed cell preparing is filled in the space of preliminary three-dimensional structure to three-dimensional structure in the middle of obtaining; 6) prepare polymeric isolation layer and polymer shell: in step 4) and step 5) successively in print procedure, after printing one deck or some layers, utilize the synthetic macromolecular solution of compound many shower nozzles 3 D-printing equipment spraying and extract organic solvent, or melt extrude synthesized polymer material, form polymeric isolation layer, obtain the alternating structure body of celliferous hydrogel layer and polymeric isolation layer; In the peripheral spraying of alternating structure body, synthesize macromolecular solution and extract organic solvent, or melt extruding synthesized polymer material, forming polymer shell, obtaining personalized bionical composite structure precursor; 7) to the bionical composite structure precursor of above-mentioned personalization, provide somatomedin, realize the vascularization in vascular tissue district, make functional organization's district's formative tissue, the cell in Bing Shi vascular tissue district and functional organization district produces contact, finally obtains the described bionical composite structure of personalization.

Preparation method described in this, described compound many shower nozzles 3 D-printing equipment employing melt extrudes forming technique or low temperature depositing forming technique is prepared branch vessel support, polymeric isolation layer and polymer shell; When employing melt extrudes forming technique and prepares branch vessel support, polymeric isolation layer and polymer shell, described synthesized polymer material is at least one in acrylonitrile-butadiene-styrene (ABS), urethane, tetrafluoroethylene and carbohydrate; When adopting low temperature depositing forming technique to prepare branch vessel support, polymeric isolation layer and polymer shell, the solute of described synthetic macromolecular solution is at least one in urethane, polycarbonate, polyureas ester, PGA, poly-succinic ester, polyether ester, polyoxyethylene glycol, poly(lactic acid), polycaprolactone, Poly(D,L-lactide-co-glycolide, polyester and polyhydroxy acid ester, the solvent of described synthetic macromolecular solution is TEG or Isosorbide-5-Nitrae-dioxane.The mass body volume concentrations of described synthetic macromolecular solution is 1%～30%; In described synthetic macromolecular solution, be compounded with anticoagulin, this anticoagulin is heparin or taxol.

The bionical composite structure of a kind of personalization provided by the invention comprises the steps: 1 for the method for drug screening) if the histocyte in functional organization district adopts adult tissue's cell or adult stem cell, the bionical composite structure of personalization is supplied with to the nutrient solution containing pathogenic medicine, make the bionical composite structure of above-mentioned personalization produce pathological symptom; If the histocyte in functional organization district adopts cancer cells, do not do pathogenic processing; 2) the bionical composite structure of personalization is supplied with to the nutrient solution containing different sorts and various dose curative drug, real-time or follow-up collection nutrient solution; 3) analyze the above-mentioned nutrient solution of having collected, the kind of obtaining medical treatment property medicine, the effect of dosage to described personalized composite structure, by detecting, contrast biological indicator, the effect of judgement medicine.

The method of drug screening of the present invention, described biological indicator is at least one in carbohydrate lipid metabolism index, liver function index, renal function index, cancer cells index and vascularization index.Described pathological symptom is diabetes, organ inflammation, organ failure or tumour.

Enumerate several specific embodiments below, further to understand the present invention.

Embodiment 1: prepare the bionical vascularization hepatic tissue of a kind of personalization and be applied to hepatitis medicament screening

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC) and liver cell (Hep), wherein prepare cell suspending liquid after ADSC subculture; By natural polymer powder and (DMEM, dulbecco's modified eagle medium) nutrient solution mixes, obtain mass body volume concentrations and be the fibrinogen solution that 10% gelatin solution, sodium alginate soln that mass body volume concentrations is 2% and mass body volume concentrations are 1%, will above-mentioned gelatin solution, sodium alginate soln and fibrinogen solution volume ratio 2:2:1 obtain natural polymer solution for later use after mixing; ADSC and Hep suspension is centrifugal, mix with natural polymer solution respectively, obtaining cell concn is 1 * 10⁶the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁶the natural polymer solution containing Hep of individual/mL.

2) preparation of branch vessel support: Solidworks software design branch vessel three-dimensional model, after delamination software is analyzed, by 3D printing technique melt extruded molten state tetrafluoroethylene (TEFLON), obtains synthetic fractionated polymer branch vessel support.

3) preparation of polymer shell and polymeric isolation layer: adopt equally 3D printing technique screw extruding molten state TEFLON to prepare shell and sealing coat; Shell 10 μ m thickness, separation layer thickness 0.5 μ m.

4) the final establishment of external bionic model: utilize compound many shower nozzles three-dimensional printing technology will be containing Hep natural polymer along branch vessel support printing-forming, 50% space be reserved at interval; Utilize compound many shower nozzles three-dimensional printing technology that the natural polymer containing ADSC is shaped along above-mentioned reserving space, molding inside can reserving hole channel; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; By the said structure vitro culture of pulsing, alternately change DMEM containing VEGF, containing DMEM and the general DMEM of HGF, realize hepatocellular organized stable, the development of vascularization and the contact between the two.

5) bringing out and drug screening of hepatitis: utilize pulsation system to supply with the DMEM induce drug hepatitis containing pathogenic medicine (as bisatin, methyldopa, Zoofurin etc.), destroy liver cell, detection record biological indicator (saccharase as amino in albumin, G-6-Pase and tyrosine and vascularization expression); The DMEM that utilizes pulsation system to supply with containing curative drug (as VITAMIN, reduced glutathion, Potenlini preparation or medicine newly developed) treats, and the above-mentioned biological indicator of record analysis, obtains drug screening result.

Embodiment 2: prepare the bionical vascularization hepatic tissue of a kind of personalization and be applied to hepatitis medicament screening

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC) and embryonic stem cell (ESC), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with DMEM nutrient solution, obtain mass body volume concentrations and be the fibrinogen solution that 20% gelatin solution, sodium alginate soln that mass body volume concentrations is 4% and mass body volume concentrations are 2%, more than state clearly after sol solution, sodium alginate soln and fibrinogen solution volume ratio 1:2:1 mix and obtain natural polymer solution for later use; ADSC and ESC suspension is centrifugal respectively, mix with natural polymer solution respectively, obtaining cell concn is 3 * 10⁶the natural polymer solution containing ADSC of individual/mL, and cell concn is 1 * 10⁵the natural polymer solution containing ESC of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) prepare natural polymer collagen branch vessel support.

3) step 3 of the preparation of polymer shell and polymeric isolation layer: embodiment 1) prepare natural polymer collagen shell and sealing coat.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of ESC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel, the CaCl that is 5% with mass body volume concentrations by molding₂the crosslinked 2min of thrombin solution of solution and 100U/mL; Utilize pulsation system to supply with stage by stage inductor and make ESC be converted into liver cell, inductor is containing aFGF, HGF, OSM, dexamethasone, Transferrins,iron complexes etc.; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; By the said structure vitro culture of pulsing, alternately change DMEM containing VEGF, containing the DMEM of HGF, containing DMEM and the general DMEM of liver cell inductor stage by stage, realize hepatocellular organized stable, the development of vascularization and the contact between the two.

5) according to the step 4 of embodiment 1) carry out induction generation and the drug screening of hepatitis.

Embodiment 3: prepare the bionical vascularization hepatic tissue of a kind of personalization and be applied to hepatitis medicament screening

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC); By importing transcription factor (Oct3/4, Sox2, c-Myc and Klf4) reprogrammed in l cell, be induced multi-potent stem cells (induced pluripotent stem cell, iPSC), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with DMEM nutrient solution or (phosphate buffered saline) PBS solution, obtain mass body volume concentrations and be the fibrinogen solution that 10% gelatin solution, sodium alginate soln that mass body volume concentrations is 2% and mass body volume concentrations are 1%, after mixing with gelatin solution, sodium alginate soln and fibrinogen solution volume ratio 2:2:1, obtain natural polymer stand-by; ADSC and iPSC suspension are centrifugal respectively the most at last, and obtaining cell concn is 2 * 10⁵the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁶the natural polymer solution containing iPSC of individual/mL.

2) step 2 of the preparation of branch vessel support: embodiment 1) the synthetic fractionated polymer branch vessel support of preparation.

3) preparation of polymer shell and polymeric isolation layer: material is that mass body volume concentrations is 1 of 5% PLGA, 4-dioxane solution, utilizes multi-nozzle spraying nozzle assembly, by this solution spraying between hydrogel layer, the alternating structure that obtains hydrogel-PLGA layer, is sealing coat; This solution spraying is peripheral to structure the most at last, obtains PLGA shell.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of iPSC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Molding is immersed to PBS solution, and Isosorbide-5-Nitrae-dioxane organic solvent is removed in extraction; Utilize pulsation system to supply with stage by stage inductor and make iPSC be converted into liver cell, inductor is containing activin A, bFGF and HGF; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; By the said structure vitro culture of pulsing, alternately change DMEM containing VEGF, containing the DMEM of HGF, DMEM and the general DMEM of liver cell inductor stage by stage, realize hepatocellular organized stable, the development of vascularization and the contact between the two.

5) according to the step 4 of embodiment 1) carry out induction generation and the drug screening of hepatitis.

Embodiment 4: prepare the bionical vascularization liver cancer tissue of a kind of personalization and be applied to liver-cancer medicine screening

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC), buy liver cancer cell (HepG2), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with PBS solution, obtain mass body volume concentrations and be the fibrinogen solution that 10% gelatin solution, sodium alginate soln that mass body volume concentrations is 5% and mass body volume concentrations are 1%, after mixing with gelatin solution, sodium alginate soln and fibrinogen solution volume ratio 3:2:1, obtain natural polymer stand-by; ADSC and HepG2 are centrifugal respectively the most at last, and sneak into respectively in natural polymer solution, and obtaining cell concn is 3 * 10⁵the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁶the natural polymer solution containing HepG2 of individual/mL.

2) preparation of branch vessel support: Solidworks software design branch vessel three-dimensional model, after delamination software is analyzed, by 3D printing technique screw extruding molten state ABS, obtains branch vessel support.

3) polymeric isolation layer and polymer shell preparation: material is that mass body volume concentrations is the TEG solution of 5% PU, utilize multi-nozzle spraying nozzle assembly, this solution spraying, between hydrogel layer, is obtained to the alternating structure of hydrogel-PU layer, be protective layer; This solution spraying is peripheral to structure the most at last, obtains PU shell.

4) utilize compound many shower nozzles 3D printing technique along branch vessel path or the computer model of design in advance, to carry out 3 D-printing containing the natural polymer of HepG2, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Molding is immersed to PBS solution, and TEG organic solvent is removed in extraction; Utilizing pulsation system to supply with MEM (minimum essential medium), nutrient solution nutrient solution is stablized liver cancer cell makes its systematism; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change MEM containing VEGF, containing MEM and the general MEM of HGF, realize organized stable, the development of vascularization of liver cancer cell and the contact between the two.

5) liver-cancer medicine screening: the biological indicator of detection record molding (saccharase as amino in jaundice level, albumin level, indocyanine green retention rate, G-6-Pase and tyrosine and vascularization expression); Utilize pulsation system to supply with curative drug as VITAMIN, chemotherapy medicine, Xarelto or medicine newly developed, record and analyze biological indicator, carry out drug screening.

Embodiment 5: prepare the bionical vascularization pancreatic tissue of a kind of personalization and be applied to the drug screening of diabetes

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC) and beta Cell of islet (Pan β), prepare cell suspending liquid; Natural polymer powder is mixed with DMEM nutrient solution, obtain mass body volume concentrations and be 10% gelatin solution stand-by; ADSC and Pan β suspension are centrifugal respectively the most at last, sneak into respectively in natural polymer solution, obtain 3 * 10⁶the natural polymer solution containing ADSC of individual/mL, and 3 * 10⁶the natural polymer solution containing Pan β of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer PU support of preparation.

3) step 3 of the preparation of shell and sealing coat: embodiment 4) prepare PU shell and sealing coat.

4) utilize compound many shower nozzles 3D printing technique along branch vessel path or the computer model of design in advance, to carry out 3 D-printing containing the natural polymer of Pan β, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system supply nutrient solution to stablize beta Cell of islet and make its systematism; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing DMEM and the general DMEM of HGF, realize organized stable, the development of vascularization of beta Cell of islet and the contact between the two.

5) induction generation and the drug screening of diabetes: utilize pulsation system to supply with streptozotocin (Streptozotocin, STZ) directly destroy beta Cell of islet and cause diabetes B, and detection record biological indicator (as glucose consumption, Regular Insulin discharge, C peptide and vascularization expression) now; Utilize pulsation system supply with curative drug as sulfourea, biguanides, glinides or or medicine newly developed, record and analyze biological indicator, carry out drug screening.

Embodiment 6: prepare the bionical vascularization pancreatic tissue of a kind of personalization and be applied to the drug screening of diabetes

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC) and embryonic stem cell (ESC), after subculture, prepare cell suspending liquid; Natural polymer is mixed with DMEM nutrient solution, obtain mass body volume concentrations and be the fibrinogen solution that 10% gelatin solution, sodium alginate soln that mass body volume concentrations is 2% and mass body volume concentrations are 1%, after mixing with gelatin solution, sodium alginate soln and fibrinogen solution volume ratio 2:2:1, obtain natural polymer stand-by; ADSC and ESC are centrifugal respectively the most at last, and sneak into respectively in natural polymer solution, and obtaining cell concn is 3 * 10⁶the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁶the natural polymer solution containing ESC of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer PEG branch vessel support of preparation.

3) step 3 of the preparation of shell and sealing coat: embodiment 4) prepare shell and sealing coat.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of ESC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system to supply with stage by stage inductor and make ESC be converted into beta Cell of islet, inductor is containing Activin A, FGF, CYC, RA, CYC, Extendin-4, IGF and Nicotinamide etc.; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing the DMEM of HGF, containing DMEM and the general DMEM of beta Cell of islet inductor stage by stage, realize organized stable, the development of vascularization of beta Cell of islet and the contact between the two.

5) according to the step 4 of embodiment 5) carry out induction generation and the drug screening of diabetes.

Embodiment 7: prepare the bionical vascularization pancreatic tissue of a kind of personalization and be applied to the drug screening of diabetes

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC); By importing transcription factor (Oct3/4, Sox2, c-Myc and Klf4) reprogrammed in l cell, be induced multi-potent stem cells (induced pluripotent stem cell, iPSC), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with PBS solution, obtain mass body volume concentrations and be 2% sodium alginate soln, ADSC and iPSC suspension are centrifugal respectively the most at last, and obtaining cell concn is 3 * 10⁶the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁶the natural polymer solution containing iPSC of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer PCL branch vessel support of preparation.

3) step 3 of the preparation of shell and sealing coat: embodiment 1) prepare shell and sealing coat.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of iPSC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system to supply with stage by stage inductor and make iPSC be converted into beta Cell of islet, inductor is containing Activin A, FGF, CYC, RA, CYC, Extendin-4, IGF and Nicotinamide etc.; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing the DMEM of HGF, containing DMEM and the general DMEM of beta Cell of islet inductor stage by stage, realize organized stable, the development of vascularization of beta Cell of islet and the contact between the two.

5) according to the step 4 of embodiment 5) carry out induction generation and the drug screening of diabetes.

Embodiment 8: prepare the bionical vascularization Pancreatic Adenocarcinoma of a kind of personalization and be applied to the drug screening of carcinoma of the pancreas

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC), buy pancreatic cancer cell (PANC-1), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with PBS solution, obtain mass body volume concentrations and be 1% fibrinogen solution, ADSC and PANC-1 are centrifugal respectively the most at last, and sneak into respectively in natural polymer solution, and obtaining cell concn is 3 * 10⁶the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁶the natural polymer solution containing PANC-1 of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer PGA branch vessel support of preparation.

3) preparation of shell and sealing coat: embodiment 4 steps 3) prepare shell and sealing coat.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of PANC-1, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel, utilize pulsation system to supply with MEM nutrient solution and stablize pancreatic cancer cell and make its systematism; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change MEM containing VEGF, containing MEM and the general MEM of HGF, realize organized stable, the development of vascularization of pancreatic cancer cell and the contact between the two.

5) carcinoma of the pancreas drug screening: the biological indicator of detection record molding (as gpt, glucose, CEA, CA19-9 and vascularization expression); Utilize pulsation system to supply with curative drug as gemcitabine, 5-Fu, for Ji Ao etc. or medicine newly developed, record and analyze biological indicator, carry out drug screening.

Embodiment 9: prepare the bionical vascularization renal tissue of a kind of personalization and be applied to the drug screening of ephritis

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC) and Kidney of Newborn cell (NKC), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with DMEM nutrient solution, obtain mass body volume concentrations and be 20% gelatin solution, ADSC and NKC suspension is centrifugal, mix with natural polymer solution respectively, obtaining cell concn is 1 * 10⁶the natural polymer solution containing ADSC of individual/mL, and cell concn is 2 * 10⁷the natural polymer solution containing NKC of/mL.Individual

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer PHB branch vessel support of preparation.

3) preparation of shell and sealing coat: embodiment 4 steps 3) prepare shell and sealing coat.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of NKC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system supply nutrient solution to stablize nephrocyte and make its systematism; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Be connected and synthesize polymer branch support, said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing DMEM and the general DMEM of HGF, realize organized stable, the development of vascularization of nephrocyte and the contact between the two.

5) induction generation and the drug screening of ephritis: utilize pulsation system to supply with microbiotic or non-steroidal anti-inflammatory drugs etc. and destroy NKC, bring out acute interstitial nephritis, and detection record biological indicator (as glucose, urea, urine protein creatinine and vascularization expression) now; Utilize pulsation system to supply with curative drug as Zestoretic, Furosemide, hormone or medicine newly developed, record and analyze biological indicator, carry out drug screening.

Embodiment 10: prepare the bionical vascularization renal tissue of a kind of personalization and be applied to the drug screening of ephritis

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC) and embryonic stem cell (ESC), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with DMEM nutrient solution, obtain mass body volume concentrations and be the fibrinogen solution that 10% gelatin solution, sodium alginate soln that mass body volume concentrations is 2% and mass body volume concentrations are 1%, more than state clearly after sol solution, sodium alginate soln and fibrinogen solution volume ratio 2:2:1 mix and obtain natural polymer solution for later use; ADSC and ESC suspension is centrifugal, mix with natural polymer solution respectively, obtaining cell concn is 1 * 10⁵the natural polymer solution containing ADSC of individual/mL, and cell concn is 1 * 10⁵the natural polymer solution containing ESC of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer PU branch vessel support of preparation.

3) preparation of polymer shell and polymeric isolation layer: embodiment 4 steps 3) prepare shell and sealing coat.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of ESC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system to supply with stage by stage inductor and make ESC be converted into nephrocyte, inductor is containing FGF-2 and LIF etc.; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing the DMEM of HGF, containing DMEM and the general DMEM of nephrocyte inductor, realize organized stable, the development of vascularization of nephrocyte and the contact between the two.

5) according to the step 4 of embodiment 9) carry out induction generation and the ephritis drug screening of ephritis.

Embodiment 11: prepare the bionical vascularization renal tissue of a kind of personalization and be applied to the drug screening of ephritis

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC); By importing transcription factor (Oct3/4, Sox2, c-Myc and Klf4) reprogrammed in l cell, be induced multi-potent stem cells (induced pluripotent stem cell, iPSC), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with PBS solution, obtain mass body volume concentrations and be the fibrinogen solution that 10% gelatin solution, sodium alginate soln that mass body volume concentrations is 2% and mass body volume concentrations are 1%, after mixing with gelatin solution, sodium alginate soln and fibrinogen solution volume ratio 1:2:2, obtain natural polymer stand-by; ADSC and iPSC suspension are centrifugal respectively the most at last, and obtaining cell concn is 1 * 10⁷the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁷the natural polymer solution containing iPSC of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer PLGA branch vessel support of preparation.

3) step 3 of the preparation of polymer shell and polymeric isolation layer: embodiment 1) prepare shell and sealing coat.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of iPSC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system to supply with stage by stage inductor and make iPSC be converted into nephrocyte, inductor is containing FGF-2 and LIF etc.; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Be connected and synthesize polymer branch support, said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing the DMEM of HGF, containing DMEM and the general DMEM of nephrocyte inductor, realize organized stable, the development of vascularization of nephrocyte and the contact between the two.

5) according to the step 4 of embodiment 9) carry out induction generation and the ephritis drug screening of ephritis.

Embodiment 12: prepare a kind of personalization bionical vascularization renal carcinoma tissue and be applied to the drug screening of kidney

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC), buy kidney cancer cell (RCC), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with PBS solution, obtain mass body volume concentrations and be the fibrinogen solution that 10% gelatin solution and mass body volume concentrations are 1%, after mixing with gelatin solution and fibrinogen solution volume ratio 1:1, obtain natural polymer stand-by; ADSC and RCC are centrifugal respectively the most at last, and sneak into respectively in natural polymer solution, and obtaining cell concn is 3 * 10⁶the natural polymer solution containing ADSC of individual/mL, and cell concn is 3 * 10⁶the natural polymer solution containing RCC of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) prepare natural polymer sodium alginate branch vessel support, crosslinked with CaCl2.

3) step 3 of the preparation of polymer shell and polymeric isolation layer: embodiment 1) prepare natural polymer sodium alginate shell and sealing coat, crosslinked with CaCl2.

4) the final establishment of external bionic model: utilize compound many shower nozzles 3D printing technique to carry out 3 D-printing along branch vessel path or the computer model of design in advance containing the natural polymer of RCC, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system supply MEM nutrient solution to stablize kidney cancer cell and make its systematism; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change MEM containing VEGF, containing MEM and the general MEM of HGF, realize organized stable, the development of vascularization of kidney cancer cell and the contact between the two.

5) kidney drug screening: the biological indicator of detection record molding (as blood urea nitrogen, creatinine, alkaline phosphatase, serum lactic dehydrogenase and vascularization expression); Utilize pulsation system to supply with curative drug and cut down strain monoclonal antibody or medicine newly developed as Xarelto, shellfish, record and analyze biological indicator, carry out drug screening.

Embodiment 13 prepares a kind of personalization bionical vascularization lymphatic cancer tissues and is applied to the drug screening of lymphatic cancer

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC), buy lymphocytic cancer cell (EL4), after subculture, prepare cell suspending liquid; Natural polymer powder is mixed with PBS solution, obtain mass body volume concentrations and be 1% fibrinogen solution, ADSC and EL4 are centrifugal respectively the most at last, and sneak into respectively in natural polymer solution, and obtaining cell concn is 2 * 10⁵the natural polymer solution containing ADSC of individual/mL, and cell concn is 2 * 10⁵the natural polymer solution containing EL4 of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) prepare synthesis of natural macromolecular fibre proteinogen branch vessel support, with the zymoplasm polymerization of 100U/mL.

3) step 3 of the preparation of polymer shell and polymeric isolation layer: embodiment 1) prepare natural polymer Fibrinogen shell and sealing coat, with the zymoplasm polymerization of 100U/mL.

4) utilize compound many shower nozzles 3D printing technique along branch vessel path or the computer model of design in advance, to carry out 3 D-printing containing the natural polymer of EL4, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Utilize pulsation system supply training MEM nutrient solution to stablize lymphocytic cancer cell and make its systematism; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing DMEM and the general DMEM of HGF, realize organized stable, the development of vascularization of lymphocytic cancer cell and the contact between the two.

5) lymphatic cancer drug screening: the biological indicator of detection record molding (as lactate dehydrogenase, alkaline phosphatase, tumor giant cell and vascularization expression); Utilize pulsation system to supply with curative drug as Zorubicin, bleomycin, vincaleucoblastine, first nitrogen miaow ammonia or medicine newly developed, record and analyze biological indicator, carry out drug screening.

Embodiment 14: prepare the little ear tissue of the bionical vascularization of a kind of personalization

1) preparation of celliferous hydrogel: extract adipose derived stem cells of rats (ADSC) and chondrocyte (CHO), prepare cell suspending liquid; Natural polymer powder is mixed with DMEM nutrient solution, obtain mass body volume concentrations and be 5% gelatin solution, ADSC and CHO suspension are centrifugal respectively the most at last, sneak into respectively in natural polymer solution, obtain 3 * 10⁶the natural polymer solution containing ADSC of individual/mL, and 3 * 10⁶the natural polymer solution containing CHO of individual/mL.

2) preparation of branch vessel support: according to the step 2 of embodiment 1) the synthetic polymer ABS branch vessel support of preparation.

3) preparation of polymer shell and polymeric isolation layer: material is that mass body volume concentrations is Isosorbide-5-Nitrae-dioxane solution of 3% PU, sneaks into heparin and leptin in solution, utilizes multi-nozzle spraying nozzle assembly, by this solution spraying between hydrogel layer; Utilize spray nozzle spraying PBS solution, extraction, removal TEG, obtain porous PU sealing coat; The alternating structure that obtains hydrogel-PU layer, is protective layer; This solution spraying is peripheral to structure the most at last, obtains PU shell.

4) utilize compound many shower nozzles 3D printing technique along branch vessel path or the computer model of design in advance, to carry out 3 D-printing containing the natural polymer of CHO, interval reserves 50% branch vessel space and refuses cell assembling; Utilize afterwards compound many shower nozzles 3D printing technique along reserved path inside above-mentioned branch vessel, to print containing the natural polymer of ADSC, and inside printable layer reserving hole channel; Molding is immersed to PBS solution, and TEG organic solvent is removed in extraction; Utilize pulsation system supply nutrient solution to stablize chondrocyte CHO and make its systematism; Utilize pulsation system to supply with the angiogenesis factor (VEGF) and pHGF (HGF), induction ADSC is divided into capillary endothelial cell or smooth muscle cell; Said structure is proceeded to the vitro culture of pulsing, alternately change DMEM containing VEGF, containing DMEM and the general DMEM of HGF, realize organized stable, the development of vascularization of Chinese hamster ovary celI and the contact between the two.

Claims (10)

1. the bionical composite structure of personalization, is characterized in that: the bionical composite structure of described personalization comprises polymer shell (106), branch vessel support (102), functional organization district (103), vascular tissue district (104) and polymeric isolation layer (401); Described functional organization district (103) is for containing histiocytic hydrogel layer; Described vascular tissue district (104) is the hydrogel layer containing blood vessel seed cell; Arrange at space-alternating described functional organization district (103) and vascular tissue district (104); Described polymeric isolation layer (401) is intervally arranged in functional organization district (103) and vascular tissue district (104), and Bing Jiang functional organization district (103) and vascular tissue district (104) are divided into a plurality of zonules; Described branch vessel support (102) comprises arteries and vein blood vessel two portions, and arteries is containing at least one entrance and several branches, and vein blood vessel is containing at least one outlet and several branches; Described branch vessel support (102) adopts synthesized polymer material, and branch vessel support (102) is through between polymer shell (106), functional organization district (103), vascular tissue district (104) and polymeric isolation layer (401); Described polymer shell (106) adopts natural polymer or synthesized polymer material.

2. the bionical composite structure of a kind of personalization as claimed in claim 1, is characterized in that: the internal diameter of described branch vessel support is 10 μ m～10mm, and wall thickness is 100 μ m～2mm; The bed thickness of described polymeric isolation layer and polymer shell is 0.1 μ m～2mm.

3. the bionical composite structure of a kind of personalization as claimed in claim 1 or 2, it is characterized in that: the hydrogel layer containing blood vessel seed cell containing histiocytic hydrogel layer and vascular tissue district in described functional organization district is natural polymer hydrogel, and this natural polymer is at least one in gelatin, sodium alginate, Fibrinogen, collagen, matrigel, carrageenin, chitosan, agar, hyaluronic acid, matrigel, elastin and laminin; In described natural polymer hydrogel, be compounded with at least one in cell growth factor, heparin, leptin, taxol and Radix Astragali anti-cancer powder.

4. the bionical composite structure of a kind of personalization as claimed in claim 3, is characterized in that: the mass body volume concentrations of described natural polymer hydrogel is 0.1～40%; Described blood vessel seed cell and histiocytic concentration are 1 * 10³～1 * 10⁸individual/mL; The histocyte in described functional organization district is a kind of in adult tissue's cell, adult stem cell and cancer cells; The blood vessel seed cell in described vascular tissue district is at least one in vascular endothelial cell, fat stem cell, mesenchymal stem cells MSCs, navel blood stem cell, bone marrow stem cell, embryonic stem cell and induced multi-potent stem cells.

5. a preparation method for personalized bionical composite structure as claimed in claim 1, is characterized in that the method comprises the following steps:

1) design branch vessel support three-dimensional model, divides two groups and imitates respectively arteries and vein blood vessel;

2) by synthesized polymer material, prepare containing histiocytic hydrogel, being loaded in the different shower nozzles of compound many shower nozzles 3 D-printing equipment containing the hydrogel of blood vessel seed cell and the synthetic macromolecular solution preparing of preparing;

3) prepare branch vessel support: according to step 1) three-dimensional model, utilize compound many shower nozzles 3 D-printing equipment to extrude synthetic macromolecular solution, and freeze-drying removes organic solvent, or melt extrudes synthesized polymer material, obtain branch vessel support;

4) prepare functional organization district: utilize compound many shower nozzles 3 D-printing equipment successively to print to prepare containing histiocytic hydrogel, printing path is along branch vessel support, print at interval, obtains preliminary three-dimensional structure;

5) prepare vascular tissue district: utilize compound many shower nozzles 3 D-printing equipment successively to print, the hydrogel containing blood vessel seed cell preparing is filled in the space of preliminary three-dimensional structure to three-dimensional structure in the middle of obtaining;

6) prepare polymeric isolation layer and polymer shell: in step 4) and step 5) successively in print procedure, after printing one deck or some layers, utilize the synthetic macromolecular solution of compound many shower nozzles 3 D-printing equipment spraying and extract organic solvent, or melt extrude synthesized polymer material, form polymeric isolation layer, obtain the alternating structure body of celliferous hydrogel layer and polymeric isolation layer; In the peripheral spraying of alternating structure body, synthesize macromolecular solution and extract organic solvent, or melt extruding synthesized polymer material, forming polymer shell, obtaining personalized bionical composite structure precursor;

7) to the bionical composite structure precursor of above-mentioned personalization, provide somatomedin, realize the vascularization in vascular tissue district, make functional organization's district's formative tissue, the cell in Bing Shi vascular tissue district and functional organization district produces contact, finally obtains the described bionical composite structure of personalization.

6. the preparation method of the bionical composite structure of a kind of personalization as claimed in claim 5, is characterized in that: described compound many shower nozzles 3 D-printing equipment employing melt extrudes forming technique or low temperature depositing forming technique is prepared branch vessel support, polymeric isolation layer and polymer shell; When employing melt extrudes forming technique and prepares branch vessel support, polymeric isolation layer and polymer shell, described synthesized polymer material is at least one in acrylonitrile-butadiene-styrene (ABS), urethane, tetrafluoroethylene and carbohydrate; When adopting low temperature depositing forming technique to prepare branch vessel support, polymeric isolation layer and polymer shell, the solute of described synthetic macromolecular solution is at least one in urethane, polycarbonate, polyureas ester, PGA, poly-succinic ester, polyether ester, polyoxyethylene glycol, poly(lactic acid), polycaprolactone, Poly(D,L-lactide-co-glycolide, polyester and polyhydroxy acid ester, the solvent of described synthetic macromolecular solution is TEG or Isosorbide-5-Nitrae-dioxane.

7. the preparation method of the bionical composite structure of a kind of personalization as claimed in claim 5, is characterized in that: the mass body volume concentrations of described synthetic macromolecular solution is 1%～30%; In described synthetic macromolecular solution, be compounded with anticoagulin, this anticoagulin is heparin or taxol.

8. personalized bionical composite structure, for a method for drug screening, is characterized in that the method comprises the steps: as claimed in claim 1

1) if the histocyte in functional organization district adopts adult tissue's cell or adult stem cell, the bionical composite structure of personalization is supplied with to the nutrient solution containing pathogenic medicine, make the bionical composite structure of above-mentioned personalization produce pathological symptom; If the histocyte in functional organization district adopts cancer cells, do not do pathogenic processing;

2) the bionical composite structure of personalization is supplied with to the nutrient solution containing different sorts and various dose curative drug, real-time or follow-up collection nutrient solution;

3) analyze the above-mentioned nutrient solution of having collected, the kind of obtaining medical treatment property medicine, the effect of dosage to described personalized composite structure, by detecting, contrast biological indicator, the effect of judgement medicine.

9. the bionical composite structure of a kind of personalization as claimed in claim 8, for the method for drug screening, is characterized in that: described biological indicator is at least one in carbohydrate lipid metabolism index, liver function index, renal function index, cancer cells index and vascularization index.

10. the bionical composite structure of a kind of personalization as claimed in claim 8, for the method for drug screening, is characterized in that: described pathological symptom is diabetes, organ inflammation, organ failure or tumour.

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