CN102266588B - Preparation method of cell-loaded microchannel hydrogel based on sucrose fiber template - Google Patents
Preparation method of cell-loaded microchannel hydrogel based on sucrose fiber template Download PDFInfo
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Abstract
The invention relates to a preparation method of a cell-loaded microchannel hydrogel based on sucrose fiber template, which comprises the following steps: preparing sucrose fibers, coating a polymer film on the surface of the sucrose fibers, and putting the sucrose fibers with the polymer film in the middle of a polymethyl methacrylate die; preparing a mixed solution of hydrogel sol and cell culture fluid, and pouring the sol-cell mixed liquid into the polymethyl methacrylate die; changing the temperature or carrying out ultraviolet irradiation so that the hydrogel sol is crosslinked to form a hydrogel; dissolving the sucrose fibers, sucrose powder or polyvinylpyrrolidone out of the hydrogel in a phosphate buffer solution to form a cell-loaded microchannel hydrogel; and introducing endothelial cell culture fluid into microchannels of the cell-loaded microchannel hydrogel, and culturing so that the endothelial cells grow and spread to form endothelial microchannels. The cell-loaded microchannel hydrogel prepared by the method has stable microchannel structure, the endothelial cells can not easily shed, and the cell-loaded microchannel hydrogel can wrap the cells in the hydrogel crosslinking process; and the endothelial cell culture liquid is implanted after the microchannels are formed, so as to form the endothelial microchannels.
Description
Technical field
The present invention relates to biomedical engineering field, be specifically related to a kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template.
Background technology
Along with the growth of human longevity, clinical in to the demand of donor organ also in continuous growth.At present, the wretched insufficiency of donor organ makes many patients death because not obtaining alternative organ.Organ lacks has become the outstanding problem that biomedical sector needs to be resolved hurrily, and the proposition of organizational project concept and develop into this problem the most potential a kind of solution is provided.
In the tremendous development process of organizational project, hydrogel has obtained increasing attention, also is a kind of biomaterial that is most widely used in the organizational project in the last few years.In organizational project, often adopt hydrogel as the analogies of extracellular matrix, the compound system that is loaded with the hydrogel of cell can be used for pharmacology and pathological model, and also can be built at In vitro culture can be for tissue or the organ of clinical transplantation.Different types of hydrogel comprises natural and the synthetic water gel, makes up some comparatively simple organizational structure such as skin, bladder and cartilages etc. after being used to make up engineered support or directly having wrapped up cell.But, when structure has the vigorous tissue/organ of high cell concentration, metabolic activity in cells such as heart, liver, kidney, can be very fast by cell depleting from diffusion into the surface to in-house nutrient, rely on nutrient to be difficult to guarantee that from diffusion into the surface the cell of hydrogel inside obtains the prescribing adequate nutrition thing and discharges metabolite merely.The accumulation of nutraceutical shortage and metabolite all can influence the homergy activity of cell and the physiological function of engineered tissue, and how the reinforced nutrition supply has become one of problem demanding prompt solution in the organizational project.
Studies show that in recent years by introduce the microchannel network in hydrogel, can effectively be improved the chemical micro-environment of nutraceutical conveying capacity and regulating cell in the hydrogel, the 26S Proteasome Structure and Function of simulation human vas.At present, some biological micro-fabrication technologies, as template, biometric print technology, based on the patterned method of light and module construction from part etc., be used to construct the microchannel in different carrying in the cellular water gel, can form the endothelialization passage by in the microchannel, planting endotheliocyte.But, being subjected to the influence of hydrogel mechanical property and cell growth, the microchannel of carrying in the cellular water gel is structurally also unstable usually, exists the microchannel to stop up easily, split or cave in, and endotheliocyte comes off easily; In addition, stick to the possible encroached water gel inside of the endotheliocyte of microchannel surface and the macromole foreign body in the microchannel, influence normal growth and the functional expression of cell.
Summary of the invention
In order to overcome the shortcoming of above-mentioned prior art, the object of the present invention is to provide a kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template, stable by MCA in year cell microchannel hydrogel of this method preparation, endotheliocyte is not easy to come off, can wrap up cell in the hydrogel cross-linking process, the microchannel forms the back and injects the microchannel that culture fluid of endothelial cell can form endothelialization.
In order to achieve the above object, the technical scheme taked of the present invention is:
Scheme 1
A kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 2～10wt% in the polymer solution of dipping, polymer is that mass ratio is poly-left lactic acid (PLLA) and polyvinylpyrrolidone (PVP) or polylactic acid-glycolic guanidine-acetic acid copolymer (PLGA) and the polyvinylpyrrolidone (PVP) of 1～4:1, solvent is dichloromethane, chloroform or 1, the 4-dioxane, after the solvent for the treatment of the fiber surface polymeric layer under the room temperature volatilizees fully, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, light trigger, hydrogel monomer and hydrogel solvent are configured to hydrogel colloidal sol by mass ratio 0～1:3～30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone or gorgeous good solid 2959, the hydrogel monomer is agarose, collagen, hyaluronic acid, chitosan, Na-alginate, polyvinyl alcohol or Polyethylene Glycol and derivant thereof, and the hydrogel solvent is that deionized water, pure water or pH value are 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
5～10
7The cell culture fluid of cells/mL is 1～4:1 mix homogeneously by volume, and the cell in the cell culture fluid is chondrocyte, osteoblast, marrow stromal cell, hepatocyte, fibroblast, myocardial cell, mescenchymal stem cell, fat stem cell or neural precursor;
The 5th step, with the sucrose fibers encapsulation of coated polymer layer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, made it to be cross-linked to form the hydrogel that carries cell by changing temperature or ultra-vioket radiation;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and polyvinylpyrrolidone (PVP), formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 10
5～10
7Culture fluid of endothelial cell 0.1～1mL of cells/mL, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
The present invention can also realize by following method:
Scheme 2
A kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 5～15wt% in the polymer solution of dipping, polymer is that mass ratio is polyurethane and the sucrose powder of 1～4:1, solvent is dichloromethane, chloroform or 1,4-dioxane, and the particle diameter of sucrose powder is 1～10 μ m, treat under the room temperature that solvent volatilizees fully and urethane cures crosslinked after, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, light trigger, hydrogel monomer and hydrogel solvent are configured to hydrogel colloidal sol by mass ratio 0～1:3～30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone or gorgeous good solid 2959, the hydrogel monomer is agarose, collagen, hyaluronic acid, chitosan, Na-alginate, polyvinyl alcohol or Polyethylene Glycol and derivant thereof, and the hydrogel solvent is that deionized water, pure water or pH value are 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
5～10
7The cell culture fluid of cells/mL is 1～4:1 mix homogeneously by volume, and the cell in the cell culture fluid is chondrocyte, osteoblast, marrow stromal cell, hepatocyte, fibroblast, myocardial cell, mescenchymal stem cell, fat stem cell or neural precursor;
The 5th step, with the sucrose fibers encapsulation of coated polymer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, made it to be cross-linked to form the hydrogel that carries cell by changing temperature or ultra-vioket radiation;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and sucrose powder, formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 10
5～10
7Culture fluid of endothelial cell 0.1～1mL of cells/mL, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
This method uses the sucrose fiber of polymer overmold as template, and is simple, efficient height, the good stability of microchannel; Can wrap up cell in the hydrogel cross-linking process, and the introducing of polymer inner layer both can have been kept the stability of microchannel in the hydrogel, also can prevent endotheliocyte and outside macromole foreign body encroached water gel inside simultaneously; In addition, different sucrose fibers can mixing together under the situation that changes humidity or temperature, and the three-dimensional that makes this method also have quick complex structure is carried the potentiality of cell microchannel hydrogel.
The specific embodiment
The present invention is described in detail below in conjunction with concrete embodiment:
Embodiment 1
A kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 4wt% in the polymer solution of dipping, polymer is that mass ratio is poly-left lactic acid (PLLA) and the polyvinylpyrrolidone (PVP) of 2:1, solvent is chloroform, after the solvent for the treatment of the fiber surface polymeric layer under the room temperature volatilizees fully, place in the middle of polymethyl methacrylate (PMMA) mould;
In the 3rd step, preparation hydrogel colloidal sol is configured to hydrogel colloidal sol with hydrogel monomer and hydrogel solvent by mass ratio 3:100 mix homogeneously, and the hydrogel monomer is agarose, and the hydrogel solvent is that pH value is 5.7～8.0 PBS buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
62:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is fibroblast;
The 5th step, with the sucrose fibers encapsulation of coated polymer layer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, in 4 ℃ of refrigerators, place 15min, make agarose colloidal sol be cross-linked to form the hydrogel that carries cell by changing temperature;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and polyvinylpyrrolidone (PVP), formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
Embodiment 2
A kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 4wt% in the polymer solution of dipping, polymer is that mass ratio is polylactic acid-glycolic guanidine-acetic acid copolymer (PLGA) and the polyvinylpyrrolidone (PVP) of 1:1, solvent is chloroform, after the solvent for the treatment of the fiber surface polymeric layer under the room temperature volatilizees fully, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, light trigger, hydrogel monomer and hydrogel solvent are configured to hydrogel colloidal sol by mass ratio 0.5:30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone, the hydrogel monomer is polyethylene glycol dimethacrylate (PEG-DMA), and the hydrogel solvent is that pH value is 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
61:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is smooth muscle cell;
The 5th step, the sucrose fibers encapsulation of coated polymer layer in hydrogel, is poured into the mixed liquor of the 4th step preparation in second step ready polymethyl methacrylate (PMMA) mould, irradiation under ultraviolet ray 3min makes it to be cross-linked to form the hydrogel that carries cell;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and polyvinylpyrrolidone (PVP), formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
Embodiment 3
A kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 160～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 8wt% in the polymer solution of dipping, polymer is that mass ratio is polyurethane and the sucrose powder of 2:1, solvent is 1,4-dioxane, and the particle diameter of sucrose powder is 5 μ m, treat under the room temperature that solvent volatilizees fully and urethane cures crosslinked after, place in the middle of polymethyl methacrylate (PMMA) mould;
In the 3rd step, preparation hydrogel colloidal sol is configured to hydrogel colloidal sol with monomer and the hydrogel solvent of hydrogel by mass ratio 3:100 mix homogeneously, and the hydrogel monomer is agarose, and the hydrogel solvent is that pH value is 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
62:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is chondrocyte;
The 5th step, with the sucrose fibers encapsulation of coated polymer layer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, in 4 ℃ of refrigerators, place 15min, make agarose colloidal sol be cross-linked to form the hydrogel that carries cell by changing temperature;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and sucrose powder, formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
Embodiment 4
A kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 160～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 10wt% in the polymer solution of dipping, polymer is that mass ratio is polyurethane and the sucrose powder of 1:1, solvent is 1,4-dioxane, and the particle diameter of sucrose powder is 5 μ m, treat under the room temperature that solvent volatilizees fully and urethane cures crosslinked after, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, monomer and the hydrogel solvent of light trigger, hydrogel are configured to hydrogel colloidal sol by mass ratio 1:30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone, the hydrogel monomer is polyethylene glycol dimethacrylate (PEG-DMA), and the hydrogel solvent is that pH value is 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
61:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is hepatocyte;
The 5th step, the sucrose fibers encapsulation of coated polymer layer in hydrogel, is poured into the mixed liquor of the 4th step preparation in second step ready polymethyl methacrylate (PMMA) mould, irradiation under ultraviolet ray 3min makes it to be cross-linked to form the hydrogel that carries cell;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and sucrose powder, formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
Claims (6)
1. the preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template is characterized in that, may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 2～10wt% in the polymer solution of dipping, polymer is that mass ratio is poly-left lactic acid (PLLA) and polyvinylpyrrolidone (PVP) or polylactic acid-glycolic guanidine-acetic acid copolymer (PLGA) and the polyvinylpyrrolidone (PVP) of 1～4:1, solvent is dichloromethane, chloroform or 1, the 4-dioxane, after the solvent for the treatment of the fiber surface polymeric layer under the room temperature volatilizees fully, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, light trigger, hydrogel monomer and hydrogel solvent are configured to hydrogel colloidal sol by mass ratio 0～1:3～30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone or gorgeous good solid 2959, the hydrogel monomer is agarose, collagen, hyaluronic acid, chitosan, Na-alginate, polyvinyl alcohol or Polyethylene Glycol and derivant thereof, and the hydrogel solvent is that deionized water, pure water or pH value are 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
5～10
7The cell culture fluid of cells/mL is 1～4:1 mix homogeneously by volume, and the cell in the cell culture fluid is chondrocyte, osteoblast, marrow stromal cell, hepatocyte, fibroblast, myocardial cell, mescenchymal stem cell, fat stem cell or neural precursor;
The 5th step, with the sucrose fibers encapsulation of coated polymer layer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, made it to be cross-linked to form the hydrogel that carries cell by changing temperature or ultra-vioket radiation;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and polyvinylpyrrolidone (PVP), formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 10
5～10
7Culture fluid of endothelial cell 0.1～1mL of cells/mL, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
2. a kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template according to claim 1 may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 4wt% in the polymer solution of dipping, polymer is that mass ratio is poly-left lactic acid (PLLA) and the polyvinylpyrrolidone (PVP) of 2:1, solvent is chloroform, after the solvent for the treatment of the fiber surface polymeric layer under the room temperature volatilizees fully, place in the middle of polymethyl methacrylate (PMMA) mould;
In the 3rd step, preparation hydrogel colloidal sol is configured to hydrogel colloidal sol with hydrogel monomer and hydrogel solvent by mass ratio 3:100 mix homogeneously, and the hydrogel monomer is agarose, and the hydrogel solvent is that pH value is 5.7～8.0 PBS buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
62:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is fibroblast;
The 5th step, with the sucrose fibers encapsulation of coated polymer layer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, in 4 ℃ of refrigerators, place 15min, make agarose colloidal sol be cross-linked to form the hydrogel that carries cell by changing temperature;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and polyvinylpyrrolidone (PVP), formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
3. a kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template according to claim 1 may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 4wt% in the polymer solution of dipping, polymer is that mass ratio is polylactic acid-glycolic guanidine-acetic acid copolymer (PLGA) and the polyvinylpyrrolidone (PVP) of 1:1, solvent is chloroform, after the solvent for the treatment of the fiber surface polymeric layer under the room temperature volatilizees fully, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, light trigger, hydrogel monomer and hydrogel solvent are configured to hydrogel colloidal sol by mass ratio 0.5:30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone, the hydrogel monomer is polyethylene glycol dimethacrylate (PEG-DMA), and the hydrogel solvent is that pH value is 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
61:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is smooth muscle cell;
The 5th step, the sucrose fibers encapsulation of coated polymer layer in hydrogel, is poured into the mixed liquor of the 4th step preparation in second step ready polymethyl methacrylate (PMMA) mould, irradiation under ultraviolet ray 3min makes it to be cross-linked to form the hydrogel that carries cell;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and polyvinylpyrrolidone (PVP), formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
4. preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 150～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 5～15wt% in the polymer solution of dipping, polymer is that mass ratio is polyurethane and the sucrose powder of 1～4:1, solvent is dichloromethane, chloroform or 1,4-dioxane, and the particle diameter of sucrose powder is 1～10 μ m, treat under the room temperature that solvent volatilizees fully and urethane cures crosslinked after, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, light trigger, hydrogel monomer and hydrogel solvent are configured to hydrogel colloidal sol by mass ratio 0～1:3～30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone or gorgeous good solid 2959, the hydrogel monomer is agarose, collagen, hyaluronic acid, chitosan, Na-alginate, polyvinyl alcohol or Polyethylene Glycol and derivant thereof, and the hydrogel solvent is that deionized water, pure water or pH value are 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
5～10
7The cell culture fluid of cells/mL is 1～4:1 mix homogeneously by volume, and the cell in the cell culture fluid is chondrocyte, osteoblast, marrow stromal cell, hepatocyte, fibroblast, myocardial cell, mescenchymal stem cell, fat stem cell or neural precursor;
The 5th step, with the sucrose fibers encapsulation of coated polymer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, made it to be cross-linked to form the hydrogel that carries cell by changing temperature or ultra-vioket radiation;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and sucrose powder, formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 10
5～10
7Culture fluid of endothelial cell 0.1～1mL of cells/mL, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
5. a kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template according to claim 4 may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 160～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of control traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 8wt% in the polymer solution of dipping, polymer is that mass ratio is polyurethane and the sucrose powder of 2:1, solvent is 1,4-dioxane, and the particle diameter of sucrose powder is 5 μ m, treat under the room temperature that solvent volatilizees fully and urethane cures crosslinked after, place in the middle of polymethyl methacrylate (PMMA) mould;
In the 3rd step, preparation hydrogel colloidal sol is configured to hydrogel colloidal sol with monomer and the hydrogel solvent of hydrogel by mass ratio 3:100 mix homogeneously, and the hydrogel monomer is agarose, and the hydrogel solvent is that pH value is 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
62:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is chondrocyte;
The 5th step, with the sucrose fibers encapsulation of coated polymer layer in hydrogel, the mixed liquor of the 4th step preparation is poured in second step ready polymethyl methacrylate (PMMA) mould, in 4 ℃ of refrigerators, place 15min, make agarose colloidal sol be cross-linked to form the hydrogel that carries cell by changing temperature;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and sucrose powder, formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
6. a kind of preparation method of carrying cell microchannel hydrogel based on the sucrose fibrous template according to claim 4 may further comprise the steps:
The first step, preparation sucrose fiber, as the template that makes up the microchannel, sucrose crystal is heated to makes it more than the coking temperature to melt fully, after being cooled to 160～180 ° of C, up draw the sucrose melt with the stainless pipe of external diameter 2mm, the speed of traction is 0.1～1.0m/s, make the diameter of sucrose fiber at 10 μ m～1mm, the sucrose fiber for preparing is stored in the drying chamber;
Second step, apply the polymer solution that a layer thickness is 20～100 μ m at the sucrose fiber surface, polymer concentration is 10wt% in the polymer solution of dipping, polymer is that mass ratio is polyurethane and the sucrose powder of 1:1, solvent is 1,4-dioxane, and the particle diameter of sucrose powder is 5 μ m, treat under the room temperature that solvent volatilizees fully and urethane cures crosslinked after, place in the middle of polymethyl methacrylate (PMMA) mould;
The 3rd step, preparation hydrogel colloidal sol, monomer and the hydrogel solvent of light trigger, hydrogel are configured to hydrogel colloidal sol by mass ratio 1:30:100 mix homogeneously, light trigger is 2-hydroxy-2-methyl propiophenone, the hydrogel monomer is polyethylene glycol dimethacrylate (PEG-DMA), and the hydrogel solvent is that pH value is 5.7～8.0 phosphate (PBS) buffer solution;
In the 4th step, the mixed liquor of preparation hydrogel colloidal sol and cell culture fluid is 3 * 10 with hydrogel colloidal sol and the cell concentration for preparing in the 3rd step
61:1 is at 40 ℃ of mix homogeneously by volume for the cell culture fluid of cells/mL, and the cell in the cell culture fluid is hepatocyte;
The 5th step, the sucrose fibers encapsulation of coated polymer layer in hydrogel, is poured into the mixed liquor of the 4th step preparation in second step ready polymethyl methacrylate (PMMA) mould, irradiation under ultraviolet ray 3min makes it to be cross-linked to form the hydrogel that carries cell;
The 6th step dissolved in pH value is 5.7～8.0 phosphate (PBS) buffer solution carrying sucrose fiber in the hydrogel of cell and sucrose powder, formed the MCA that has polymer inner layer in the hydrogel that carries cell;
In the 7th step, injecting cell concentration in the microchannel of carrying the cellular water gel is 2 * 10
6The culture fluid of endothelial cell 0.2mL of cells/ml, cultivation is sprawled endothelial cell growth, finally forms the microchannel of endothelialization.
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