CN102488926A - Tissue engineering scaffold for urethra reconstruction and preparation method thereof - Google Patents

Tissue engineering scaffold for urethra reconstruction and preparation method thereof Download PDF

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Publication number
CN102488926A
CN102488926A CN2011104253068A CN201110425306A CN102488926A CN 102488926 A CN102488926 A CN 102488926A CN 2011104253068 A CN2011104253068 A CN 2011104253068A CN 201110425306 A CN201110425306 A CN 201110425306A CN 102488926 A CN102488926 A CN 102488926A
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tissue engineering
silk fibroin
regenerated silk
urethra
engineering bracket
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CN102488926B (en
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张耀鹏
范苏娜
邵惠丽
杭怡春
潘辉
徐月敏
冯超
谢敏凯
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Shanghai Shuixing Home Textile Co Ltd
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Donghua University
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Abstract

The invention relates to a tissue engineering scaffold for urethra reconstruction and a preparation method thereof. The preparation method comprises the steps of: (1) preparing a regenerated silk fibroin aqueous solution; (2) preparing a vascular endothelial growth factor and bovine serum albumin aqueous solution; (3) mixing the solutions prepared in the steps (1) and (2) to prepare a regenerated silk fibroin solution containing the vascular endothelial growth factor; and (4) preparing the tissue engineering scaffold by adopting an electrostatic spinning technology with the solution prepared in the step (3) as a spinning liquid and a decellularized matrix as a base layer. According to the invention, the tissue engineering scaffold is of a double-layer structure composed of a decellularized matrix base layer and a functional layer, and the functional layer is a fibrofelt which is formed by regenerated silk fibroin fiber mixed with the vascular endothelial growth factor and bovine serum albumin and has a three-dimensional staggered network structure characteristic. The tissue engineering scaffold prepared by the method not only has excellent mechanical properties, but also can be early and quickly vascularized in vivo, and can be used for repairing long-segment urethrostenosis.

Description

A kind of tissue engineering bracket that is used for the urethra reconstruction and preparation method thereof
Technical field
The invention belongs to the technical field of textile industry, material science; Be particularly related to a kind of tissue engineering bracket that is used for the urethra reconstruction and preparation method thereof; With the regenerated silk fibroin solution that contains VEGF is spinning liquid; Through electrostatic spinning process it is sprayed on the acellular matrix, has prepared and be used for the tissue engineering bracket that urethra is rebuild.
Background technology
A long section complexity urethral stricture not only can cause urinary extravasation, urine fistula, via etc. local complication, severe patient even involve other internal organs of urinary system and cause the general pathological changes has a strong impact on patient's quality of life.Be that the alternative medicine of reconstruction biomaterials has shown good prospects for application in the treatment of complexity urethral stricture previously with multiple autologous tissue; But there is the shortcoming with " the sacrifice normal structure is a cost; damaged with the operation wound repair tissue " in this therapy, inevitably causes the complication at the position of drawing materials.The rise of organizational project and fast development are that new treatment approach has been opened up in the reparation of urethra pathological changes.Owing to acellular matrix (BAMG) under acellular matrix (SIS) under the small intestinal mucosa and the bladder mucosa has non-immunogenicity, antimicrobial acivity etc.; Can obtain metabolism through biodegradation in vivo; Thereby obtain broad research and be applied clinically, but postoperative patient is prone to take place complication such as urethra restenosis, urethral diverticulum and urine fistula.Tracing it to its cause, is because of the vascularization fast in vivo of acellular matrix under acellular matrix under the simple small intestinal mucosa or the bladder mucosa.Therefore promote under the small intestinal mucosa that the early stage in vivo vascularization fast of acellular matrix is to improve the key element of its clinical application effect under acellular matrix or the bladder mucosa.
VEGF (VEGF) is a kind of effective angiogenic growth factor; Can directly act on vascular endothelial cell and promote vascular endothelial cell proliferation; Suppress endothelial cell apoptosis, increase vascular permeability, induce new vessels to form; But its short half-life, only exogenous VEGF added under the small intestinal mucosa under acellular matrix or the bladder mucosa and can not effectively and enduringly urge its vascularization in the acellular matrix.
Solution to the problems described above: the one, adopt technique for gene engineering, make seed cell carry the VEGF gene, but this method poor controllability; The 2nd, adopt timbering material that VEGF is wrapped up, make its slow release VEGF.The key of second method is to select suitable clad material.At present the degradable clad material of bibliographical information mainly contains two kinds: the one, to process by animal collagen or collagen fiber, and another kind is to be processed by polylactic acid and medical plasticizer or collagen fiber.But after these degradation materials implant, can cause the implantation region reaction that is inflamed, exist the risk of the interior rejection of body simultaneously thereby in degradation process, be prone to produce acid.
And the objective of the invention is to overcome above-mentioned shortcoming, select fibroin albumen as clad material.Fibroin albumen not only has excellent mechanical property, and has excellent biological compatibility, degradability, high ventilation poisture-penetrability, good vascularization promoting ability etc.Publication number is that to disclose a kind of be raw material with the natural silk to the Chinese invention patent " a kind of fibre for tissue engineering scaffold and preparation method thereof " of CN 13720203A; Through coming unstuck, after the operation such as dissolving, adopting wet spinning technology to prepare the method for tissue engineering bracket material.It is raw material with silkworm silk and collagen protein that the Chinese invention patent " a kind of cell culturing rack material and preparation method thereof " of publication number CN 1887362A has been announced a kind of, adopts the static spinning technique to prepare the method for three-dimensional netted fibroin albumen composite nano fiber non-woven mat.But, also do not see the correlational study of the compound preparation tissue engineering bracket of acellular matrix and regenerated silk fibroin under acellular matrix under the small intestinal mucosa or the bladder mucosa and the bibliographical information or the patent of production both at home and abroad to so far.
Summary of the invention
Technical problem to be solved by this invention provides a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction, is that the regenerated silk fibroin solution that contains VEGF is sprayed on the method on the acellular matrix through electrostatic spinning technique.Through this method being used for of making double-decker engineering rack of being made up of basic unit and functional layer of tissue engineering bracket that urethra rebuilds, wherein basic unit is an acellular matrix, and functional layer is the regenerated silk fibroin fiber felt; This fiber felt is the fiber felt with three-dimensional staggered-mesh architectural feature that is mixed with the regenerated silk fibroin fiber formation of VEGF and bovine serum albumin; VEGF is uniformly dispersed in the regenerated silk fibroin fiber, has guaranteed that it can discharge and produce short angiogenesis function gradually, and acellular matrix then gives support excellent mechanical property; Simultaneously; This preparation process is simple, easy operating, environmentally safe.
The object of the present invention is to provide that a kind of good biocompatibility, biological degradability are excellent, mechanical property is excellent, simultaneously can be in vivo tissue engineering bracket of vascularization and preparation method thereof fast.
The double-decker engineering rack that a kind of tissue engineering bracket that is used for the urethra reconstruction of the present invention is made up of basic unit and functional layer; The quality of wherein said VEGF accounts for 3.5 * 10 of fiber felt gross mass -5~9.5 * 10 -5%, the quality of described bovine serum albumin accounts for 1.5 * 10 of fiber felt gross mass -3~4 * 10 -3%.
As optimized technical scheme:
Aforesaid a kind of tissue engineering bracket that urethra is rebuild that is used for, described acellular matrix are a kind of in the acellular matrix under acellular matrix or the bladder mucosa under the small intestinal mucosa.
Aforesaid a kind of tissue engineering bracket that urethra is rebuild that is used for, described acellular matrix is translucent cellular thin film.
Aforesaid a kind of tissue engineering bracket that urethra is rebuild that is used for, the diameter range of described regenerated silk fibroin fiber is 200nm~5 μ m.
Aforesaid a kind of tissue engineering bracket that urethra is rebuild that is used for, the porosity of described fiber felt is 50~90%.
The present invention also provides a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction, the steps include:
(1) with Bombyx bombycis be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 30~40% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.3~0.7% aqueous sodium carbonate comes unstuck that Bombyx bombycis is used mass percent; Be dissolved in the lithium bromide water solution that molar concentration is 9.0~9.5mol/L, obtain regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 30~40% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 15~25 μ g/ml, the mass concentration of bovine serum albumin is 0.6~1mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 60: 1~130: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning technique that it is sprayed on the acellular matrix, and preparation is used for the tissue engineering bracket that urethra is rebuild;
Aforesaid a kind of method for preparing that is used for the tissue engineering bracket that urethra rebuilds, described Bombyx bombycis are a kind of in mulberry cocoon, tussah cocoon or the Semen Ricini Bombyx bombycis.
Aforesaid a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction, the spinning ambient temperature when described electrostatic spinning technique prepares the tissue engineering bracket that is used for the urethra reconstruction is 2~6 ℃.
Aforesaid a kind of method for preparing that is used for the tissue engineering bracket that urethra rebuilds, described acellular matrix are acellular matrix under acellular matrix or the bladder mucosa under the small intestinal mucosa.
Further, the described tissue engineering bracket that is used for the urethra reconstruction is carried out post processing; Described post processing is meant soaks 30~90min with the tissue engineering bracket that is used for the urethra reconstruction at the alcohol-water mixed solution.
Aforesaid a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction, ethanol accounts for 75~95% of alcohol-water mixed solution cumulative volume in the described alcohol-water mixed solution.
Beneficial effect:
Proposed to the invention property to be used for the method for preparing of the tissue engineering bracket that urethra rebuilds.Under the prerequisite that guarantees the regenerated silk fibroin higher concentration, avoided the use of organic solvent (like formic acid, hexafluoroisopropanol etc.), not only reduced production cost, and avoided injury human body.The tissue engineering bracket that is used for the urethra reconstruction that makes according to method of the present invention not only has excellent mechanical property, and vascularization fast in vivo, satisfies the instructions for use that long section urethral stricture is repaired.
The specific embodiment
Below in conjunction with the specific embodiment, further set forth the present invention.Should be understood that these embodiment only to be used to the present invention is described and be not used in the restriction scope of the present invention.Should be understood that in addition those skilled in the art can do various changes or modification to the present invention after the content of having read the present invention's instruction, these equivalent form of values fall within the application's appended claims institute restricted portion equally.
The double-decker engineering rack that a kind of tissue engineering bracket that is used for the urethra reconstruction of the present invention is made up of basic unit and functional layer; Wherein basic unit is an acellular matrix; Functional layer is the regenerated silk fibroin fiber felt; Wherein the regenerated silk fibroin fiber felt is the fiber felt with three-dimensional staggered-mesh architectural feature that is mixed with the regenerated silk fibroin fiber formation of VEGF and bovine serum albumin, and VEGF and bovine serum albumin be homodisperse in the regenerated silk fibroin fiber; The quality of described VEGF accounts for 3.5 * 10 of said regenerated silk fibroin total fiber mass -5~9.5 * 10 -5%, the quality of described bovine serum albumin accounts for 1.5 * 10 of said regenerated silk fibroin total fiber mass -3~4 * 10 -3%.
Described acellular matrix is a kind of in the acellular matrix under acellular matrix or the bladder mucosa under the small intestinal mucosa.
Described acellular matrix is translucent cellular thin film.
The diameter range of the regenerated silk fibroin fiber of described formation fiber felt is 200nm~5 μ m.
The porosity of described fiber felt is 50~90%.
Tissue engineering bracket of the present invention is used for the reparation of long section urethral stricture.
Embodiment 1
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) with the mulberry cocoon be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 30% regenerated silk fibroin water solution;
The preparation process of said regeneration pure silk fibroin aqueous solution is following: it is after 0.3% aqueous sodium carbonate comes unstuck that mulberry cocoon is used mass percent; Be dissolved in the lithium bromide water solution that molar concentration is 9.0mol/L, obtain regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 30% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 15 μ g/ml, the mass concentration of bovine serum albumin is 0.6mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 130: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning technique it to be sprayed under the small intestinal mucosa on the acellular matrix down at 2 ℃, and preparation is used for the tissue engineering bracket that urethra is rebuild; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 200nm~1 μ m, and the porosity of described fiber felt is 90%.
Embodiment 2
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) with the tussah cocoon be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 33% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.4% aqueous sodium carbonate comes unstuck, to be dissolved in the lithium bromide water solution that molar concentration is 9.1mol/L that tussah cocoon is used mass percent, obtains regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 33% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 17 μ g/ml, the mass concentration of bovine serum albumin is 0.7mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 70: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning technique it to be sprayed under the bladder mucosa on the acellular matrix down at 4 ℃, and preparation is used for the tissue engineering bracket that urethra is rebuild; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 500nm~2 μ m, and the porosity of described fiber felt is 81%.
Embodiment 3
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) be raw material with Semen Ricini Bombyx bombycis, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 35% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.5% aqueous sodium carbonate comes unstuck that Semen Ricini Bombyx bombycis is used mass percent; Be dissolved in the lithium bromide water solution that molar concentration is 9.2mol/L, obtain regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 35% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 20 μ g/ml, the mass concentration of bovine serum albumin is 0.8mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 90: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning technique it to be sprayed under the bladder mucosa on the acellular matrix down at 6 ℃, and preparation is used for the tissue engineering bracket that urethra is rebuild; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 800nm~3 μ m, and the porosity of described fiber felt is 72%.
Embodiment 4
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) be raw material with Semen Ricini Bombyx bombycis, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 37% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.6% aqueous sodium carbonate comes unstuck that Semen Ricini Bombyx bombycis is used mass percent; Be dissolved in the lithium bromide water solution that molar concentration is 9.3mol/L, obtain regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 37% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 22 μ g/ml, the mass concentration of bovine serum albumin is 0.9mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 110: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning technique it to be sprayed under the small intestinal mucosa on the acellular matrix down at 4 ℃, and preparation is used for the tissue engineering bracket that urethra is rebuild; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 1 μ m~4 μ m, and the porosity of described fiber felt is 59%.
Embodiment 5
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) with the mulberry cocoon be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 40% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.7% aqueous sodium carbonate comes unstuck, to be dissolved in the lithium bromide water solution that molar concentration is 9.5mol/L that mulberry cocoon is used mass percent, obtains regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 40% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 25 μ g/ml, the mass concentration of bovine serum albumin is 1.0mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 60: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning technique it to be sprayed under the small intestinal mucosa on the acellular matrix down at 6 ℃, and preparation is used for the tissue engineering bracket that urethra is rebuild; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 1.5 μ m~5 μ m, and the porosity of described fiber felt is 50%.
Embodiment 6
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) with the mulberry cocoon be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 35% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.7% aqueous sodium carbonate comes unstuck, to be dissolved in the lithium bromide water solution that molar concentration is 9.2mol/L that mulberry cocoon is used mass percent, obtains regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 35% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 22 μ g/ml, the mass concentration of bovine serum albumin is 0.9mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 90: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning techniques it to be sprayed under the bladder mucosa on the acellular matrix preparation tissue engineering bracket down at 2 ℃; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 800nm~3 μ m, and the porosity of described fiber felt is 72%;
(5) tissue engineering bracket of step (4) preparation is soaked 90min in the ethanol volume fraction is 75% alcohol-water mixed solution, promptly finally be used for the tissue engineering bracket that urethra is rebuild.
Embodiment 7
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) with the tussah cocoon be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 37% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.7% aqueous sodium carbonate comes unstuck, to be dissolved in the lithium bromide water solution that molar concentration is 9.0mol/L that tussah cocoon is used mass percent, obtains regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 37% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 17 μ g/ml, the mass concentration of bovine serum albumin is 0.7mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 110: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning techniques it to be sprayed under the small intestinal mucosa on the acellular matrix preparation tissue engineering bracket down at 4 ℃; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 1 μ m~4 μ m, and the porosity of described fiber felt is 61%;
(5) tissue engineering bracket of step (4) preparation is soaked 60min in the ethanol volume fraction is 85% alcohol-water mixed solution, promptly finally be used for the tissue engineering bracket that urethra is rebuild.
Embodiment 8
A kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction the steps include:
(1) with the tussah cocoon be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 33% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.5% aqueous sodium carbonate comes unstuck, to be dissolved in the lithium bromide water solution that molar concentration is 9.1mol/L that tussah cocoon is used mass percent, obtains regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 33% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 19 μ g/ml, the mass concentration of bovine serum albumin is 0.8mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 70: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning techniques it to be sprayed under the small intestinal mucosa on the acellular matrix preparation tissue engineering bracket down at 6 ℃; The diameter range of the regenerated silk fibroin fiber of described formation tissue engineering bracket functional layer is 500nm~2 μ m, and the porosity of described fiber felt is 81%;
(5) tissue engineering bracket of step (4) preparation is soaked 30min in the ethanol volume fraction is 95% alcohol-water mixed solution, promptly finally be used for the tissue engineering bracket that urethra is rebuild.

Claims (10)

1. one kind is used for the tissue engineering bracket that urethra is rebuild; It is characterized in that: the double-decker engineering rack that the described tissue engineering bracket that is used for the urethra reconstruction is made up of basic unit and functional layer; Wherein basic unit is an acellular matrix, and functional layer is the regenerated silk fibroin fiber felt; Described regenerated silk fibroin fiber felt is the fiber felt with three-dimensional staggered-mesh architectural feature that is mixed with the regenerated silk fibroin fiber formation of VEGF and bovine serum albumin; The quality of described VEGF accounts for 3.5 * 10 of fiber felt gross mass -5~9.5 * 10 -5%, the quality of described bovine serum albumin accounts for 1.5 * 10 of fiber felt gross mass -3~4 * 10 -3%.
2. a kind of tissue engineering bracket that urethra is rebuild that is used for according to claim 1 is characterized in that, described acellular matrix is a kind of in the acellular matrix under acellular matrix or the bladder mucosa under the small intestinal mucosa.
3. a kind of tissue engineering bracket that urethra is rebuild that is used for according to claim 1 is characterized in that described acellular matrix is translucent cellular thin film.
4. a kind of tissue engineering bracket that urethra is rebuild that is used for according to claim 1 is characterized in that the diameter range of the regenerated silk fibroin fiber of described formation fiber felt is 200nm~5 μ m.
5. a kind of tissue engineering bracket that urethra is rebuild that is used for according to claim 1 is characterized in that the porosity of described fiber felt is 50~90%.
6. a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction according to claim 1 is characterized in that may further comprise the steps:
(1) with Bombyx bombycis be raw material, after coming unstuck, dissolve, dialyse, concentrating, preparation quality percentage ratio is 30~40% regenerated silk fibroin water solution;
The preparation process of said regenerated silk fibroin water solution is following: it is after 0.3~0.7% aqueous sodium carbonate comes unstuck that Bombyx bombycis is used mass percent; Be dissolved in the lithium bromide water solution that molar concentration is 9.0~9.5mol/L, obtain regenerated silk fibroin-lithium bromide water solution; With this solution centrifugal, filtration, dialysis with concentrate, obtain mass percent and be 30~40% regenerated silk fibroin water solution;
(2) with water be solvent, preparation VEGF and bovine serum albumin aqueous solution, wherein the mass concentration of VEGF is 15~25 μ g/ml, the mass concentration of bovine serum albumin is 0.6~1mg/ml;
(3) said regenerated silk fibroin water solution is mixed with 60: 1~130: 1 volume ratio with said VEGF and bovine serum albumin aqueous solution, preparation contains the regenerated silk fibroin solution of VEGF;
(4) the regenerated silk fibroin solution that contains VEGF with step (3) preparation is spinning liquid, adopts electrostatic spinning technique that it is sprayed on the acellular matrix, prepares to be used for the tissue engineering bracket that urethra is rebuild.
7. a kind of method for preparing that is used for the tissue engineering bracket that urethra rebuilds according to claim 6 is characterized in that, described Bombyx bombycis is a kind of in mulberry cocoon, tussah cocoon or the Semen Ricini Bombyx bombycis; Described acellular matrix is an acellular matrix under acellular matrix or the bladder mucosa under the small intestinal mucosa.
8. a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction according to claim 6 is characterized in that the spinning ambient temperature when described electrostatic spinning technique prepares the tissue engineering bracket that is used for the urethra reconstruction is 2~6 ℃.
9. a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction according to claim 6 is characterized in that, further, the described tissue engineering bracket that is used for the urethra reconstruction is carried out post processing; Described post processing is meant soaks 30~90min with the tissue engineering bracket that is used for the urethra reconstruction at the alcohol-water mixed solution.
10. a kind of method for preparing that is used for the tissue engineering bracket of urethra reconstruction according to claim 9 is characterized in that ethanol accounts for 75~95% of alcohol-water mixed solution cumulative volume in the described alcohol-water mixed solution.
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CN104473706A (en) * 2014-12-09 2015-04-01 金华市人民医院 Biodegradable composite type tubular urethral stent and preparation method
CN105505883A (en) * 2016-01-12 2016-04-20 河南师范大学 Optimized culture method for primary rat hepatocytes
CN106075586A (en) * 2016-07-19 2016-11-09 东华大学 The cultural method of artificial liver repair materials based on adult stem cell regenerated silk fibroin tissue engineering bracket
CN107551322A (en) * 2017-10-31 2018-01-09 无锡中科光远生物材料有限公司 A kind of compound support frame material for Bladder repair
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CN109847101A (en) * 2018-12-28 2019-06-07 广州市妇女儿童医疗中心 A kind of Tissue Engineering Urethra bracket and its preparation process
CN110538348A (en) * 2018-06-05 2019-12-06 广州市妇女儿童医疗中心 Urethra tissue engineering scaffold and preparation method thereof
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CN115487359A (en) * 2022-09-26 2022-12-20 中国人民解放军总医院第一医学中心 Novel bladder tissue engineering patch for promoting angiogenesis, and preparation method and application thereof
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CN106075586A (en) * 2016-07-19 2016-11-09 东华大学 The cultural method of artificial liver repair materials based on adult stem cell regenerated silk fibroin tissue engineering bracket
CN106075586B (en) * 2016-07-19 2019-05-24 东华大学 Cultural method based on adult stem cell-regenerated silk fibroin tissue engineering bracket artificial liver repair materials
CN107551322A (en) * 2017-10-31 2018-01-09 无锡中科光远生物材料有限公司 A kind of compound support frame material for Bladder repair
CN110538348A (en) * 2018-06-05 2019-12-06 广州市妇女儿童医疗中心 Urethra tissue engineering scaffold and preparation method thereof
CN109248341A (en) * 2018-11-20 2019-01-22 北京航空航天大学 A kind of soft tissue repair sticking patch and preparation method thereof
CN109276759A (en) * 2018-11-27 2019-01-29 北京航空航天大学 A kind of bionical antibacterial stomach wall repair materials
CN109276759B (en) * 2018-11-27 2020-11-24 北京航空航天大学 Bionic antibacterial abdominal wall repair material
CN109847101A (en) * 2018-12-28 2019-06-07 广州市妇女儿童医疗中心 A kind of Tissue Engineering Urethra bracket and its preparation process
WO2023284329A1 (en) * 2021-07-15 2023-01-19 南通纺织丝绸产业技术研究院 Urethral stent having long-acting antibacterial and anti-stenotic functions and preparation method therefor
CN114949322A (en) * 2022-04-13 2022-08-30 齐齐哈尔医学院 Preparation method of tissue-induced nanofiber biomaterial capable of trapping host cells by graphene quantum dot tweezers and homing
CN115414533A (en) * 2022-09-20 2022-12-02 中国人民解放军总医院第一医学中心 Tissue engineering scaffold for simulating physiological structure of bladder wall and preparation method thereof
CN115414533B (en) * 2022-09-20 2024-02-23 中国人民解放军总医院第一医学中心 Tissue engineering scaffold simulating physiological structure of bladder wall and preparation method thereof
CN115487359A (en) * 2022-09-26 2022-12-20 中国人民解放军总医院第一医学中心 Novel bladder tissue engineering patch for promoting angiogenesis, and preparation method and application thereof

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