CN113713178A - Regeneration promoting and displacement preventing artificial nucleus prosthesis and preparation method thereof - Google Patents
Regeneration promoting and displacement preventing artificial nucleus prosthesis and preparation method thereof Download PDFInfo
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/18—Macromolecular materials obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
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- A61L27/16—Macromolecular materials obtained by reactions only involving carbon-to-carbon unsaturated bonds
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- A—HUMAN NECESSITIES
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- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/20—Polysaccharides
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/222—Gelatin
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/227—Other specific proteins or polypeptides not covered by A61L27/222, A61L27/225 or A61L27/24
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
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- A—HUMAN NECESSITIES
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/54—Biologically active materials, e.g. therapeutic substances
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/58—Materials at least partially resorbable by the body
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
- A61L2300/412—Tissue-regenerating or healing or proliferative agents
- A61L2300/414—Growth factors
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/38—Materials or treatment for tissue regeneration for reconstruction of the spine, vertebrae or intervertebral discs
Abstract
The invention discloses an artificial nucleus pulposus prosthesis capable of promoting regeneration and preventing displacement and a preparation method thereof. The nucleus pulposus prosthesis is designed into a structure with an inner layer which can be injected with in-situ polymerized hydrogel and an outer layer which is a zipper-type embedded composite capsule. The specific method comprises the steps of obtaining a degradable regeneration promoting film and a non-degradable elastic film with a microcolumn structure on one side by using a template method, carrying out embedding and hot pressing on the two films to obtain a composite capsule, and then injecting hydrogel. The structure of hydrogel is wrapped by the capsule body, so that the overflow of the hydrogel injected into the capsule body can be effectively prevented. The capsule is designed into a zipper-type embedded structure, firstly, the outer membrane of the capsule can release growth factors to promote the endogenous growth of nucleus pulposus cells in the degradation process; and secondly, the gradually exposed capsule inner membrane with the concave-convex structure can increase the friction force between the nucleus pulposus prosthesis and the inner wall of the fibrous ring, plays a role in fixing, and solves the problems of instability and displacement in the disc.
Description
Technical Field
The invention relates to the field of implant materials, in particular to an artificial nucleus pulposus prosthesis capable of promoting regeneration and preventing displacement and a preparation method thereof.
Background
Degenerative disc disease is the leading cause of most low back pain and is one of the chronic diseases that accompany aging. Most degenerative disc disease is usually due to degeneration, bulging of the nucleus pulposus and rupture of the annulus fibrosus. Aging and apoptosis of nucleus pulposus cells can lead to degenerative changes such as water loss, poor elasticity and even fibrosis. At the same time, this is likely to cause increased pressure and even rupture of the peripheral annulus, eventually leading to herniated discs and compression of nerves and thus pain.
The treatment of degenerative diseases such as disc herniation should take into account both the nucleus pulposus and the annulus fibrosus. The common treatment mode in the existing semi-molded artificial nucleus pulposus is to inject the in-situ solidifiable high molecular hydrogel into the vertebral body after the balloon is placed into the vertebral body so as to expand the balloon. The addition of the balloon can effectively reduce the possibility of hydrogel leakage from the incision of the annulus fibrosus. However, the success of the implantation of artificial nuclei in terms of reparative properties is limited because they do not modify the underlying disease nor promote tissue integration and subsequent primary biomechanics. Moreover, artificial nucleus prostheses have a variety of problems after implantation, such as instability and displacement within the disc due to the size of the nucleus prosthesis not being compatible with the size of the nucleus cavity.
In view of the above problems, there are also related patents which propose solutions, for example, patent (patent publication: CN 201810457276.0) designs an artificial nucleus pulposus carrying fullerene xerogel wrapped by an electrospun fibrous membrane. The addition of fullerene can reduce the content of free radicals in nucleus pulposus to a certain extent, so as to achieve the purpose of slowing down the degeneration of intervertebral disc, but does not achieve the effect of promoting the growth of original tissues. The patent (patent publication: CN 201310308397.6) utilizes 3D printing technology to customize an artificial nucleus pulposus that meets the requirements according to the nucleus pulposus size of different patients. The problems of intradiscal instability and displacement caused by the fact that the size of the nucleus pulposus prosthesis is not consistent with the size of the nucleus pulposus cavity are effectively solved, but the personalized customization mode is too limited and is not suitable for general application. In addition, the above patents only slow down the degeneration or limitation of the tissues within the disc to solve the problem of instability in the disc after the nucleus is implanted, and do not fundamentally promote the endogenous growth of nucleus cells and solve the general problem of the occurrence of displacement after the artificial nucleus is implanted.
The invention designs the artificial nucleus prosthesis into a structure with an inner layer which can be injected with in-situ polymerized hydrogel and an outer layer which is a zipper-type embedded composite capsule. The capsule body is designed into a 'zipper type' embedded structure, the outer membrane of the capsule body can release growth factors to promote the endogenous growth of nucleus pulposus cells in the degradation process, and meanwhile, the exposed inner membrane with the concave-convex structure can increase the friction force between the nucleus pulposus prosthesis and the inner wall of the fiber ring, so that the fixing effect is realized. Therefore, the invention can promote the endogenous regeneration of nucleus pulposus cells to a certain extent and can prevent the displacement of the nucleus pulposus prosthesis after being implanted.
Disclosure of Invention
It is an object of the present invention to provide a regeneration-promoting, anti-migration artificial nucleus prosthesis in order to solve the above-mentioned technical problems.
The invention also aims to provide a preparation method of the regeneration promoting and displacement preventing artificial nucleus prosthesis.
The invention is realized by the following technical scheme:
an artificial nucleus pulposus prosthesis for promoting regeneration and preventing displacement and a preparation method thereof are carried out according to the following operation steps:
(1) preparation of Polydimethylsiloxane (PDMS) template
And pouring a certain amount of PDMS on the template by taking the photoresist with the micro-column characteristics as the template, and curing to obtain the PDMS groove template with the shape opposite to that of the column array.
In order to better realize the invention, further, the thickness of the obtained template groove of the PDMS template is 350-.
(2) Preparation of 'zipper type' embedded composite film
Dissolving 0.03-0.07 g of hyaluronic acid in 15 mL of deionized water, and stirring for a period of time until the hyaluronic acid is completely dissolved; dissolving 0.5-1.2 g of gelatin in 15 mL of deionized water, and stirring until the gelatin is dissolved; mixing the two solutions, adding 0.3-0.5g of soybean protein isolate and 0.2-0.5% of growth factor by mass, and stirring uniformly. And pouring the mixed solution into a PDMS groove template, removing the redundant part and curing to obtain the degradable regeneration promoting film with the microcolumns on one side. Pouring the polyurethane into the PDMS groove template, removing the redundant part and curing to obtain the nondegradable high-elasticity film with the microcolumns on one side.
Embedding and hot-pressing the degradable regeneration promoting film and the non-degradable elastic film with the microcolumns at the temperature of 40-100 ℃ and the pressure of 2-25 MPa to obtain the zipper type embedded composite film. The outer membrane loaded with the growth factors has good biocompatibility and degradability, and can release the growth factors to promote the endogenous growth of nucleus pulposus cells in the degradation process; the inner layer is a polyurethane film, which can provide certain support and high elasticity.
In order to better implement the invention, further, the molecular weight of hyaluronic acid is (10-13), (80-100), (130) -150) multiplied by 104 One of daltons (Da).
In order to better realize the invention, the growth factor is one or more of insulin-like growth factor 1, exogenous transforming growth factor beta 1, growth differentiation factor 5 and fibroblast growth factor 18.
To better implement the present invention, further, the film thickness of the resulting "zipper-type" embedded composite film is 480 μm to 2000 μm.
In the examples of the present invention, the growth factors are insulin-like growth factor 1 and hyaluronic acid with molecular weight of 130-4 Da。
(3) Preparation of hydrogels
The injectable in-situ polymerized hydrogel can be prepared into one or more of polyvinyl alcohol hydrogel, polyacrylamide hydrogel and polyacrylic hydrogel.
And pumping out air in the composite capsule body by using an injector, implanting the capsule body into the intervertebral disc space by using a conveying tool, injecting 0.5-5.0 mL of the hydrogel polymer into the capsule body, and obtaining the artificial nucleus pulposus prosthesis after in-situ forming. The structure of the balloon-wrapped hydrogel prevents the risk of spillage of the internally injected hydrogel when it expands to completely fill the outer balloon.
The invention has the advantages of
(1) The artificial nucleus pulposus prosthesis is designed into a structure that the inner layer is injectable in-situ polymerized hydrogel and the outer layer is a zipper-type embedded composite capsule, so that the phenomenon that the injected hydrogel overflows when the injected hydrogel is expanded to completely fill the outer capsule can be effectively prevented.
(2) The capsule body is designed into a zipper-type embedded composite double-layer membrane structure by utilizing a template method, and after the capsule body is implanted, growth factors can be released from the outer membrane of the capsule body in the degradation process to promote the endogenous growth of nucleus pulposus cells; meanwhile, the surface concave-convex structure of the gradually exposed capsule inner membrane can increase the friction force between the nucleus pulposus prosthesis and the inner wall of the fibrous ring, so that the fixation effect is achieved, and the problems of instability and displacement in the disc are solved.
Drawings
FIG. 1 is a schematic view of a process for preparing a "zipper type" embedded composite film.
FIG. 2 is a schematic view of a regeneration promoting, displacement preventing artificial nucleus prosthesis.
Detailed Description
The present invention will be described in further detail with reference to specific examples. The description is illustrative of the invention and is not to be construed as limiting.
Preparation of regeneration promoting and displacement preventing artificial nucleus pulposus prosthetic material
Example 1:
the PDMS template is obtained by a template method, the thickness of the groove of the obtained template is 350 μm, the height of the micro-column is 140 μm, the distance between the columns is 130 μm, and the diameter of the columns is 130 μm. Dissolving 0.03 g of hyaluronic acid in 15 mL of deionized water, and stirring for 1 h until the hyaluronic acid is completely dissolved; dissolving 0.5g of gelatin in 15 mL of deionized water, and stirring at 40 ℃ until the gelatin is dissolved; mixing the above two solutions, adding 0.3 g of soybean protein isolate and 0.2% of growth factor, and stirring. And pouring the mixed solution into a PDMS groove template, removing the redundant part and curing to obtain the degradable regeneration promoting film with the microcolumns on one side. And pouring the polyurethane solution into the PDMS groove template, removing the redundant part and curing to obtain the nondegradable high-elasticity film with the microcolumns on one side. Embedding the degradable regeneration promoting film and the non-degradable elastic film with the micro-columns at 65 ℃ and 15MPa, and hot-pressing to obtain the 'zipper type' embedded composite film with the thickness of 480 mu m.
Dispersing 0.02 g of GO in 20 mL of deionized water, adjusting the pH value of a GO aqueous solution to 10, stirring uniformly, and performing ultrasonic dispersion for 45 min; 0.4 g of PVA is dissolved in 10mL of deionized water, and the mixture is heated to 80 ℃ until the PVA is dissolved to obtain a PVA aqueous solution with the mass fraction of 4%. And (2) uniformly mixing the two solutions, adding 0.9 g of AM monomer, 1 mL of 2 mg/mL MBA serving as a cross-linking agent and 4.5 mg of APS serving as an initiator into the solution, continuously stirring for 2 hours, ultrasonically removing bubbles, and heating and polymerizing at 40 ℃ to obtain the PVA-GO-PAM hydrogel.
And pumping out air in the composite capsule body by using an injector, implanting the capsule body into the intervertebral disc space by using a conveying device, injecting 3 mL of hydrogel polymer into the capsule body, and obtaining the artificial nucleus pulposus prosthesis after forming.
Example 2:
the PDMS template is obtained by a template method, the thickness of the groove of the obtained template is 350 μm, the height of the micro-column is 140 μm, the distance between the columns is 130 μm, and the diameter of the columns is 130 μm. Dissolving 0.07 g of hyaluronic acid in 15 mL of deionized water, and stirring for 1 h until the hyaluronic acid is completely dissolved; dissolving 1.2 g of gelatin in 15 mL of deionized water, and stirring at 40 ℃ until the gelatin is dissolved; mixing the above two solutions, adding 0.5g of soybean protein isolate and 0.5% of growth factor, and stirring. And pouring the mixed solution into a PDMS groove template, removing the redundant part and curing to obtain the degradable regeneration promoting film with the microcolumns on one side. And pouring the polyurethane solution into the PDMS groove template, removing the redundant part and curing to obtain the nondegradable high-elasticity film with the microcolumns on one side. Embedding the degradable regeneration promoting film and the non-degradable elastic film with the micro-columns at 65 ℃ and 15MPa, and hot-pressing to obtain the 'zipper type' embedded composite film with the thickness of 480 mu m.
Dispersing 0.05 g of GO in 20 mL of deionized water, adjusting the pH value of a GO aqueous solution to 10, stirring uniformly, and performing ultrasonic dispersion for 45 min; 1.0 g of PVA is dissolved in 10mL of deionized water, and the mixture is heated to 80 ℃ until the PVA is dissolved to obtain a PVA aqueous solution with the mass fraction of 10%. And (2) uniformly mixing the two solutions, adding 1.8 g of AM monomer, 1 mL of 2 mg/mL MBA serving as a cross-linking agent and 9 mg of APS serving as an initiator into the solution, continuously stirring for 2 hours, ultrasonically removing bubbles, and heating and polymerizing at 40 ℃ to obtain the PVA-GO-PAM hydrogel.
And pumping out air in the composite capsule body by using an injector, implanting the capsule body into the intervertebral disc space by using a conveying device, injecting 3 mL of hydrogel polymer into the capsule body, and obtaining the artificial nucleus pulposus prosthesis after forming.
Example 3:
the PDMS template is obtained by a template method, the thickness of the groove of the obtained template is 350 μm, the height of the micro-column is 140 μm, the distance between the columns is 130 μm, and the diameter of the columns is 130 μm. Dissolving 0.05 g of hyaluronic acid in 15 mL of deionized water, and stirring for 1 h until the hyaluronic acid is completely dissolved; dissolving 0.8 g of gelatin in 15 mL of deionized water, and stirring at 40 ℃ until the gelatin is dissolved; mixing the above two solutions, adding 0.4 g of soybean protein isolate and 0.3% of growth factor, and stirring. And pouring the mixed solution into a PDMS groove template, removing the redundant part and curing to obtain the degradable regeneration promoting film with the microcolumns on one side. And pouring the polyurethane solution into the PDMS groove template, removing the redundant part and curing to obtain the nondegradable high-elasticity film with the microcolumns on one side. Embedding the degradable regeneration promoting film and the non-degradable elastic film with the micro-columns at 65 ℃ and 15MPa, and hot-pressing to obtain the 'zipper type' embedded composite film with the thickness of 480 mu m.
Dispersing 0.03 g of GO in 20 mL of deionized water, adjusting the pH value of a GO aqueous solution to 10, stirring uniformly, and performing ultrasonic dispersion for 45 min; 0.7 g of PVA is dissolved in 10mL of deionized water, and the mixture is heated to 80 ℃ until the PVA is dissolved to obtain a PVA aqueous solution with the mass fraction of 7%. And (2) uniformly mixing the two solutions, adding 1.5 g of AM monomer, 1 mL of 2 mg/mL MBA serving as a cross-linking agent and 7.5 mg of APS serving as an initiator into the solution, continuously stirring for 2 hours, ultrasonically removing bubbles, and heating and polymerizing at 40 ℃ to obtain the PVA-GO-PAM hydrogel.
And pumping out air in the composite capsule body by using an injector, implanting the capsule body into the intervertebral disc space by using a conveying device, injecting 3 mL of hydrogel polymer into the capsule body, and obtaining the artificial nucleus pulposus prosthesis after forming.
Comparative example 1
Dissolving 0.07 g of hyaluronic acid in 15 mL of deionized water, and stirring for 1 h until the hyaluronic acid is completely dissolved; dissolving 1.2 g of gelatin in 15 mL of deionized water, and stirring at 40 ℃ until the gelatin is dissolved; mixing the above two solutions, adding 0.5g of soybean protein isolate and 0.5% of growth factor, and stirring. And obtaining the degradable regeneration promoting film by using a tape casting method. Dispersing 0.05 g of GO in 20 mL of deionized water, adjusting the pH value of a GO aqueous solution to 10, stirring uniformly, and performing ultrasonic dispersion for 45 min; 1.0 g of PVA is dissolved in 10mL of deionized water, and the mixture is heated to 80 ℃ until the PVA is dissolved to obtain a PVA aqueous solution with the mass fraction of 10%. And (2) uniformly mixing the two solutions, adding 1.8 g of AM monomer, 1 mL of 2 mg/mL MBA serving as a cross-linking agent and 9 mg of APS serving as an initiator into the solution, continuously stirring for 2 hours, ultrasonically removing bubbles, and heating and polymerizing at 40 ℃ to obtain the PVA-GO-PAM hydrogel. And pumping out air in the composite capsule body by using an injector, implanting the capsule body into the intervertebral disc space by using a conveying device, injecting 3 mL of hydrogel polymer into the capsule body, and obtaining the artificial nucleus pulposus prosthesis after forming.
Comparative example 2
The non-degradable elastic film is prepared by taking polyurethane as a raw material. Dispersing 0.05 g of GO in 20 mL of deionized water, adjusting the pH value of a GO aqueous solution to 10, stirring uniformly, and performing ultrasonic dispersion for 45 min; 1.0 g of PVA is dissolved in 10mL of deionized water, and the mixture is heated to 80 ℃ until the PVA is dissolved to obtain a PVA aqueous solution with the mass fraction of 10%. And (2) uniformly mixing the two solutions, adding 1.8 g of AM monomer, 1 mL of 2 mg/mL MBA serving as a cross-linking agent and 9 mg of APS serving as an initiator into the solution, continuously stirring for 2 hours, ultrasonically removing bubbles, and heating and polymerizing at 40 ℃ to obtain the PVA-GO-PAM hydrogel. And pumping out air in the composite capsule body by using an injector, implanting the capsule body into the intervertebral disc space by using a conveying device, injecting 3 mL of hydrogel polymer into the capsule body, and obtaining the artificial nucleus pulposus prosthesis after forming.
Adaptation, safety and efficacy testing of an artificial nucleus prosthesis
The method comprises the following steps: the artificial nucleus prosthesis obtained in example 3 was subjected to an adaptation study. The tail cone of the newly slaughtered adult cattle is taken for processing and standby. A small incision is made on the annulus fibrosus with a scalpel, and the diameter of the incision is 2-3 mm. Digging out 3-4 mL of nucleus pulposus by using the nucleus pulposus spoon, implanting the composite capsule body by using the conveying device, injecting hydrogel into the cavity of the capsule body, and completing the implantation of the artificial nucleus pulposus prosthesis after the hydrogel is polymerized.
The result shows that the artificial nucleus pulposus prosthesis is implanted into the nucleus pulposus cavity in a way that the hydrogel is wrapped by the balloon, the wound is small, after the hydrogel is expanded, the external balloon can fill the whole nucleus pulposus cavity, and the artificial nucleus pulposus prosthesis has excellent adaptability.
The artificial nucleus pulposus prostheses obtained in the examples 1 to 3 and the comparative examples 1 to 2 are implanted into the nucleus pulposus defect of the intervertebral disc and then cultured in vitro for one week, and then tested for safety and effectiveness.
The method comprises the following steps: and (3) performing cell survival rate characterization on the artificial nucleus pulposus prosthesis by adopting an MTT method. The artificial nucleus pulposus prostheses obtained in examples 1 to 3 and comparative examples 1 to 2 were cultured in a 96-well plate, and then the viability of the nucleus pulposus cells in the nucleus pulposus prostheses was examined by using the MTT method. After one week of culture, the dead cells were removed by removing the old medium and washing with Phosphate Buffered Saline (PBS), and 200. mu.L of serum-free medium and 50. mu.L of MTT solution were added to each well and incubated at 37 ℃ for 4 hours. Then, 200. mu.L of dimethyl sulfoxide (DMSO) was added to each well of the old medium to dissolve the formazan crystal sufficiently, and 100. mu.L of the solution was measured for absorbance at a wavelength of 500 nm by a microplate reader.
According to GB16886.6 "biological evaluation of medical devices: inflammation degree test is required in a local reaction test after implantation; characterizing the degradation degree of the outer membrane of the capsule body by a weighing method; the bonding strength between the capsule body of the artificial nucleus pulposus prosthesis and the inner wall of the fibrous ring is represented by a tensile machine, and the test result is represented by peel strength; the compression strength and the elastic modulus of the nucleus prosthesis are tested by adopting a biomechanics testing machine, the compression height ratio is set to be 85 percent, and the compression speed is 3 mm/min; the results are summarized in the following table.
The following conclusions can be drawn from the above experimental data:
the capsule body of the comparative example 1 only consists of the degradable membrane, the degradation rate can reach 38 +/-4%, the MTT absorbance is 1.35 +/-0.5, growth factors can be released in the degradation process to promote the proliferation of nucleus pulposus cells, but the capsule body has lower compressive strength, elastic modulus and peel strength and relatively poorer mechanical property. Comparative example 2 the capsule body consists of polyurethane only, has relatively high mechanical strength, particularly has an elastic modulus of 3.62 +/-1 MPa, but cannot promote the proliferation of nucleus pulposus cells.
The compressive strength and elastic modulus of examples 1-3 show that the average compressive strength and elastic modulus are 3.07 + -1.55 MPa and 3.39 + -1 MPa after one week of nucleus pulposus implantation, which are improved to some extent compared with the defective nucleus pulposus; meanwhile, the average degradation degree of the embodiments 1 to 3 is 36 +/-4%, the MTT absorbance is 1.25 +/-0.5 and the peel strength is 0.15 +/-0.01 MPa, and the analysis of experimental data shows that the growth factors released by the degradable membrane in the degradation process promote the proliferation of nucleus pulposus cells, and meanwhile, the exposed inner membrane with the concave-convex structure increases the friction force between the artificial nucleus pulposus prosthesis and the fibrous ring.
The result shows that the artificial nucleus pulposus prosthesis with the zipper-type embedded composite capsule structure has the advantages of promoting the growth of nucleus pulposus cells and simultaneously preventing the displacement of the nucleus pulposus prosthesis.
While the present invention has been described in detail with reference to the preferred embodiments, it should be understood that the above description should not be taken as limiting the invention. Various modifications and alterations to this invention will become apparent to those skilled in the art upon reading the foregoing description. Accordingly, the scope of the invention should be determined from the following claims.
Claims (8)
1. The artificial nucleus pulposus prosthesis is characterized in that the artificial nucleus pulposus prosthesis is designed into a structure with an inner layer which can be injected with in-situ polymerized hydrogel and an outer layer which is a zipper-type embedded composite capsule body, the composite capsule body is formed by embedding two structures of a degradable regeneration promoting film and a non-degradable elastic film, and the preparation of the artificial nucleus pulposus prosthesis is carried out according to the following steps:
(1) taking photoresist with micro-column characteristics as a template, pouring Polydimethylsiloxane (PDMS) on the template and curing to obtain a PDMS groove template with a shape opposite to that of the column array;
(2) stirring hyaluronic acid solution, gelatin solution, soy protein isolate and growth factors until the hyaluronic acid solution, the gelatin solution, the soy protein isolate and the growth factors are uniformly mixed, pouring a proper amount of mixed solution into the PDMS template obtained in the step (1), and curing to obtain a degradable regeneration promoting film with a micro-column structure on one side;
(3) pouring polyurethane into the PDMS template obtained in the step (1) and curing to obtain a non-degradable high-elasticity film with a micro-column structure on one side;
(4) embedding and hot-pressing the two films obtained in the step (2) and the step (3) by utilizing a single-side micro-column structure to obtain a zipper type embedded composite film with regeneration promoting and displacement preventing functions;
(5) preparing injectable in-situ polymerized hydrogel;
(6) and (3) pumping out air in the composite capsule body by using an injector, implanting the capsule body into the intervertebral disc space by using a conveying tool, injecting the hydrogel polymer obtained in the step (5) into the cavity of the capsule body, and obtaining the artificial nucleus pulposus prosthesis after the hydrogel is polymerized and formed in situ.
2. The method for preparing an artificial nucleus pulposus prosthesis with regeneration promotion and displacement prevention functions as claimed in claim 1, wherein the groove thickness of the PDMS template in the step (1) is 520 μm-.
3. The method for preparing the regeneration-promoting and displacement-preventing artificial nucleus pulposus prosthesis according to claim 1, wherein the mass fractions of the hyaluronic acid solution, the gelatin solution, the isolated soy protein and the growth factor in the step (2) are 0.1% -0.23%, 1.6% -4%, 1.0% -1.6% and 0.2% -0.5%, respectively.
4. The method for preparing a regeneration-promoting and displacement-preventing artificial nucleus pulposus prosthesis according to claim 1, wherein the growth factor in the step (2) is one or more of insulin-like growth factor 1, exogenous transforming growth factor beta 1, growth differentiation factor 5 and fibroblast regeneration factor 18.
5. The method for preparing the regeneration-promoting and displacement-preventing artificial nucleus pulposus prosthesis according to the claim 1, wherein the parameters of the embedding and hot pressing of the zipper-type embedded composite membrane in the step (4) are as follows: the temperature is 40-100 ℃, and the pressure is 2-25 MPa.
6. The method for preparing the regeneration-promoting and displacement-preventing artificial nucleus pulposus prosthesis according to claim 1, wherein the film thickness of the "zipper-type" embedded composite film in the step (4) is 480 μm to 2000 μm.
7. The method for preparing a regeneration-promoting and displacement-preventing artificial nucleus pulposus prosthesis according to claim 1, wherein the injectable in-situ polymerized hydrogel in the step (5) is one or a combination of polyvinyl alcohol, polyacrylamide and polyacrylic hydrogel.
8. The method of claim 1, wherein the hydrogel polymer in the infusion balloon of step (6) has a volume of 0.5-5.0 mL.
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