CN107233143A - Remove cell corneal stroma lens and preparation method thereof - Google Patents
Remove cell corneal stroma lens and preparation method thereof Download PDFInfo
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- CN107233143A CN107233143A CN201710403038.7A CN201710403038A CN107233143A CN 107233143 A CN107233143 A CN 107233143A CN 201710403038 A CN201710403038 A CN 201710403038A CN 107233143 A CN107233143 A CN 107233143A
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
- A61F2/02—Prostheses implantable into the body
- A61F2/14—Eye parts, e.g. lenses, corneal implants; Implanting instruments specially adapted therefor; Artificial eyes
- A61F2/16—Intraocular lenses
- A61F2/1602—Corrective lenses for use in addition to the natural lenses of the eyes or for pseudo-phakic eyes
-
- 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
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3604—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix characterised by the human or animal origin of the biological material, e.g. hair, fascia, fish scales, silk, shellac, pericardium, pleura, renal tissue, amniotic membrane, parenchymal tissue, fetal tissue, muscle tissue, fat tissue, enamel
-
- 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
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3641—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix characterised by the site of application in the body
-
- 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
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3683—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment
- A61L27/3687—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment characterised by the use of chemical agents in the treatment, e.g. specific enzymes, detergents, capping agents, crosslinkers, anticalcification agents
-
- 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
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3683—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment
- A61L27/3691—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment characterised by physical conditions of the treatment, e.g. applying a compressive force to the composition, pressure cycles, ultrasonic/sonication or microwave treatment, lyophilisation
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
- A61F2240/00—Manufacturing or designing of prostheses classified in groups A61F2/00 - A61F2/26 or A61F2/82 or A61F9/00 or A61F11/00 or subgroups thereof
- A61F2240/001—Designing or manufacturing processes
- A61F2240/002—Designing or making customized prostheses
-
- 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
- A61L2430/16—Materials or treatment for tissue regeneration for reconstruction of eye parts, e.g. intraocular lens, cornea
Abstract
Cell corneal stroma lens and preparation method thereof are removed the invention provides one kind, are related to the technical field of organizational engineering.What the present invention was provided goes the preparation method of cell corneal stroma lens, cell component in matrix can not only effectively be removed, reduce the immunogenicity of corneal stroma lens, improve the mechanical strength of corneal lens, more can effectively keep the original form and transparency of corneal stroma, it is obtained go cell corneal stroma lens have that security is higher, transparency more preferably with performance it is more stable the characteristics of；And the support of growth can be bred as keratocyte, finally combined together with auto corneal, permanent implanted lens can be used as；In addition it is possible to avoid the appearance of the phenomenons such as immunological rejection.
Description
Technical field
The present invention relates to organizational engineering technical field, cell corneal stroma lens and its preparation are removed more particularly, to one kind
Method.
Background technology
Long sight is that parallel rays enters a kind of refractive status after retina after intraocular, when the refractive power of eyeball is not enough
Or just produce long sight during its axiallength deficiency.Long sight is used as one kind of ophthalmology disease, the strong influence life matter of people
Amount, especially with the increase at age, the occurrence probability of long sight can also be raised.
Long sight is clinical common ametropia illness in eye, and the correction of hyperpresbyopia is always the problem of refractive therapy.At present
The method of distance vision correction treatment mainly includes frame eyeglasses, contact lens or refractive surgery.
It is ametropia that presbyopia is that a kind of physiological phenomenon is not that pathological state is also not belonging to, and it is inevitable after person in middle and old age to be that people step into
The visual problem of appearance.With advancing age, there is the patient of presbyopia and must increase convex lens could obtaining clearly near-sighted
Power.
At present, it is made more than the contact lens of implanted of synthetic material, can be according to the refractive status of different patients
The lens of the various number of degrees and model size are produced, Postoperative visual acuity recovers fast, but its drawback is also obvious, such as：It is postoperative can
Astigmatism increase can occur, the vaporific muddiness of cornea is likely to occur in early days；And its applicable disease is limited, and there is the related wind of cornea flap
Danger, is likely to occur corneal injury, the situation that visual quality declines during being chronically implanted；In addition, its security is relatively low, belong to
Permanent foreign body, is likely to occur immunological rejection during being chronically implanted.Therefore, for hyperpresbyopia, presbyopia and merging
Anisometropic patient is, it is necessary to find that indication is wider, security is higher, predictability is stronger, stability more preferably lens material.
For the material needed for corneal stroma is transplanted, good optical characteristics is not required nothing more than, is also required and human eye
Tissue has good histocompatbility, the drawbacks of synthetic material then has very big in biological tissue's compatibility, and utilizes dynamic
The cornea of material resource, then the problem of can largely alleviating the histocompatbility difference of synthetic material.
But using animal derived cornea there is also some problems, for example：(1) although the tissue compatible of animal derived cornea
Property preferably, but if its own cell removes incomplete, then can have the risk for producing immune response；(2) in addition, although at present
The method of some existing removal keratocytes, but existing cornea goes cellular matrix preparation technology to be easily caused cornea suction
Oedema is swollen, causes arrangement of collagen fibers or conformation in matrix to change, and declines corneal stroma transparency, original form,
Curvature etc. changes；(3) moreover, traditional corneal lens is typically to be made with special keratome cutting, technique is relatively crude
Rough, it is not smooth enough to be likely to result in the face of cutting, or lens component is lost, and predictive and security is poor, and postoperative short-term is concurrent
Disease, such as astigmatism incidence is higher.
Therefore, the corneal stroma lens that security is higher, transparency is more stable more preferably with performance how to be obtained, it is still necessary to enter
Row constantly research.The processing method of safe and effective people's corneal stroma, in practical clinical, is significant.
In view of this, it is special to propose the present invention.
The content of the invention
First purpose of the present invention is to provide a kind of preparation method for removing cell corneal stroma lens, and of the invention the
Two purposes are that cell corneal stroma lens are removed in offer, low, saturating to alleviate corneal lens security present in prior art
Luminosity equation and the unstable technical problem of performance.
The invention provides a kind of preparation method for removing cell corneal stroma lens, including：By the cornea base with cell
Matter lens carry out cell cracking and crosslinking Treatment successively, then obtain described removing cell corneal stroma lens after sterilizing.
Further, the method for the cell cracking includes：With 0.9% life containing 0.1%-3%TritonX-100
Salt solution or phosphate buffer solution immersion 12-48h are managed, then 48- is rinsed with 0.9% physiological saline or phosphate buffer solution
96h。
Further, the method for the crosslinking includes：It is molten with 0.9% physiological saline or phosphate-buffered containing crosslinking agent
Soaked in liquid, then rinse 24-96h with 0.9% physiological saline or phosphate buffer solution, the crosslinking agent is 1- (3- bis-
Methylaminopropyl) -3- ethyl carbodiimides, N- hydroxy thiosuccinimides, Geniposide or glutaraldehyde.
Further, the mass ratio of the usage amount of the crosslinking agent and corneal stroma lens is 1:1-1:15.
Further, the mass ratio of the usage amount of the crosslinking agent and corneal stroma lens is 1:5-1:10.
Further, before cell cracking processing, in addition to disinfect, the method for the sterilization includes：With
The physiological saline immersion 1-5h that containing concentration be 0.01~0.1mg/ml penicillin and concentration is 0.05~0.5mg/ml streptomysins,
Then rinsed with 0.9% physiological saline or phosphate buffer solution.
Further, after the crosslinking Treatment, in addition to sterilization treatment, the method for the sterilizing includes：Penetrated with γ
Line is irradiated, and irradiation dose is 20~30kGy.
Further, the corneal stroma lens with cell are obtained by full femtosecond laser technology.
Further, the preparation method includes：
Step (a), sterilization：The corneal stroma that the exact thickness with cell is obtained by full femtosecond laser technology is saturating
Mirror, with being 0.01~0.1mg/ml penicillin containing concentration and physiological saline that concentration is 0.05~0.5mg/ml streptomysins soaks
1-5h, is then rinsed with 0.9% physiological saline or phosphate buffer solution；
Step (b), cell cracking：It is slow with 0.9% physiological saline or phosphate containing 0.1%-3%TritonX-100
Solution immersion 12-48h is rushed, then 48-96h is rinsed with 0.9% physiological saline；
Step (c), crosslinking：Soaked with 0.9% physiological saline or phosphate buffer solution containing crosslinking agent, Ran Houyong
0.9% physiological saline or phosphate buffer solution rinsing 24-96h, the crosslinking agent is 1- (3- dimethylamino-propyls) -3- second
Base carbodiimide, or N- hydroxy thiosuccinimides or glutaraldehyde or Geniposide；The usage amount of the crosslinking agent with it is described
The mass ratio that the corneal stroma lens of processing are cracked by cell is 1:1-1:15；
Step (d), sterilizing：Irradiated with gamma-rays, irradiation dose is 20~30kGy.
In addition, removing cell corneal stroma lens present invention also offers what is obtained according to described preparation method.
What the present invention was provided goes the preparation method of cell corneal stroma lens, can not only effectively remove in matrix cell into
The immunogenicity of point reduction matrix, improves the mechanical strength of cornea, more can effectively keep the original form of corneal stroma and transparent
Degree, it is obtained go cell corneal stroma lens have that security is higher, transparency more preferably with performance it is more stable the characteristics of.And
And, value-added support can be grown as keratocyte after the implantation of corneal stroma lens, keratocyte can be in lens
Migration and grow, finally combined together with auto corneal, can be clinically hyperpresbyopia and dioptric as permanent implanted lens
Irregular correction provides new treatment method.In addition, synthesis can be avoided the occurrence of using the lens as implanted contact lens
The permanent foreign matter situation of material class corneal lens, it is to avoid the appearance of the phenomenon such as immunological rejection.
Brief description of the drawings
, below will be to specific in order to illustrate more clearly of the specific embodiment of the invention or technical scheme of the prior art
The accompanying drawing used required in embodiment or description of the prior art is briefly described, it should be apparent that, in describing below
Accompanying drawing is some embodiments of the present invention, for those of ordinary skill in the art, before creative work is not paid
Put, other accompanying drawings can also be obtained according to these accompanying drawings.
Fig. 1 is the HE coloration result figures of the corneal stroma lens before cell；
Fig. 2 be Cell extraction after corneal stroma lens HE coloration result figures；
Fig. 3 is cell and the HE coloration result figures of corneal stroma lens after being crosslinked；
Fig. 4 is that the light transmittance of the corneal stroma lens before and after cell compares figure.
Embodiment
Technical scheme is clearly and completely described below in conjunction with embodiment, it is clear that described reality
It is a part of embodiment of the invention to apply example, rather than whole embodiments.Based on the embodiment in the present invention, the common skill in this area
The every other embodiment that art personnel are obtained under the premise of creative work is not made, belongs to the model that the present invention is protected
Enclose.
Existing corneal stroma lens go cell technique still immature, and obtained corneal stroma lens material has safety
The problem of lower, poor transparency of property and unstable performance.The present invention is carried aiming at these problems present in prior art
The improvement gone out.
The invention provides the preparation method for removing cell corneal stroma lens.
Wherein, the 0.1%-3%TritonX-100 being related in the method for cell cracking, for example can be, but be not limited to
0.1%TritonX-100,0.3%TritonX-100,0.5%TritonX-100,0.8%TritonX-100,1%
TritonX-100,1.5%TritonX-100,2%TritonX-100,2.5%TritonX-100 or 3%TritonX-
100。
Wherein, the crosslinking agent being related in the method for crosslinking for example can be, but be not limited to 1- (3- dimethylamino-propyls) -3-
Ethyl carbodiimide, either N- hydroxy thiosuccinimides or Geniposide, or glutaraldehyde.
Wherein, the usage amount for the crosslinking agent being related in the method for crosslinking is 1 with the mass ratio of corneal stroma lens:1-1:
15, its mass ratio such as can be, but be not limited to 1:1、1:2、1:3、1:4、1:5、1:6、1:7、1:8、1:9、1:10、1:10、
1:11、1:12、1:13、1:14 or 1:15.
Wherein, the concentration being related in the method for sterilization is 0.01~0.1mg/ml penicillin and concentration is 0.05~0.5mg/
The physiological saline or phosphate buffer solution of ml streptomysins, wherein penicillin for example can be, but be not limited to 0.01mg/ml,
0.03mg/ml、0.05mg/ml、0.07mg/ml、0.1mg/ml；Its streptomycin for example can be, but be not limited to 0.05mg/
ml、0.06mg/ml、0.07mg/ml、0.08mg/ml、0.09mg/ml、0.1mg/ml、0.2mg/ml、0.3mg/ml、0.4mg/
ml、0.5mg/ml。
Wherein, the dose of radiation being related in the method for sterilizing be 20~30kGy, for example can be, but be not limited to 20kGy,
21kGy, 22kGy, 23kGy, 24kGy, 25kGy, 26kGy, 27kGy, 28kGy, 29kGy or 30kGy.
Wherein, the corneal stroma lens with cell come from people or other animals (other animals for example can be, but
It is not limited to pig, ox or sheep etc.), it is preferred that the corneal stroma lens with cell come from people.
Wherein, the full femtosecond laser technology being related to, refers to utilize the small incision corneal matrix lens removal surgery of full femtosecond laser
The technology that (Small Incision Lenticule Extraction, SMILE) is cut.SMILE is to apply ultrashort pulse
Laser completes the scanning of two subpulses in cornea, makes in corneal stroma and is drawn off after eyeglass, can reduce corneal nerve
Damage and the biological influence with mechanics.
In addition, the corneal stroma lens with cell being related in the present invention refer to it is undressed, directly from people or
The fresh corneal stroma lens taken out in other animal eyes.
It should be noted that the method provided according to the present invention, results in the accurate lens of multi-thickness, it is preferred that
Thickness is 10-90 μm.
In order to contribute to the clearer understanding present invention, now present disclosure is carried out by specific embodiment detailed
Introduction.Pointed out as being not known, the Examination on experimental operation being related in following examples is conventional molecular biology or medical science behaviour
Make method, reagent, the instrument being related to are conventional commercial reagent or instrument.
Embodiment 1 goes the preparation method of cell corneal stroma lens
Step (a), sterilization：The corneal stroma of the exact thickness with cell obtained by full femtosecond laser technology is saturating
Mirror, with being 0.01mg/ml penicillin containing concentration and physiological saline that concentration is 0.1mg/ml streptomysins soaks 3h, Ran Houyong
0.9% physiological saline rinsing；
Step (b), cell cracking：24h is soaked with 0.9% physiological saline containing 0.5%TritonX-100, then
96h is rinsed with 0.9% physiological saline；
Step (c), crosslinking：Soaked with 0.9% physiological saline containing crosslinking agent, then use 0.9% physiological saline
24h is rinsed, crosslinking agent is 1- (3- dimethylamino-propyls) -3- ethyl carbodiimides (EDC)；The usage amount of crosslinking agent with through meticulous
The mass ratio of the corneal stroma lens of cellular lysate processing is 1:5；
Step (d), sterilizing：Corneal lens after crosslinking is irradiated with gamma-rays, irradiation dose is 25kGy；
It should be noted that in the present embodiment 1, the corneal stroma lens with cell come from people.
In addition, the performance for removing cell corneal stroma lens that inventor prepares to the method according to embodiment 1 is carried out
Analysis, specific experiment is as follows.
HE Coloration experiments
Respectively cell cornea base is removed to the corneal stroma lens with cell, according to what the method for embodiment 1 was prepared
Matter lens carry out HE dyeing, as a result respectively as shown in Figure 1, Figure 2 and Figure 3, are dyed from the HE of the corneal stroma lens with cell
Scheme to can see with the presence of more cell in (Fig. 1), and cell cornea base is removed from what the method according to embodiment 1 was prepared
Then it is substantially not visible in matter lens HE colored graphs (Fig. 2) between cell, but matrix fiber and there are some spaces；And according to embodiment 1
Method be crosslinked after obtain go in cell corneal stroma lens HE colored graphs (Fig. 3) be then substantially not visible space, explanation
The method that the present invention is provided can effectively remove the cell in corneal stroma lens, and keep corneal stroma lens morphology steady
It is fixed.
Light transmittance test experience
Using spectrophotometer visible light wave range (390nm-780nm) respectively to the corneal stroma lens with cell,
What the method according to embodiment 1 was prepared goes cell corneal stroma lens to carry out light transmittance analysis, as a result as shown in figure 4, pressing
The visible light transmittance rate of what the method according to embodiment 1 was prepared go cell corneal stroma lens substantially with fresh cornea matrix
(the corneal stroma lens i.e. with cell) unanimously, illustrating the light transmission of the diagonal membrane matrix lens of the technique influences smaller, energy
It is allowed to keep preferable light transmittance.
Wherein, in Fig. 4, it is cell corneal stroma lens to remove cell corneal stroma；Fresh cornea matrix is to carry
The corneal stroma lens of cell.
Embodiment 2 goes the preparation method of cell corneal stroma lens
Step (a), sterilization：The corneal stroma of the exact thickness with cell obtained by full femtosecond laser technology is saturating
Mirror, with being 0.05mg/ml penicillin containing concentration and physiological saline that concentration is 0.2mg/ml streptomysins soaks 2h, Ran Houyong
0.9% physiological saline rinsing；
Step (b), cell cracking：48h is soaked with 0.9% physiological saline containing 0.3%TritonX-100, then
96h is rinsed with 0.9% physiological saline；
Step (c), crosslinking：Soaked with 0.9% physiological saline containing crosslinking agent, then use 0.9% physiological saline
Rinse 24h, crosslinking agent N- hydroxy thiosuccinimides (NHS)；The usage amount of crosslinking agent and the angle that processing is cracked by cell
The mass ratio of membrane matrix lens is 1:9；
Step (d), sterilizing：Corneal lens after crosslinking is irradiated with gamma-rays, irradiation dose is 25kGy；
It should be noted that in the present embodiment 2, the corneal stroma lens with cell come from people.
Embodiment 3 goes the preparation method of cell corneal stroma lens
Step (a), sterilization：It is 0.1mg/ml moulds with containing concentration by the fresh corneal stroma lens with cell
Element and concentration soak 1h for the physiological saline of 0.5mg/ml streptomysins, then rinsed with 0.9% physiological saline；
Step (b), cell cracking：36h is soaked with 0.9% physiological saline containing 0.4%TritonX-100, then
72h is rinsed with 0.9% physiological saline；
Step (c), crosslinking：Soaked with 0.9% physiological saline containing crosslinking agent, then use 0.9% physiological saline
24h is rinsed, crosslinking agent is 1- (3- dimethylamino-propyls) -3- ethyl carbodiimides；The usage amount of crosslinking agent by cell with splitting
The mass ratio for solving the corneal stroma lens of processing is 1:10；
Step (d), sterilizing：Corneal lens after crosslinking is irradiated with gamma-rays, irradiation dose is 25kGy；
It should be noted that in the present embodiment 3, the corneal stroma lens with cell come from people.
The preparation method for removing cell corneal stroma lens provided according to the present invention, it is obtained to remove cell corneal stroma lens
With advantages below：
First, cell cracking and crosslinking are carried out in the diagonal membrane matrix lens of physiological saline environment, it is to avoid corneal stroma lens
There is water suction/dewatering state, change conformation and the arrangement of collagenous fibres, corneal stroma lens is effectively kept original
Form, thickness and curvature.The cell in diagonal membrane matrix lens is cracked and is crosslinked in this state, can not only effectively be removed
Go cell component in matrix to reduce the immunogenicity of matrix, improve the mechanical strength of cornea, more effectively corneal stroma can be kept saturating
The original form and transparency of mirror, so as to obtain the corneal stroma lens that security is higher, transparency is more stable more preferably with performance.
Secondly, value-added 3-dimensional support, keratocyte can be grown as keratocyte after the implantation of corneal stroma lens
It can migrate and grow in lens, finally be combined together with auto corneal, can be clinically high as permanent implanted lens
Spend long sight and anisometropic correction provides new treatment method.In addition, can be with using the lens as implanted contact lens
Avoid the occurrence of the permanent foreign matter situation of synthetic material class corneal lens, it is to avoid the appearance of the phenomenon such as immunological rejection.
In addition, full femtosecond laser technology can accurately be cut so that the preparation of corneal lens is more accurate, it can avoid passing
The corneal lens cutting face that system Cutting Process is caused is not smooth enough or lens component is lost；And the application then make use of full femtosecond
Laser technology is cut, and preparation technology is more accurate, predictive stronger.
Therefore, what the present invention was provided go, and cell corneal stroma lens have that security is higher, transparency more preferably with performance more
Stable the characteristics of.
Finally it should be noted that：Various embodiments above is merely illustrative of the technical solution of the present invention, rather than to the present invention's
Limitation；Although the present invention is described in detail with reference to foregoing embodiments, one of ordinary skill in the art should manage
Solution：It can still modify to the technical scheme described in foregoing embodiments, or to which part or whole skills
Art feature carries out equivalent substitution；And these modifications or replacement, the essence of appropriate technical solution is departed from each reality of the invention
Apply the scope of a technical scheme.
Claims (10)
1. a kind of preparation method for removing cell corneal stroma lens, it is characterised in that including：Corneal stroma with cell is saturating
Mirror carries out cell cracking and crosslinking Treatment successively, then obtains described removing cell corneal stroma lens after sterilizing.
2. preparation method according to claim 1, it is characterised in that the method for the cell cracking includes：With containing
0.1%-3%TritonX-100 0.9% physiological saline or phosphate buffer solution immersion 12-48h, then with 0.9% life
Manage salt solution or phosphate buffer solution rinsing 48-96h.
3. preparation method according to claim 2, it is characterised in that the method for the crosslinking includes：With containing crosslinking agent
0.9% physiological saline or phosphate buffer solution in soak, then with 0.9% physiological saline or phosphate buffer solution drift
Wash 24-96h, the crosslinking agent be 1- (3- dimethylamino-propyls) -3- ethyl carbodiimides, N- hydroxy thiosuccinimides,
Geniposide or glutaraldehyde.
4. preparation method according to claim 3, it is characterised in that the usage amount of the crosslinking agent and corneal stroma lens
Mass ratio be 1:1-1:15.
5. preparation method according to claim 4, it is characterised in that the usage amount of the crosslinking agent and corneal stroma lens
Mass ratio be 1:5-1:10.
6. preparation method according to claim 4, it is characterised in that before cell cracking processing, in addition to disappear
Poison processing, the method for the sterilization includes：Be 0.01~0.1mg/ml penicillin containing concentration and concentration be 0.05~
The physiological saline immersion 1-5h of 0.5mg/ml streptomysins, is then rinsed with 0.9% physiological saline or phosphate buffer solution.
7. preparation method according to claim 6, it is characterised in that after the crosslinking Treatment, in addition at sterilizing
Reason, the method for the sterilizing includes：Irradiated with gamma-rays, irradiation dose is 20~30kGy.
8. preparation method according to claim 7, it is characterised in that the corneal stroma lens with cell are to pass through
What full femtosecond laser technology was obtained.
9. preparation method according to claim 8, it is characterised in that the preparation method includes：
Step (a), sterilization：The corneal stroma lens of the exact thickness with cell will be obtained by full femtosecond laser technology, used
The physiological saline immersion 1-5h that containing concentration be 0.01~0.1mg/ml penicillin and concentration is 0.05~0.5mg/ml streptomysins,
Then rinsed with 0.9% physiological saline or phosphate buffer solution；
Step (b), cell cracking：It is molten with 0.9% physiological saline or phosphate-buffered containing 0.1%-3%TritonX-100
Immersion steeps 12-48h, then rinses 48-96h with 0.9% physiological saline；
Step (c), crosslinking：Soaked with 0.9% physiological saline or phosphate buffer solution containing crosslinking agent, Ran Houyong
0.9% physiological saline or phosphate buffer solution rinsing 24-96h, the crosslinking agent is 1- (3- dimethylamino-propyls) -3- second
Base carbodiimide, or N- hydroxy thiosuccinimides or glutaraldehyde or Geniposide；The usage amount of the crosslinking agent with it is described
The mass ratio that the corneal stroma lens of processing are cracked by cell is 1:1-1:15；
Step (d), sterilizing：Irradiated with gamma-rays, irradiation dose is 20~30kGy.
10. remove cell corneal stroma lens according to what the preparation method described in claim any one of 1-9 was obtained.
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| PCT/CN2018/082470 WO2018219045A1 (en) | 2017-05-31 | 2018-04-10 | Decellularized corneal stromal lens and preparation method therefor |
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| WO2018219045A1 (en) * | 2017-05-31 | 2018-12-06 | 广州新诚生物科技有限公司 | Decellularized corneal stromal lens and preparation method therefor |
| CN109568663A (en) * | 2018-11-27 | 2019-04-05 | 中山大学中山眼科中心 | The preparation method and cornea scaffold of cornea scaffold |
| CN111803437A (en) * | 2020-06-03 | 2020-10-23 | 广州新诚生物科技有限公司 | Medicine slow-release animal corneal stroma lens and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN115103695A (en) * | 2019-12-27 | 2022-09-23 | 盖鲍厄-克洛波泰克佩坦特弗瓦尔屯格斯-尤格公司(有限责任) | Stabilization of collagen scaffolds |
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