CN100579588C - Intravascular adult stem cell rack for preventing and treating angiostenosis and preparation method thereof - Google Patents
Intravascular adult stem cell rack for preventing and treating angiostenosis and preparation method thereof Download PDFInfo
- Publication number
- CN100579588C CN100579588C CN 200610095351 CN200610095351A CN100579588C CN 100579588 C CN100579588 C CN 100579588C CN 200610095351 CN200610095351 CN 200610095351 CN 200610095351 A CN200610095351 A CN 200610095351A CN 100579588 C CN100579588 C CN 100579588C
- Authority
- CN
- China
- Prior art keywords
- antibody
- cytokine
- solution
- protein
- stent
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Images
Abstract
The intravascular adult stem cell rack for preventing and treating angiostenosis includes one bare metal rack and one protein coating coated to the surface of the bare metal rack to promote the adhesion, proliferation and directional differentiation into endothelial cell of the adult stem cell. There may be also one polymer coating in between. The protein coating includes one antibody layer, one polypeptide/medicine layer and one cell factor layer. The present invention can form endothelium in the surface of the rack, decrease injury to endothelium, and prevent and treat angiostenosis.
Description
(1), technical field
The present invention relates to a kind of endovascular stent of cell envelope, particularly a kind of endovascular stent of adult stem cell bag quilt.
(2), background technology
Intravascular stent is as the important carrier of Minimally Invasive Surgery treatment angiostenosis, short-period clinical after it is implanted has obtained extensive approval, since the eighties development, become the routine treatment of running neck and neck as the percutaneous transluminal angio plasty (PTA) of technical foundation with medicine and other surgical operation therapy coronary heart disease.But regrettably, bare metal stent insert and fail fully " healing " vascular restenosis for a long time, but also cause appearance---in-stent restenosis (the In-stent restenosis of a new problem; ISR).According to clinical statistics, insert that the vascular restenosis incidence rate is 15-20% behind the support, and complication with diabetes, pathological changes is filled the air with the incidence rate of blood vessel smaller in-stent restenosis up to 30-50%.In order to overcome this limitation, strengthen the anti-restenosis ability of support, farthest prolong the curative effect time limit of support, people make many-sided improvement to support technology.For example, improve the materials used and the version of support; Antithrombotic class medicine is coated on support; Make rack surface have the radioactivity effect; Develop and no sheath support etc.As if improve in the effort at these, coating stent of medicine is becoming an important development direction of support technology, the newtype drug support is exactly the antithrombotic medicine of coating one deck on the common metal bare bracket, can make ISR be reduced to 10% once.But bracket for eluting medicament exists shortcoming and defect: (1) bracket for eluting medicament also just postpones the generation of restenosis, rather than suppresses restenosis; (2) being used for the polymer of drug loading can induce chronic inflammatory disease, injury repairing and untoward reaction such as endothelialization delay again; (3) limitation of medicine, the medicine that applies on support has at present also suppressed the hypertrophy of vascular endothelial cell in the outgrowth while of suppressing vascular smooth muscle cell, causes the formation of thrombosis and the generation that support is inserted back middle and late stage restenosis.
At top problem, the someone proposes that endothelialization is carried out on the endovascular stent surface and modifies.Generally believe anticoagulation and even improve the optimal approach of vascular graft blood compatibility and should be in the plantation of the surface of biomaterial, cultivate endotheliocyte, i.e. endotheliocyte immobilization now.Yet, this imagination of endovascular stent endothelialization still fails to become conventional clinical treatment means so far, tracing it to its cause is the problem that can run into a lot of reality in concrete operations, be mainly the problem of the aspects such as transportation, storage of the biological function of adhesion, cell of source, the cell of seed cell and endothelialization support, wherein the most key is the problem in seed cell source.In the cytoskeleton research in the early stage, what seed cell adopted is mature endothelial cell.But, in the past in 30 years, still externally in the body attempt all not obtain to count on one's card in the method for artificial graft's blood vessel, support and politef blood vessel graft surface plantation endotheliocyte.Its reason mainly contains: (1) endotheliocyte has strong immunogenicity, thereby the application of xenogenesis or allosome endotheliocyte can not realize basically; (2) from the body endotheliocyte, mainly take from blood capillary in autogenous vein or the fatty tissue, source and number are limited, and are difficult to use in acute and subacute cardiovascular patient; (3) endothelial cell seeding can not effectively keep and survive behind balloon expandable in rack surface.Because there is above-mentioned defective in endotheliocyte as seed cell, therefore seeking new seed cell becomes urgent problem, and stem-cell research day by day deeply may provide a kind of new cell source for us.
The endothelial progenitor cells that derives from bone marrow is considered to support the integrity of blood vessel endothelium endothelialization.Jacob etc. obtain ample evidence and confirm that the quick endotheliosis in the support is partly relevant with the endothelium of damage, and this is to reduce (the Jacob George that causes by the quantity of EPCs or its function, Itzhak Herz, Emil Goldstein, et al.Number and Adhesiveproperties of circulating endothelial progenitor cells inpatients with in-stent restenosis.Arterioscler Thromb VascBiol, 2003,23 (12): the quantity and the adhesion characteristics of 57-60. in-stent restenosis patient body internal recycle endothelial progenitor cells).Then, Shi etc. find that at first the subgroup of bone marrow CD34 (+) hemopoietic forebody cell has participated in endothelialization (the Shi Q of Dacron support of the sealing of canine model intravascular stent treatment, Rafii S, Wu MH, et al.Evidence for circulatingbone marrow-derived endothelial cells.Blood.1998; 92:362-367. the evidence of the circulation endothelium cell of derived from bone marrow).Subsequently, Bhattacharya etc. discover at polyethylene terephthalate (polyethylene terephthalate, PET) autologous transplanting bone marrow can quicken the endothelialization of support on the support, confirmed it is that CD34+ cell in the bone marrow has strengthened intravascular stent endothelialization degree (Vishwanath Bhattacharya simultaneously, Peter A.McSweeney, Qun Shi, endothelialization and blood capillary that et al.Enhanced endothelializationand microvessel formation in polyester grafts seeded with CD34+bone marrow cells.Blood.2000,95:581-585. strengthen plantation CD34+ medullary cell polyester graft form).2005, after utilization angiography technology such as Jiro are observed in the support of catching EPCs is implanted the coronary artery patient body, observe whether rapidly endothelialization of stainless steel stent, thereby the formation of thrombosis and minimizing restenosis rate in the inhibition support, proof EPCs crown support is safe and feasible (Jiro Aoki to treating newborn coronary artery disease, Patrick W.Serruys, Heleen van Beusekom, etal.Endothelial Progenitor Cell Capture by Stents Coated WithAntibody Against CD34.Journal of the American College ofCardiology, 2005,45 (10): 1574-1579. wraps by CD34 antibody support capturing endothelial ancestral cell.)。
Above-mentioned result of study shows, although quickened the formation that the support endothelium covered and reduced thrombosis in the support, but the effect of anti-restenosis does not reach initial imagination, the reason that may exist comprises: (1) is also thoroughly not clear and definite to the differentiation characteristic of EPC, the EPC of CD34+ may not have best endothelium differentiation potential, even may be divided into smooth muscle cell and increase the weight of restenosis.(2) specificity of CD34+ antibody capture is still waited to improve, and also has sophisticated endotheliocyte or even inflammatory cell except that EPCs.Although still have many problem demanding prompt solutions, but the adult stem cell support combines stem cell and bionic technology, generation from unique angle control restenosis, also further improve on processing technology if its mechanism is done further investigation, this support will be an impressive progress of coronary heart disease control behind drug stent.
(3), summary of the invention
One of purpose of the present invention just provides the adult stem cell endovascular stent of restenosis in a kind of treatment of vascular, it can make the rapid endothelialization of rack surface, minimizing is to the damage of endothelium, prevent and treat percutaneous transluminal coronary angioplasty (PTCA) back vascular restenosis or endovascular stent restenosis, have significant effect.
Two of purpose of the present invention just provides the adult stem cell endovascular stent of restenosis in a kind of treatment of vascular, and it is on the basis of above-mentioned purpose one, and protein coat is coated on the outer surface of bare metal stent more easily uniformly and stably.
Three of purpose of the present invention is exactly the support in the relative above-mentioned purpose one, and the preparation method of the adult stem cell endovascular stent of restenosis in a kind of treatment of vascular is provided.
Four of purpose of the present invention is exactly the support in the relative above-mentioned purpose two, and the preparation method of the adult stem cell endovascular stent of restenosis in a kind of treatment of vascular is provided.
One of purpose of the present invention is to realize by such technical scheme, it includes bare metal stent, it is characterized in that: at the outer surface of bare metal stent one deck is set and facilitates the soma cell adhesion, propagation and directed differentiation are the protein coat of endotheliocyte, protein coat includes antibody layer, layer polypeptide and cytokine layer, it is according to the following outer surface that is set in sequence in bare metal stent: with antibody layer, layer polypeptide and cytokine layer optionally carry out assembled arrangement, with this unit of primordial, a plurality of such elementary cells are evenly distributed on the outer surface of bare metal stent.
With support of the present invention, directly implant patient vessel's lesion, perhaps In vitro culture patient's adult stem cell, because the outer surface of bare metal stent is provided with protein coat, therefore, can the specific adhesion adult stem cell, promote propagation of stem cell and the directed differentiation of promotion stem cell, thereby make the rapid endothelialization of rack surface, reduce damage, prevent and treat percutaneous transluminal coronary angioplasty (PTCA) back vascular restenosis or endovascular stent restenosis endothelium.Described protein coat includes a kind of or several in antibody, cytokine, polypeptide, statins and the estrogen or all.Wherein antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes, and its effect is for the specific adhesion adult stem cell; Cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes, promotes the differentiation of stem cell; Polypeptide promptly is ring-type Arg-Gly-Asp polypeptide, and is identical with statins and estrogen action, promotes the propagation of stem cell.
Two of purpose of the present invention is to realize by such technical scheme, on the basis of above-mentioned purpose one, is provided with the polymer coating of being convenient to apply protein coat between the outer surface of bare metal stent and protein coat.
Because the affinity of protein and metal surface is poor, therefore, causes protein coat inhomogeneous easily, poor stability as easy as rolling off a logly comes off from the metal surface.In order to address this problem, make full use of polymer not only had with the metal surface associativity by force but also have characteristics high with proteinic affinity, outer surface at bare metal stent applies one layer of polymeric earlier, back coating protein coat, not only can make like this that protein coat is evenly distributed, stability is strong, and guaranteed the reliability that combines between protein coat and the metal, thereby prevented proteinic coming off.Described polymer coating includes polymeric material and solvent, wherein, polymeric material is by any one or wherein two kinds or wherein multiple composition the in polylactic acid, politef, gelatin, high density polyethylene (HDPE), polyurethane and the polyethylene terephthalate; Solvent is acetone or oxolane.Effect is for protein coat more easily being applied, keeping the uniformity and the stability of protein coat.
The preparation method of the support of above-mentioned purpose one is that to facilitate soma cell adhesion, propagation and directed differentiation be the protein solution of endotheliocyte in preparation earlier; Then, the bare metal stent after sterile-processed is soaked in the protein solution, perhaps the mode of protein solution with spraying is coated on the outer surface of bare metal stent, to make the endovascular stent of band protein coat; Described protein solution is prepared as follows: protein solution includes antibody and cytokine, with antibody and cytokine respectively with after the PBS buffer mixes, remix is made solution together, perhaps with antibody and cytokine together with after the PBS buffer mixes, make solution, be protein solution; Wherein, described antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes; Described cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes; The ultimate density of antibody and cytokine is than (3～6): (2～3).
Protein coat includes antibody layer, layer polypeptide and cytokine layer, it is according to the following outer surface that is set in sequence in bare metal stent: antibody layer, layer polypeptide and cytokine layer are optionally carried out assembled arrangement, with this unit of primordial, a plurality of such elementary cells are evenly distributed on the outer surface of bare metal stent, help adhesion, propagation and the directed differentiation of adult stem cell.The implementing of this method is beneficial to outer surface at bare metal stent and one deck is set to facilitate soma cell adhesion, propagation and directed differentiation be the protein coat of endotheliocyte, increases the specificity of stem cell and guaranteed that directed differentiation is the ability and the endothelium differentiation potential of endotheliocyte.
The preparation method of the support of above-mentioned purpose two is carried out as follows:
(1), preparation is convenient to apply the polymer solution of protein coat; Described polymer solution includes polymeric material and solvent, wherein, polymeric material is by any one or wherein two kinds or wherein multiple composition the in polylactic acid, politef, gelatin, high density polyethylene (HDPE), polyurethane and the polyethylene terephthalate; Solvent is acetone or oxolane; Polymer solution is preparation like this: 20 ℃-40 ℃ of room temperatures, utilize the magnetic agitation instrument to stir, and at least 15 minutes time, fully to mix, preparation concentration is the polymer material solution of 1mg/ml-5mg/ml;
(2), the bare metal stent after sterile-processed is soaked in the polymer solution, perhaps the mode of polymer solution with spraying is coated on the outer surface of bare metal stent, to make polymer coating;
(3), to facilitate soma cell adhesion, propagation and directed differentiation be the protein solution of endotheliocyte in preparation; Described protein solution is prepared as follows: protein solution includes antibody and cytokine, with antibody and cytokine respectively with after the PBS buffer mixes, remix is made solution together, perhaps with antibody and cytokine together with after the PBS buffer mixes, make solution, be protein solution; Wherein, described antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes; Described cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes; The ultimate density of antibody and cytokine is than (3～6): (2～3);
(4), the support that obtains in the step (2) is soaked in the protein solution, perhaps the mode of protein solution with spraying is coated on the outer surface of polymer coating, making protein coat, thereby makes endovascular stent.
Owing to adopted technique scheme, the present invention to have following advantage:
1. cell derives from the patient from body, can not produce the allosome immunological rejection, and safety is guaranteed.
2. the rapid endothelialization of rack surface reduces the damage to endothelium.
3. avoid the inductive inflammatory reaction of polymer, effectively reduce restenosis in the blood vessel.
4. the selection of specific antibody and inducible factor increases the specificity of stem cell and has guaranteed that directed differentiation is the ability and the endothelium differentiation potential of endotheliocyte.
Domestic present clinical practice drug-eluting stent mainly is external endovascular stent product, makes it hold at high price.And, also do not have formal commodity listing both at home and abroad in the cytoskeleton field.Therefore,, be expected to shorten China and developed country gap in cardiovascular intervention apparatus field by development and application of the present invention, will for homemade support move towards the international market and at home market strive for suitable share.
(4), description of drawings
Description of drawings of the present invention is as follows:
Fig. 1 is a schematic appearance of the present invention;
Fig. 2 is the cutaway view of first kind of embodiment of A-A among Fig. 1;
Fig. 3 is the cutaway view of second kind of embodiment of A-A among Fig. 1;
Fig. 4 is the cutaway view of the third embodiment of A-A among Fig. 1;
Fig. 5 is the cutaway view of first kind of embodiment of B-B among Fig. 4;
Fig. 6 is the cutaway view of second kind of embodiment of B-B among Fig. 4;
Among the figure, 1. bare metal stent; 2. protein coat; 3. polymer coating; 4. antibody layer; 5. layer polypeptide; 6. cytokine layer.
(5), the specific embodiment
The invention will be further described below in conjunction with drawings and Examples:
As Fig. 1,2,3,4,5, shown in 6, the present invention includes bare metal stent 1, at the outer surface of bare metal stent 1 one deck is set and facilitates the soma cell adhesion, propagation and directed differentiation are the protein coat 2 of endotheliocyte, protein coat 2 includes antibody layer 4, layer polypeptide 5 and cytokine layer 6, it is according to the following outer surface that is set in sequence in bare metal stent 1: with antibody layer 4, layer polypeptide 5 and cytokine layer 6 optionally carry out assembled arrangement, with this unit of primordial, a plurality of such elementary cells are evenly distributed on the outer surface of bare metal stent 1.
With support of the present invention, directly implant patient vessel's lesion, perhaps In vitro culture patient's adult stem cell, because the outer surface of bare metal stent is provided with protein coat, therefore, can the specific adhesion adult stem cell, promote propagation of stem cell and the directed differentiation of promotion stem cell, thereby make the rapid endothelialization of rack surface, reduce damage, prevent and treat percutaneous transluminal coronary angioplasty (PTCA) back vascular restenosis or endovascular stent restenosis endothelium.Described protein coat includes a kind of or several in antibody, cytokine, polypeptide, statins and the estrogen or all.Wherein antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes, and its effect is for the specific adhesion adult stem cell; Cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes, promotes the differentiation of stem cell; Polypeptide promptly is ring-type Arg-Gly-Asp polypeptide, and is identical with statins and estrogen action, promotes the propagation of stem cell.
Shown in Fig. 1,4, between the outer surface of bare metal stent 1 and protein coat 2, can be provided with the polymer coating 3 of being convenient to apply protein coat.
Because the affinity of protein and metal surface is poor, therefore, causes protein coat inhomogeneous easily, poor stability as easy as rolling off a logly comes off from the metal surface.In order to address this problem, make full use of polymer not only had with the metal surface associativity by force but also have characteristics high with proteinic affinity, outer surface at bare metal stent applies one layer of polymeric earlier, back coating protein coat, not only can make like this that protein coat is evenly distributed, stability is strong, and guaranteed the reliability that combines between protein coat and the metal, thereby prevented proteinic coming off.Described polymer coating includes polymeric material and solvent, wherein, polymeric material is by any one or wherein two kinds or wherein multiple composition the in polylactic acid, politef, gelatin, high density polyethylene (HDPE), polyurethane and the polyethylene terephthalate; Solvent is acetone or oxolane.Effect is for protein coat more easily being applied, keeping the uniformity and the stability of protein coat.
As shown in Figure 2, after protein coat can be all substances mix homogeneously of forming, the whole layer that is evenly distributed.
As shown in Figure 3, protein coat also can be to be made of antibody layer 4, layer polypeptide 5 and cytokine layer 6, they carry out assembled arrangement with form longitudinally, with this unit of primordial, a plurality of such elementary cells are evenly distributed on the outer surface of bare metal stent 1.
In conjunction with Fig. 4,5 as can be known, after protein coat can be all substances mix homogeneously of forming, the whole layer that is evenly distributed.
In conjunction with Fig. 4,6 as can be known, protein coat also can be to be made of antibody layer 4, layer polypeptide 5 and cytokine layer 6, they carry out assembled arrangement with horizontal form, with this unit of primordial, a plurality of such elementary cells are evenly distributed on the outer surface of bare metal stent 1.
The thickness of polymer coating 3 is 1 μ m～500 μ m; Optimum thickness is 10 μ m～100 μ m.The thickness of protein coat 2 is 1 μ m～500 μ m; Optimum thickness is 10 μ m～100 μ m.
The preparation method of above-mentioned support can be that to facilitate soma cell adhesion, propagation and directed differentiation be the protein solution of endotheliocyte in preparation earlier; Then, the bare metal stent after sterile-processed is soaked in the protein solution first, perhaps the mode of protein solution with spraying is coated on the outer surface of bare metal stent, to make the endovascular stent of band protein coat; Described protein solution is prepared as follows: protein solution includes antibody and cytokine, with antibody and cytokine respectively with after the PBS buffer mixes, remix is made solution together, perhaps with antibody and cytokine together with after the PBS buffer mixes, make solution, be protein solution; Wherein, described antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes; Described cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes; The ultimate density of antibody and cytokine is than (3～6): (2～3).
Protein coat includes antibody layer, layer polypeptide and cytokine layer, it is according to the following outer surface that is set in sequence in bare metal stent: antibody layer, layer polypeptide and cytokine layer are optionally carried out assembled arrangement, with this unit of primordial, a plurality of such elementary cells are evenly distributed on the outer surface of bare metal stent, help adhesion, propagation and the directed differentiation of adult stem cell.The implementing of this method is beneficial to outer surface at bare metal stent and one deck is set to facilitate soma cell adhesion, propagation and directed differentiation be the protein coat of endotheliocyte, increases the specificity of stem cell and guaranteed that directed differentiation is the ability and the endothelium differentiation potential of endotheliocyte.
The preparation method of above-mentioned support also can be carried out as follows:
(1), preparation is convenient to apply the polymer solution of protein coat; Described polymer solution includes polymeric material and solvent, wherein, polymeric material is by any one or wherein two kinds or wherein multiple composition the in polylactic acid, politef, gelatin, high density polyethylene (HDPE), polyurethane and the polyethylene terephthalate; Solvent is acetone or oxolane; Polymer solution is preparation like this: 20 ℃-40 ℃ of room temperatures, utilize the magnetic agitation instrument to stir, and at least 15 minutes time, fully to mix, preparation concentration is the polymer material solution of 1mg/ml-5mg/ml;
(2), the bare metal stent after sterile-processed is soaked in the polymer solution, perhaps the mode of polymer solution with spraying is coated on the outer surface of bare metal stent, to make polymer coating;
(3), to facilitate soma cell adhesion, propagation and directed differentiation be the protein solution of endotheliocyte in preparation; Described protein solution is prepared as follows: protein solution includes antibody and cytokine, with antibody and cytokine respectively with after the PBS buffer mixes, remix is made solution together, perhaps with antibody and cytokine together with after the PBS buffer mixes, make solution, be protein solution; Wherein, described antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes; Described cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes; The ultimate density of antibody and cytokine is than (3～6): (2～3);
(4), the support that obtains in the step (2) is soaked in the protein solution, perhaps the mode of protein solution with spraying is coated on the outer surface of polymer coating, making protein coat, thereby makes endovascular stent.
In the said method, protein solution also includes ring-type Arg-Gly-Asp polypeptide; The ultimate density of antibody, ring-type Arg-Gly-Asp polypeptide, cytokine is than (3～6): (1～3): (2～3).
In the said method, protein coat also includes statins or/and estrogen; Antibody, statins or/and the ultimate density of estrogen, cytokine than (3～6): (1～3): (2～3).
In the said method, protein coat also includes ring-type Arg-Gly-Asp polypeptide; Antibody, ring-type Arg-Gly-Asp polypeptide, statins or/and the ultimate density of estrogen, cytokine than (3～6): (1～3): (1～3): (2～3).
(1) the selection of material 316L type stainless steel stent, ultrasonic cleaning 5 minutes in ethanol, acetone, distilled water successively, in vacuum drying oven dry 60 minutes then, baking temperature was 45 degree, utilizes ultra violet lamp to carry out disinfection in 30 minutes at last;
(2) preparation coating material, matrix material is polylactic acid and polyvinyl fluoride (1: 1), and solvent is an acetone, and polymer concentration in its solvent is 2 mg/ml, under 25 degree room temperatures, utilizes the magnetic agitation instrument to stir mix homogeneously 30 minutes;
(3) ultrasonic atomizatio spraying preparation polymer coating, the matrix material solution that makes is added the solution bottle of spray equipment, form uniform medication coat by spray equipment at rack surface, spray time is 1 minute, operate in sterilizing room, final coating thickness is 20 μ m;
(4) will be coated with and be placed in the vacuum drying oven, 24 hours drying times, baking temperature 40 degree;
(5) CD133 antibody, ring-type Arg-Gly-Asp polypeptide and the VEGF factor three class drug solutions are mixed (2: 1: 1), the support that step (4) prepares is put into this mixed solution (3-5ml), soak 10min, oven dry, same operation three times, vacuum drying forms protein layer on the polymeric layer surface, and thickness is 20 μ m;
(6) cryopreservation, 40 degrees below zero, standby.
(1) the selection of material NiTi alloy bracket, ultrasonic cleaning 5 minutes in ethanol, acetone, distilled water successively, in vacuum drying oven dry 60 minutes then, baking temperature was 45 degree, utilizes ultra violet lamp to carry out disinfection in 30 minutes at last;
(2) preparation coating material, matrix material is high density polyethylene (HDPE) and gelatin (1: 1), and solvent is an acetone, and polymer concentration in its solvent is 3 mg/ml, under 30 degree temperature, utilizes the magnetic agitation instrument to stir mix homogeneously 30 minutes;
(3) bare bracket is put into the polymer coating solution that step (2) prepares, soaked 10min, oven dry, three times so repeatedly, form polymer coating, thickness is 30 μ m;
(4) CD34 antibody, statins and the TGF factor three class drug solutions are mixed (2: 1: 1), the support that step (4) prepares is put into this mixed solution (3-5ml), soaks 10min, oven dry, same operation three times, vacuum drying, last coating layer thickness are 30 μ m;
(5) cryopreservation, subzero 30 degree, standby.
(1) select stainless steel stent for use, ultrasonic cleaning 5 minutes in ethanol, acetone, distilled water successively, in vacuum drying oven dry 60 minutes then, baking temperature was 45 degree, utilizes ultra violet lamp to carry out disinfection in 30 minutes at last;
(2) preparation coating material, matrix material is polylactic acid and polyurethane (2: 1), and solvent is an acetone, and polymer concentration in its solvent is 4 mg/ml, under 35 degree temperature, utilizes the magnetic agitation instrument to stir mix homogeneously 60 minutes;
(3) ultrasonic atomizatio spraying preparation medication coat adds the solution bottle of spray equipment with the matrix material solution that makes, and by spray equipment, forms uniform medication coat at rack surface, and coating layer thickness is 20 μ m;
(4) prepare A, B and C three class mixed solutions respectively, category-A solution C D133 antibody wherein: CD34 antibody: CD14 antibody is 1: 1: 1, category-B solution ring-type Arg-Gly-Asp polypeptide: statins: estrogen is 1: 1: 1, and the C class solution VEGF factor: the bFGF factor: the TGF factor is 1: 1: 1.By spraying technology, on the support that step (2) prepares, be coated with the band of three class mixed solutions respectively, every bandwidth 0.1-0.5mm, thickness are 25 μ m, last vacuum drying;
(5) separation and purification endothelial progenitor cells from patient's peripheral blood, amplification in vitro, cell density reaches 10
6, on the support of the sterilization that inoculating cell prepares to step (4), cultivate 3-4d, cell covers rack surface fully.
(6) with the stem cell support for preparing, put into storage liquid (culture fluid: serum: glycerol=7: 2: 1), place liquid nitrogen container to preserve then, standby.
Claims (9)
1. the adult stem cell endovascular stent of restenosis in the treatment of vascular, include bare metal stent (1), it is characterized in that: at the outer surface of bare metal stent (1) one deck is set and facilitates the soma cell adhesion, propagation and directed differentiation are the protein coat (2) of endotheliocyte, protein coat (2) includes antibody layer (4), layer polypeptide (5) and cytokine layer (6), it is according to the following outer surface that is set in sequence in bare metal stent (1): with antibody layer (4), layer polypeptide (5) and cytokine layer (6) optionally carry out assembled arrangement, with this unit of primordial, a plurality of such elementary cells are evenly distributed on the outer surface of bare metal stent (1).
2. the adult stem cell endovascular stent of restenosis in the treatment of vascular as claimed in claim 1 is characterized in that: be provided with the polymer coating (3) of being convenient to apply protein coat between the outer surface of bare metal stent (1) and protein coat (2).
3. the adult stem cell endovascular stent of restenosis in the treatment of vascular as claimed in claim 2, it is characterized in that: the thickness of polymer coating (3) is 1 μ m～500 μ m.
4. as the adult stem cell endovascular stent of restenosis in claim 1, the 2 or 3 described treatment of vascular, it is characterized in that: the thickness of protein coat (2) is 1 μ m～500 μ m.
5. the preparation method of the adult stem cell endovascular stent of restenosis in the treatment of vascular as claimed in claim 1, it is that to facilitate soma cell adhesion, propagation and directed differentiation be the protein solution of endotheliocyte in preparation earlier; Then, the bare metal stent after sterile-processed is soaked in the protein solution, perhaps the mode of protein solution with spraying is coated on the outer surface of bare metal stent, to make the endovascular stent of band protein coat; Described protein solution is prepared as follows: protein solution includes antibody and cytokine, with antibody and cytokine respectively with after the PBS buffer mixes, remix is made solution together, perhaps with antibody and cytokine together with after the PBS buffer mixes, make solution, be protein solution; Wherein, described antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes; Described cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes; The ultimate density of antibody and cytokine is than (3～6): (2～3).
6. the preparation method of the adult stem cell endovascular stent of restenosis in the treatment of vascular as claimed in claim 2, it carries out as follows:
(1), preparation is convenient to apply the polymer solution of protein coat; Described polymer solution includes polymeric material and solvent, wherein, polymeric material is by any one or wherein two kinds or wherein multiple composition the in polylactic acid, politef, gelatin, high density polyethylene (HDPE), polyurethane and the polyethylene terephthalate; Solvent is acetone or oxolane; Polymer solution is preparation like this: 20 ℃-40 ℃ of room temperatures, utilize the magnetic agitation instrument to stir, and at least 15 minutes time, fully to mix, preparation concentration is the polymer material solution of 1mg/ml-5mg/ml;
(2), the bare metal stent after sterile-processed is soaked in the polymer solution, perhaps the mode of polymer solution with spraying is coated on the outer surface of bare metal stent, to make polymer coating;
(3), to facilitate soma cell adhesion, propagation and directed differentiation be the protein solution of endotheliocyte in preparation; Described protein solution is prepared as follows: protein solution includes antibody and cytokine, with antibody and cytokine respectively with after the PBS buffer mixes, remix is made solution together, perhaps with antibody and cytokine together with after the PBS buffer mixes, make solution, be protein solution; Wherein, described antibody is by one in CD133 antibody, CD34 antibody and the CD14 antibody or two or all constitutes; Described cytokine is by one in the VEGF factor, the bFGF factor and the TGF factor or two or all constitutes; The ultimate density of antibody and cytokine is than (3～6): (2～3);
(4), the support that obtains in the step (2) is soaked in the protein solution, perhaps the mode of protein solution with spraying is coated on the outer surface of polymer coating, making protein coat, thereby makes endovascular stent.
7. as the preparation method of the adult stem cell endovascular stent of restenosis in claim 5 or the 6 described treatment of vascular, it is characterized in that: protein solution also includes ring-type Arg-Gly-Asp polypeptide; The ultimate density of antibody, ring-type Arg-Gly-Asp polypeptide, cytokine is than (3～6): (1～3): (2～3).
8. as the preparation method of the adult stem cell endovascular stent of restenosis in claim 5 or the 6 described treatment of vascular, it is characterized in that: protein coat also includes statins or/and estrogen; Antibody, statins or/and the ultimate density of estrogen, cytokine than (3～6): (1～3): (2～3).
9. the preparation method of the adult stem cell endovascular stent of restenosis in the treatment of vascular as claimed in claim 8, it is characterized in that: protein coat also includes ring-type Arg-Gly-Asp polypeptide; Antibody, ring-type Arg-Gly-Asp polypeptide, statins or/and the ultimate density of estrogen, cytokine than (3～6): (1～3): (1～3): (2～3).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200610095351 CN100579588C (en) | 2006-12-26 | 2006-12-26 | Intravascular adult stem cell rack for preventing and treating angiostenosis and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200610095351 CN100579588C (en) | 2006-12-26 | 2006-12-26 | Intravascular adult stem cell rack for preventing and treating angiostenosis and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101020087A CN101020087A (en) | 2007-08-22 |
| CN100579588C true CN100579588C (en) | 2010-01-13 |
Family
ID=38708013
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 200610095351 Expired - Fee Related CN100579588C (en) | 2006-12-26 | 2006-12-26 | Intravascular adult stem cell rack for preventing and treating angiostenosis and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN100579588C (en) |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20110046747A1 (en) * | 2009-02-19 | 2011-02-24 | Kelvin Wai Kwok Yeung | Antibacterial surface and method of fabrication |
| CN102648988A (en) * | 2011-02-28 | 2012-08-29 | 刘斌 | Preparation method of stent coated with CD34 antibody |
| CN105327399B (en) * | 2015-10-27 | 2017-12-22 | 苏州大学 | A kind of construction method of artificial blood vessel |
| CN114904062B (en) * | 2022-04-29 | 2023-04-28 | 中国科学院金属研究所 | Vascular stent with selective biological functionalization and preparation method thereof |
-
2006
- 2006-12-26 CN CN 200610095351 patent/CN100579588C/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| CN101020087A (en) | 2007-08-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Pacelli et al. | Strategies to develop endogenous stem cell-recruiting bioactive materials for tissue repair and regeneration | |
| Jaganathan et al. | Biomaterials in cardiovascular research: applications and clinical implications | |
| US9289533B2 (en) | Collagen scaffold modified by covalent grafting of adhesion molecules, associated methods and use thereof for cardiovascular and thoracic cell therapy and contractile tissue engineering | |
| CN100455275C (en) | Medical device with coating that promotes endothelial cell adherence and differentiation | |
| Lam et al. | Biomaterial applications in cardiovascular tissue repair and regeneration | |
| Jabbarzadeh et al. | Induction of angiogenesis in tissue-engineered scaffolds designed for bone repair: a combined gene therapy–cell transplantation approach | |
| Ozawa et al. | Optimal biomaterial for creation of autologous cardiac grafts | |
| Kuss et al. | Short-term hypoxic preconditioning promotes prevascularization in 3D bioprinted bone constructs with stromal vascular fraction derived cells | |
| Isenberg et al. | Small-diameter artificial arteries engineered in vitro | |
| Miyagi et al. | Biodegradable collagen patch with covalently immobilized VEGF for myocardial repair | |
| Murphy et al. | Bone regeneration via a mineral substrate and induced angiogenesis | |
| CN104013996B (en) | The drug eluting implantable medical device of progenitor endothelial cell capturing | |
| Schultheiss et al. | Biological vascularized matrix for bladder tissue engineering: matrix preparation, reseeding technique and short-term implantation in a porcine model | |
| CN101391115B (en) | Preparation method of biological activity protein or polypeptides coating biology bracket | |
| Dahan et al. | Dynamic autologous reendothelialization of small-caliber arterial extracellular matrix: a preclinical large animal study | |
| Proulx et al. | Fibrin microthreads support mesenchymal stem cell growth while maintaining differentiation potential | |
| Tan et al. | Tissue-engineered trachea: History, problems and the future | |
| Sun et al. | Composite scaffolds of mineralized natural extracellular matrix on true bone ceramic induce bone regeneration through Smad1/5/8 and ERK1/2 pathways | |
| CN100579588C (en) | Intravascular adult stem cell rack for preventing and treating angiostenosis and preparation method thereof | |
| Ghorbel et al. | Reconstruction of the pulmonary artery by a novel biodegradable conduit engineered with perinatal stem cell-derived vascular smooth muscle cells enables physiological vascular growth in a large animal model of congenital heart disease | |
| Di Franco et al. | Biomaterials and heart recovery: cardiac repair, regeneration and healing in the MCS era: a state of the “heart” | |
| Arrigoni et al. | Rotating versus perfusion bioreactor for the culture of engineered vascular constructs based on hyaluronic acid | |
| CN101589971A (en) | Third generation PCI therapeutic saccule support system, preparation method and application | |
| Kong et al. | The use of heparin, bFGF, and VEGF 145 grafted acellular vascular scaffold in small diameter vascular graft | |
| Hui et al. | Preparation and in vivo evaluation of surface heparinized small diameter tissue engineered vascular scaffolds of poly (ε-caprolactone) embedded with collagen suture |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C17 | Cessation of patent right | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20100113 Termination date: 20121226 |