Artificial blood vessels and methods for manufacturing the same

Abstract

To provide a small-diameter artificial blood vessel that exhibits adequate antithrombotic efficacy and is suitable for practical use, and a method for producing the same.SOLUTION: An artificial blood vessel 101 includes a tubular structure 100 composed of nanofibers, and a hydrophilic polymer coating that is formed on the nanofiber surfaces, constituting at least the inner peripheral surface of the tubular structure 100. The artificial blood vessel has an inner diameter of 1 mm or more and 5 mm or less.SELECTED DRAWING: Figure 7

Description

Technical Field

[0001] The present disclosure relates to an artificial blood vessel and a method for manufacturing the same.

Background Art

[0002] In atherosclerotic diseases such as ischemic heart disease and lower limb obstructive arteriosclerosis, in severe cases, surgical revascularization such as coronary artery bypass surgery or lower limb artery bypass surgery is required. For this treatment, a substitute blood vessel with a diameter of less than 5 mm is required to bypass the occluded blood vessel. Currently, since an artificial blood vessel with a diameter of less than 5 mm does not even have short-term patency, autologous vascular grafts collected from the patient himself / herself are used for the treatment. However, there are problems such that the treatment itself cannot be performed when the length of the autologous vascular graft is insufficient or the properties are inappropriate for the treatment, and the development of a stockable small-diameter artificial blood vessel that can withstand practical use is desired.

[0003] What is expected of an ideal substitute blood vessel is the ability to integrate with the patient's tissue and behave like an autologous blood vessel. In order to achieve this with an artificial blood vessel, it is necessary for autologous cells to engraft and regenerate into a tissue-like state after transplantation, and biodegradable nanofibers on which vascular endothelial cells and smooth muscle cells that form blood vessels engraft have attracted attention as materials for artificial blood vessels (Non-Patent Documents 1 to 3).

[0004] When biodegradable nanofibers are used as a material for an artificial blood vessel, when a layer (intima) of vascular endothelial cells and smooth muscle cells that exhibit antithrombotic properties regenerates, as the biodegradable nanofibers decompose, the artificial blood vessel will ultimately be replaced by a complete autologous blood vessel, and long-term patency can be expected. However, since biodegradable nanofibers have high hydrophobicity, they easily adsorb platelets and cause thrombosis. In particular, the smaller the diameter of the artificial blood vessel, the easier thrombosis occurs, and it becomes difficult to have short-term patency until the intima is regenerated. Therefore, in order to obtain a small-diameter artificial blood vessel that can withstand practical use, it is essential to impart antithrombotic properties to the artificial blood vessel.

[0005] Attempts have been made to use hydrophilic polymers as materials for artificial blood vessels in order to impart antithrombotic properties to them (Patent Documents 1-3). However, hydrophilic polymers dissolve when they come into contact with water, so they cannot be used as is as materials for artificial blood vessels. Therefore, the techniques described in Patent Documents 1-3 attempt to impart hydrophilicity to artificial blood vessels while preventing the elution of the hydrophilic polymer by methods such as graft polymerization of a hydrophilic polymer onto an artificial blood vessel made of a hydrophobic polymer (Patent Document 1), blending it into the material beforehand (Patent Document 2), or hydrogelizing and adsorbing it (Patent Document 3). [Prior art documents] [Patent Documents]

[0006] [Patent Document 1] Japanese Patent Publication No. 60-227763 [Patent Document 2] Japanese Patent Publication No. 11-226113 [Patent Document 3] Japanese Patent Publication No. 04-038960 [Non-patent literature]

[0007] [Non-Patent Document 1] Biomaterials. 2012 33(1) 38-47 [Non-Patent Document 2] Pharmaceuticals (Basel). 2020 13(5) 101 [Non-Patent Document 3] Biomaterials. 1996 17(2) 115-124 [Overview of the project] [Problems that the invention aims to solve]

[0008] However, the technologies described in Patent Documents 1 to 3 have problems such as complicating the manufacturing process and increasing manufacturing costs, and although there are many research and development examples, they have not yet been put into practical use.

[0009] Therefore, the objective of this disclosure is to provide a small-diameter artificial blood vessel with sufficient antithrombotic properties that is suitable for practical use, and a method for manufacturing the same. [Means for solving the problem]

[0010] To solve the above problems, one embodiment of an artificial blood vessel disclosed herein comprises a tubular structure made of nanofibers and a coating of a hydrophilic polymer formed on the surface of the nanofibers constituting at least the inner circumferential surface of the tubular structure, characterized in that the inner diameter is 1 mm or more and 5 mm or less.

[0011] The ratio of the coating to the tubular structure is preferably 2.5% by mass or more and 10% by mass or less.

[0012] When distilled water is passed through the artificial blood vessel, the outflow ratio of the hydrophilic polymer is preferably 25% or less.

[0013] The water contact angle of the inner surface of the artificial blood vessel is preferably 120° or less.

[0014] Preferably, the coating is also formed on the surface of the nanofibers that constitute the outer surface of the tubular structure.

[0015] The water contact angle of the outer surface of the artificial blood vessel is preferably 120° or less.

[0016] In the platelet adsorption test, the platelet adsorption inhibition rate, expressed as a percentage of the number of platelet granules attached to the tubular structure without the coating minus the number of platelet granules attached to the artificial blood vessel, is preferably 30% or more.

[0017] For the artificial blood vessel with an inner diameter of 2 mm or more and 5 mm or less, the compression recovery obtained by compression testing is preferably 63% or more.

[0018] For the artificial blood vessel with an inner diameter of more than 1 mm and less than 2 mm, the compression recovery obtained by the compression test is preferably 41% or more.

[0019] The maximum load measured by the three-point bending test is preferably 0.0055 N or more.

[0020] The wall thickness of the artificial blood vessel is preferably 0.2 mm or more and 1.2 mm or less.

[0021] The tubular structure has a laminated structure formed by laminating a plurality of long nanofiber ribbons, and it is preferable that the end portions in the width direction of the nanofiber ribbons are arranged in a spiral shape on the inner peripheral surface side of the tubular structure.

[0022] The tubular structure has a laminated structure formed by laminating a plurality of nanofiber layers, and it is preferable that it further includes a suture disposed between two adjacent nanofiber layers.

[0023] The hydrophilic polymer is preferably at least one selected from the group consisting of polyvinyl alcohol, polyacrylic acid, sodium polyacrylate, polyacrylamide, polyvinylpyrrolidone, dextran, carboxymethyl cellulose, and polyethylene oxide.

[0024] The raw material resin of the nanofiber is preferably at least one selected from the group consisting of polycaprolactone, polylactic acid, polyglycolic acid, polylactic acid-glycolic acid, polyurethane, polyhydroxybutyric acid, and silk.

[0025] One embodiment of a method for manufacturing an artificial blood vessel disclosed herein is a method for manufacturing an artificial blood vessel comprising a tubular structure made of nanofibers and a coating of a hydrophilic polymer formed on the surface of the nanofibers constituting at least the inner circumferential surface of the tubular structure, wherein the inner diameter of the artificial blood vessel is 1 mm or more and 5 mm or less, and the method comprises a tubular structure formation step for forming the tubular structure and a coating formation step for forming the coating by passing a solution containing a hydrophilic polymer through the lumen of the tubular structure.

[0026] Preferably, the tubular structure formation step comprises a first step of producing a nanofiber sheet by electrospinning, a second step of cutting the nanofiber sheet to produce a long nanofiber ribbon, and a third step of winding the nanofiber ribbon around a core rod at a predetermined angle and removing the core rod to form the tubular structure.

[0027] Preferably, the third step includes step A of winding the first nanofiber ribbon around the core rod from one side to the other at a predetermined angle, and step B of winding the second nanofiber ribbon around the core rod around which the first nanofiber ribbon is wound, from the other side to the one side at a predetermined angle, and is a step of forming the tubular structure by repeating steps A and B.

[0028] The third step preferably includes a step of forming a nanofiber sheet on the surface of the core rod by electrospinning before winding the nanofiber ribbon onto the core rod.

[0029] The thickness of the nanofiber ribbon is preferably 5 μm or more and 100 μm or less.

[0030] The width of the nanofiber ribbon is preferably 50 mm or less.

[0031] The length of the nanofiber ribbon is preferably 30 cm or more.

[0032] One embodiment of the artificial blood vessel disclosed herein is an artificial blood vessel manufactured by any of the above-described manufacturing methods. [Effects of the Invention]

[0033] This disclosure provides a small-diameter artificial blood vessel with sufficient antithrombotic properties that is suitable for practical use, as well as a method for manufacturing the same. [Brief explanation of the drawing]

[0034] [Figure 1] A diagram showing an example of a tubular structure for an artificial blood vessel related to this disclosure. [Figure 2] Figure 1 shows a cross-sectional view along line AA and a modified example thereof. [Figure 3] Figure 1 shows a cross-sectional view along line BB and a modified example thereof. [Figure 4] A flowchart illustrating an example of a method for manufacturing an artificial blood vessel related to this disclosure. [Figure 5] This diagram illustrates the specific steps involved in each process shown in Figure 4. [Figure 6] Digital camera photographs of the tubular structures of Examples 1-4. [Figure 7] SEM image of the artificial blood vessel in Example 4. [Figure 8] SEM image of the artificial blood vessel in Example 2. [Figure 9] SEM images of the inner circumferential surface of the artificial blood vessels of Comparative Example 1 and Example 1. [Figure 10] Graphs showing the results of compression tests on artificial blood vessels from Examples 1 and 2 and Comparative Examples 1 and 3. [Figure 11] A diagram illustrating a method for calculating compression linearity, compression work, and compression recovery based on compression test results. [Figure 12] A diagram illustrating the sample preparation method for manufacturing examples 1 and 2 in contact angle measurement test 1. [Figure 13] Photographs of the contact angle measurement of samples from manufacturing examples 1 and 2 in contact angle measurement test 1. [Figure 14] SEM images of grafts from Example 4 and Comparative Example 2 in the platelet adsorption test. [Figure 15] A graph showing the results of a platelet adsorption test. [Figure 16] Fluorescence micrograph of tissue immunostaining of the radial section of the graft from Example 4 in Artificial Blood Vessel Transplantation Experiment 1. [Figure 17] Digital and optical microscope images of the graft from Example 4 in Artificial Blood Vessel Transplantation Experiment 2. [Figure 18] Fluorescence micrograph of tissue immunostaining of the graft from Example 4 in Artificial Blood Vessel Transplantation Experiment 2. [Modes for carrying out the invention]

[0035] Embodiments of the present disclosure will be described in detail below with reference to the drawings. The following description of preferred embodiments is illustrative in nature and is not intended to limit the present disclosure, its applications, or its uses in any way.

[0036] <Artificial blood vessels> The artificial blood vessel relating to this disclosure comprises a tubular structure and a coating of a hydrophilic polymer.

[0037] [Tubular structure] The tubular structure is made of nanofibers (hereinafter also referred to as "NF").

[0038] NF has an extremely fine average fiber diameter and a large specific surface area, resulting in characteristics such as being lightweight, thin, soft, stretchable, and breathable and waterproof.

[0039] The average fiber diameter of NF is not particularly limited and may be a general fiber diameter, but from the viewpoint of ensuring the strength and flexibility of the tubular structure, it can be set to, for example, 50 nm to 1000 nm. The average fiber diameter of NF can be determined, for example, from the average fiber diameter of 10 arbitrary NF fiber-shaped portions obtained by scanning electron microscopy (SEM) observation.

[0040] The method for manufacturing NF, or tubular structures, is not particularly limited, and known methods can be employed. Specifically, for example, from the viewpoint of improving the uniformity of the fiber diameter of NF, tubular structures can be manufactured using electrospinning. Melt spinning may also be used.

[0041] NF may be in the form of a nonwoven fabric, woven fabric, or knitted fabric, or a combination thereof, and is preferably a nonwoven fabric.

[0042] The raw material resin of NF serves as the matrix material that forms the fibrous shape of NF. The raw material resin of NF is not particularly limited, and known materials used in artificial blood vessels can be used as appropriate. The raw material resins include biocompatible materials, biodegradable materials, and bio-derived materials, and specifically include, for example, polycaprolactone (PCL), polylactic acid, polyglycolic acid, polyhydroxybutyric acid, polypropylene fumarate, polypropylene carbonate, polyvinylidene fluoride, polyurethane, nylon, polyacrylonitrile, polyester, polyvinyl alcohol, poly(ethylene vinyl alcohol) copolymer, polystyrene, polyvinyl carbonate, polyamide, polyaniline, polyethylene oxide, polyvinylidene chloride, polyethersulfone and copolymers thereof, silk, wool, agarose, alginate, cellulose, oxidized cellulose, collagen, gelatin, albumin, alastin, keratin, chitin, chitosan, elastin, agar, etc. From the viewpoint of ensuring biodegradability, strength, and flexibility of artificial blood vessels, the preferred raw material resin is at least one selected from the group consisting of polycaprolactone (PCL), polylactic acid, polyglycolic acid, polylactic acid-glycolic acid, polyurethane, polyhydroxybutyric acid, and silk.

[0043] The weight-average molecular weight Mw of the raw resin is 10,000 to 500,000, more preferably 30,000 to 200,000. If the weight-average molecular weight Mw is less than 10,000, the strength of the NF may be insufficient, and if it is greater than 5,000,000, the flexibility of the NF may decrease.

[0044] NF may contain additives other than the raw resin as needed. The additives are not particularly limited, and known additives used in artificial blood vessels can be used. Furthermore, salts may be added to facilitate the production of NF, as described later. These additives can be used individually or in combination of two or more.

[0045] Figures 1 to 3 schematically show an example of the tubular structure 100 related to this disclosure.

[0046] As shown in Figures 2(a) and 3(a), the tubular structure 100 has a laminated structure, for example, made up of multiple nanofiber layers 110 stacked on top of each other, and has an inner lumen 120. In this example, three nanofiber layers 110 are shown for simplicity, but the number of layers is not particularly limited and can be, for example, 2 to 1000 layers, preferably 10 to 800 layers.

[0047] In this example, each nanofiber layer 110 consists of a long nanofiber ribbon. As will be described later, the nanofiber ribbon is wound at a predetermined angle to form each nanofiber layer 110. As shown in Figures 2(a) and 3(a), it is preferable that the winding direction of the nanofiber ribbon is opposite for the innermost and outermost nanofiber layers 110 and for the inner nanofiber layer 110.

[0048] As shown by the reference numeral 110C in Figure 3(a), the ends of the nanofiber ribbon in the width direction, i.e., the ends extending in the length direction, are arranged in a spiral pattern on the inner circumferential surface 112 of the tubular structure 100. As will be described in detail later, this arrangement is the result of winding the innermost layer of nanofiber ribbon at a predetermined angle while overlapping it.

[0049] Figures 2(b) and 3(b) show one modified example of the tubular structure 100 shown in Figures 2(a) and 3(a).

[0050] In this modified tubular structure 100, a nanofiber layer 111 made of a nanofiber sheet is arranged in the innermost layer. As will be described in detail later, the nanofiber sheet is not ribbon-shaped. A single nanofiber sheet is formed or wound directly onto a core rod, and then a nanofiber ribbon is wound around the outside of it.

[0051] As shown in Figure 3(b), a nanofiber layer 111 made of nanofiber sheets is arranged on the inner circumferential surface 112 of the tubular structure 100. Since the nanofiber layer 111 is thin, when observed with an electron microscope or the like, the ends of the nanofiber ribbons in the width direction, which are located on the outside of the nanofiber layer 111 (towards the back of the paper in Figure 3(b)), can be seen arranged in a spiral shape.

[0052] Thus, in this specification, "the ends of the nanofiber ribbon in the width direction are arranged helically on the inner circumferential surface of the tubular structure" means the case in which the ends of the nanofiber ribbon in the width direction are arranged helically on the inner circumferential surface 112 of the tubular structure 100, as shown in Figure 3(a), and the case in which the ends of the nanofiber ribbon arranged helically in the width direction can be observed when the inner circumferential surface 112 of the tubular structure 100 is observed with an electron microscope or the like, as shown in Figure 3(b).

[0053] Figures 2(c) and 3(c) show one of the further modifications of the tubular structure 100 shown in Figures 2(b) and 3(b).

[0054] As shown in Figures 2(c) and 3(c), this modified example further includes sutures 130 as reinforcing material placed between two adjacent nanofiber layers 110. The inclusion of sutures 130 improves the strength of the artificial blood vessel.

[0055] The thickness of the nanofiber layer 110 is preferably 5 μm to 100 μm, more preferably 5 μm to 50 μm, more preferably 7 μm to 50 μm, even more preferably 10 μm to 35 μm, and particularly preferably 10 μm to 20 μm, from the viewpoint of ensuring the strength and flexibility of the artificial blood vessel.

[0056] The thickness of the nanofiber layer 111 is preferably 5 μm to 50 μm, more preferably 10 μm to 40 μm, and particularly preferably 15 μm to 25 μm, from the viewpoint of ensuring the strength and flexibility of the artificial blood vessel while ensuring the smoothness of the inner circumferential surface of the artificial blood vessel.

[0057] [Hydrophilic polymer coating] The hydrophilic polymer coating is formed on the surface of the NF that constitutes at least the inner circumferential surface 112 of the tubular structure 100.

[0058] Furthermore, it is preferable that the coating is also formed on the surface of the NF constituting the outer circumferential surface 114 of the tubular structure 100.

[0059] By incorporating a hydrophilic polymer coating, the artificial blood vessel is given antithrombotic properties, improving its patency rate.

[0060] Furthermore, since the rigidity of the resulting NF varies depending on the type and molecular weight of the raw material resin, the rigidity of the tubular structure may be insufficient depending on the raw material resin, especially when considering its use as an artificial blood vessel. By applying a hydrophilic polymer coating, the tubular structure is given rigidity suitable for use as an artificial blood vessel. In addition, the presence of this coating improves the handling properties (anastomosis, sutureability, fray resistance) and kink resistance of the artificial blood vessel.

[0061] The hydrophilic polymer is preferably, though not intended to be limiting, at least one selected from the group consisting of polyvinyl alcohol, polyacrylic acid, sodium polyacrylate, polyacrylamide, polyvinylpyrrolidone, dextran, carboxymethylcellulose, and polyethylene oxide.

[0062] The ratio of the coating to the tubular structure 100 is preferably 2.5% by mass or more and 10% by mass or less, more preferably 3% by mass or more and 8% by mass or less, and particularly preferably 4% by mass or more and 7% by mass or less.

[0063] [Size of artificial blood vessels] Since the aforementioned coating is formed thinly on the surface of the NF, in this specification, the size of the artificial blood vessel is treated as being the same as the size of the tubular structure.

[0064] The inner diameter of the artificial blood vessel according to this disclosure is 1 mm or more and 5 mm or less, preferably 1 mm or more and 4.5 mm or less, and more preferably 1 mm or more and 4 mm or less.

[0065] The outer diameter of the artificial blood vessel relating to this disclosure is, for example, 1.2 mm or more and 6.2 mm or less, preferably 1.3 mm or more and 6.0 mm or less, and more preferably 1.4 mm or more and 5.8 mm or less, without the intention of limiting it.

[0066] The wall thickness of the artificial blood vessel is, for example, 0.2 mm to 1.2 mm, preferably 0.3 mm to 1.0 mm, and more preferably 0.4 mm to 0.8 mm, although this is not intended to be limiting.

[0067] [Compression recovery] For artificial blood vessels with an inner diameter of 2 mm to 5 mm, preferably 2 mm to 4 mm, the compression recovery obtained by the compression test described later is preferably 63% or more, more preferably 63% to 95%, even more preferably 70% to 93%, and particularly preferably 80% to 90%.

[0068] Furthermore, for artificial blood vessels with an inner diameter of 1 mm or more and less than 2 mm, the compressive recovery obtained by the compression test is preferably 41% or more, more preferably 50% to 95%, even more preferably 55% to 93%, and particularly preferably 60% to 90%.

[0069] According to this configuration, the artificial blood vessel can have excellent shape retention and excellent kink resistance.

[0070] [Bending stiffness] The maximum load measured by the three-point bending test described later is preferably 0.0055 N or more, more preferably 0.006 N to 0.05 N, and particularly preferably 0.007 N to 0.02 N, from the viewpoint of ensuring excellent kink resistance of the artificial blood vessel.

[0071] [Retention performance of hydrophilic polymers] One indicator of the retention performance of hydrophilic polymers in artificial blood vessels is the ratio of hydrophilic polymer outflow when distilled water is passed through the artificial blood vessel. The ratio of hydrophilic polymer outflow is expressed as a percentage, representing the ratio of the amount of hydrophilic polymer outflowing from the artificial blood vessel after distilled water is passed through it to the amount of hydrophilic polymer attached to the artificial blood vessel before distilled water is passed through it. The smaller this value, the higher the retention performance of the hydrophilic polymer in the artificial blood vessel.

[0072] In the artificial blood vessel relating to this disclosure, the hydrophilic polymer outflow rate calculated by the hydrophilic polymer outflow rate evaluation test described later is preferably 25% or less, more preferably 20% or less, and particularly preferably 19.5% or less.

[0073] When the outflow ratio exceeds the upper limit, the patency rate of the artificial blood vessel decreases. In the artificial blood vessel according to this disclosure, the hydrophilic polymer has high retention performance, so excellent antithrombotic properties can be maintained over a long period of time, and the patency rate can be improved.

[0074] [Water contact angle] The water contact angle of the NF surface of the nanofiber sheet (with coating), as measured by the contact angle measurement test 1 described later, is preferably 130° or less, more preferably 100° or less, even more preferably 50° or less, and particularly preferably 30° or less.

[0075] The water contact angle of the inner surface of the artificial blood vessel, as measured by the contact angle measurement test 2 described later, is preferably 120° or less, more preferably 100° or less, and particularly preferably 30° or less. If it is difficult to measure the water contact angle after dropping a droplet onto the inner surface of the artificial blood vessel and the droplet is absorbed, the time from dropping the droplet to absorption is preferably less than 5 seconds, more preferably 3 seconds or less, and particularly preferably 1 second or less.

[0076] The water contact angle of the outer surface of the artificial blood vessel, as measured by the contact angle measurement test 2 described later, is preferably 120° or less, more preferably 100° or less, and particularly preferably 30° or less. If it is difficult to measure the water contact angle after dropping a droplet onto the outer surface of the artificial blood vessel and the droplet is absorbed, the time from dropping the droplet to absorption is preferably 15 seconds or less, more preferably 5 seconds to 12 seconds, and particularly preferably 5 seconds to 10 seconds.

[0077] It is preferable that the hydrophilicity of the inner surface of the artificial blood vessel is higher than that of the outer surface.

[0078] In other words, it is preferable that the water contact angle of the inner surface of the artificial blood vessel is smaller than the water contact angle of the outer surface.

[0079] Furthermore, in the contact angle test, when droplets are dropped onto the inner and outer surfaces of the artificial blood vessel, if the droplets are absorbed, it is preferable that the time required for absorption of the droplets on the inner surface is shorter than the time required for absorption of the droplets on the outer surface.

[0080] [Platelet adsorption suppression rate] One indicator of the antithrombotic properties of artificial blood vessels is the platelet adsorption inhibition rate, calculated from the results of the platelet adsorption test described later. The platelet adsorption inhibition rate is an indicator that shows how much the number of platelet granules adsorbed by an artificial blood vessel (a coated tubular structure) is suppressed compared to a tubular structure without a coating, in the platelet adsorption test described later. Specifically, the platelet adsorption inhibition rate X can be defined as, for example, the ratio of the number of platelet granules N1 attached to a tubular structure without a coating to the number of platelet granules N2 attached to the artificial blood vessel (a coated tubular structure), expressed as a percentage (X = [(N1-N2) / N1] × 100). The larger this value, the higher the antithrombotic properties of the artificial blood vessel.

[0081] From the viewpoint of ensuring excellent antithrombotic properties of the artificial blood vessel, the platelet adsorption inhibition rate of the artificial blood vessel is preferably 30% or more, more preferably 40% or more, even more preferably 50% or more, and particularly preferably 54% or more.

[0082] [Patency rate] In the artificial blood vessel transplantation experiment 1 described later, the patency rate of the artificial blood vessel two weeks after transplantation of the heparin-coated artificial blood vessel into the abdominal working aorta of a rat is preferably 84% or higher, more preferably 87% or higher, and particularly preferably 90% or higher. Furthermore, in the same transplantation experiment 1, the patency rate of the artificial blood vessel eight weeks after transplantation is preferably 76% or higher, more preferably 80% or higher, and particularly preferably 83% or higher.

[0083] In the artificial blood vessel transplantation experiment 2 described later, the patency rate of the artificial blood vessel 8 weeks after transplantation of an uncoated artificial blood vessel into the working abdominal aorta of a rat is preferably 80% or more, more preferably 83% or more.

[0084] Having the above-mentioned patency rate, the artificial blood vessel according to this disclosure can actually function as an artificial blood vessel when implanted in the body and can be said to be fully suitable for practical use.

[0085] <Manufacturing method for artificial blood vessels> The following describes an example of a method for manufacturing artificial blood vessels related to this disclosure. However, the method for manufacturing artificial blood vessels related to this disclosure is not limited to the following.

[0086] The method for manufacturing an artificial blood vessel specifically includes, for example, a tubular structure formation step S1 and a coating formation step S2, as shown in Figure 4.

[0087] [Tubular structure formation process] Step S1, the tubular structure formation step, is the step of forming a tubular structure made of nanofibers. The method for forming the tubular structure is not particularly limited as described above, but for example, electrospinning can be used. For example, the tubular structure may be formed by directly electrospinning a core rod.

[0088] From the viewpoint of improving the kink resistance of artificial blood vessels, a more preferred method includes, for example, a nanofiber sheet fabrication step S11 (first step), a nanofiber ribbon fabrication step S12 (second step), and a winding step S13 (third step).

[0089] -Nanofiber sheet fabrication process- As shown in Figure 5(a), in the nanofiber sheet fabrication step S11, the nanofiber sheet 111A is fabricated by electrospinning. The shape of the nanofiber sheet 111A is not particularly limited, but it is preferably rectangular.

[0090] First, the raw material resin for the nanofiber sheet 111A is dissolved in a solvent to prepare a resin solution 51. The resin solution 51 is prepared, for example, by weighing the raw material resin into a container, adding a solvent and additives, and stirring with a known stirring means (not shown), such as a stirrer, to dissolve and disperse the resin and additives in the solvent. Heating or ultrasonic waves may be applied simultaneously during stirring.

[0091] The solvent dissolves the resin, adjusts the viscosity of the resin solution 51, and evaporates during spinning to promote the formation of the fiber shape. Specific examples of solvents include dimethylformamide (DMF), dimethylacetamide (DMAc), dimethyl sulfoxide (DMSO), N-methyl-2-pyrrolidone (NMP), toluene, benzotrifluoride, 10-camphor sulfonic acid, isopropyl alcohol (IPA), hexafluoroisopropyl alcohol (HFIP), methyl ethyl ketone (MEK), acetone, tetrahydrofuran (THF), diethyl ether, dichloromethane, chloroform, ethanol, formic acid, hydrochloric acid, trifluoroacetic acid, ethyl acetate, and water. One or more of these solvents may be used. Note that these solvents evaporate during spinning and are therefore not contained in the nanofiber sheet 111A.

[0092] Additives that can be included in the resin solution 51 include, for example, salts of sodium chloride, sodium acetate, lithium chloride, calcium chloride, magnesium chloride, etc., from the viewpoint of improving the solubility and dispersibility of the resin and promoting the uniform formation of fine fiber shapes.

[0093] The concentration of the resin in the resin solution 51 is preferably 5% by mass or more and 50% by mass or less, more preferably 7% by mass or more and 40% by mass or less, and particularly preferably 10% by mass or more and 30% by mass or less, from the viewpoint of obtaining a nanofiber sheet 111A with a uniform and fine fiber shape. Furthermore, when the above salt is added, the concentration of the salt is preferably 2% by mass or less, more preferably 1% by mass or less, and particularly preferably 0.5% by mass or less, from the viewpoint of obtaining a nanofiber sheet 111A with a uniform and fine fiber shape. Furthermore, the concentration of additives other than salt can be, depending on the type of additive, for example, 0.5% by mass or more and 3% by mass or less, depending on the purpose.

[0094] Next, the resin solution 51 is spun using an electrolytic spinning method to obtain a nanofiber sheet 111A.

[0095] Spinning by electrospinning can be performed, for example, using the electrospinning apparatus shown in Figure 5(a). Specifically, for example, the resin solution 51 is sprayed from the tip of a syringe 33 onto the surface of a rotary drum-type collector 35. However, the method of spinning by electrospinning is not limited to the method shown in Figure 5(a), and any known method can be used as appropriate.

[0096] As shown in Figure 5(a), a high-voltage power supply 34 is connected to the syringe 33 and the collector 35, and a predetermined high voltage is applied between them. The resin solution 51 in the syringe 33 can be at room temperature, for example, 15°C to 30°C.

[0097] The voltage applied by the high-voltage power supply 34 can be, for example, 5kV to 80kV, preferably 10kV to 50kV, from the viewpoint of obtaining nanofibers with a uniform fiber shape and minute diameter.

[0098] The distance from the tip of the syringe 33 to the collector 35 can be, for example, 50 mm to 200 mm, from the viewpoint of obtaining nanofibers with a uniform fiber shape and minute diameter.

[0099] The thickness of the nanofiber sheet 111A obtained in this way (and the nanofiber ribbon 110A manufactured in the next step) is preferably 5 μm to 100 μm, more preferably 5 μm to 50 μm, more preferably 7 μm to 50 μm, even more preferably 10 μm to 35 μm, and particularly preferably 10 μm to 20 μm.

[0100] -Nanofiber ribbon fabrication process- As shown in Figure 5(b), in the nanofiber ribbon fabrication step S12, the nanofiber sheet 111A obtained as described above is cut to produce a long nanofiber ribbon 110A.

[0101] Figure 5(b) illustrates an example of cutting using scissors, but the method of cutting the nanofiber sheet 111A is not particularly limited, and other known means such as a cutting machine may be used.

[0102] Furthermore, it is preferable that the width and length of the nanofiber ribbon 110A are uniform. Specifically, for example, the width of the nanofiber ribbon 110A is preferably 50 mm or less, preferably 5 mm to 40 mm, and more preferably 10 mm to 35 mm.

[0103] Furthermore, the length of the nanofiber ribbon 110A is preferably 30 cm or more, more preferably 30 cm to 100 cm, and particularly preferably 40 cm to 80 cm.

[0104] -Winding process- In the winding step S13, as shown in Figure 5(c), nanofiber ribbons 110A, preferably multiple nanofiber ribbons 110A, are wound around the core rod 150 at a predetermined angle i. Finally, the core rod 150 is removed to form the tubular structure 100.

[0105] The predetermined angle i is the angle between the axial direction of the tubular structure 100 and the longitudinal direction of the nanofiber ribbon 110A, and is an angle greater than 0° and less than 90°. The predetermined angle i can be, for example, 10° or more and 80° or less, preferably 20° or more and 75° or less, and more preferably 50° or more and 70° or less.

[0106] It is preferable to wind the nanofiber ribbon 110A while overlapping it with the already wound portion. Specifically, for example, when winding the nanofiber ribbon 110A around the core rod 150, it is preferable to wind it so that one end 110B (end) in the width direction of the nanofiber ribbon 110A is visible from the outside, and the other end 110C (end) is hidden by the nanofiber ribbon 110A. This ensures sufficient flexibility and strength of the artificial blood vessel.

[0107] Furthermore, in the winding process S13, it is desirable to wind multiple nanofiber ribbons 110A alternately in opposite directions around the core rod 150.

[0108] Specifically, for example, in step A, the first nanofiber ribbon 110A is wound around the core rod 150 from one side to the other at a predetermined angle i. Then, in step B, the second nanofiber ribbon 110A is wound around the core rod 150 on which the first nanofiber ribbon 110A is wound, from the other side to the other at a predetermined angle i. Then, by repeating steps A and B described above, a tubular structure 100 is formed.

[0109] This forms the tubular structure 100 shown in Figures 2(a) and 3(a).

[0110] In this way, by winding a long nanofiber ribbon 110A around a core rod 150 to fabricate a tubular structure 100, the thickness of the tubular structure 100 can be easily increased compared to the case where nanofibers are directly electrospun onto the core rod 150 and laminated. This reduces the manufacturing time of the tubular structure 100. Furthermore, nanofibers have an enormous specific surface area due to their extremely fine fiber diameter, and by winding the nanofiber ribbons in layers, a strong van der Waals force derived from this enormous specific surface area can be obtained. As a result, the cylindrical shape of the tubular structure 100 can be maintained without the use of adhesive.

[0111] The tubular structure 100 formed in this manner is excellent in both strength and flexibility, making it suitable as a base material for artificial blood vessels.

[0112] For example, to form the tubular structure 100 shown in Figures 2(b) and 3(b), the core rod 150 can be set as the collector 35 in Figure 5(a) before winding the nanofiber ribbon 110A around the core rod 150, and the nanofiber layer 111 can be directly formed on the core rod 150 by electrospinning.

[0113] Alternatively, before winding the nanofiber ribbon 110A onto the core rod 150, the nanofiber sheet 111A manufactured in the nanofiber sheet manufacturing process S1 may be wound onto the core rod 150. At this time, one or more nanofiber sheets 111A may be wound, but from the viewpoint of ensuring the flexibility of the artificial blood vessel, it is preferable to use one sheet.

[0114] To form the tubular structure 100 shown in Figures 2(c) and 3(c), a predetermined number of nanofiber ribbons 110A can be wound around the core rod 150, followed by winding suture thread 130, and then winding another predetermined number of nanofiber ribbons 110A. Alternatively, the suture thread 130 may be wound around the core rod 150 before winding the nanofiber ribbons 110A. If necessary, multiple suture threads 130 may be wound around the core rod 150 in a crisscross pattern.

[0115] For example, in a method of forming a tubular structure 100 by electrospinning nanofibers directly onto a core rod, depending on the nanofiber lamination rate, it may take a long time to laminate them to a wall thickness suitable for use in artificial blood vessels. Also, in a method of simply forming nanofiber sheets into a cylindrical shape and laminating them to create a tubular structure, adhesives or the like must be applied to the overlapping parts of the nanofiber sheets in order to maintain the shape. This complicates the manufacturing process of artificial blood vessels, and the adhesive may peel off when the artificial blood vessel is anastomosed to the target blood vessel for transplantation.

[0116] As described above, the method of forming a tubular structure 100 by winding a long nanofiber ribbon 110A allows for easy thickness creation of the tubular structure and reduces manufacturing time. Furthermore, NF has an enormous specific surface area due to its extremely fine fiber diameter, and by winding multiple nanofiber ribbons 110A, a strong van der Waals force derived from this enormous specific surface area can be obtained. As a result, the shape can be maintained without the use of adhesives. Moreover, this method can impart appropriate elasticity to the artificial blood vessel and improve its kink resistance.

[0117] [Coating Formation Process] In the coating formation step S2, as shown in Figure 5(d), a solution containing a hydrophilic polymer is passed through the lumen 120 of the tubular structure 100 to form a hydrophilic polymer coating.

[0118] In detail, when a solution containing a hydrophilic polymer is passed through the lumen 120 of the tubular structure 100, the solution penetrates the walls of the tubular structure 100. Thus, the surface of the nanofibers in the tubular structure 100 is coated with the hydrophilic polymer.

[0119] A hydrophilic polymer solution consists of a hydrophilic polymer and a solvent. The solvent is not particularly limited as long as the solubility of the hydrophilic polymer can be ensured, and is, for example, water, a mixed solvent of water and a water-soluble organic solvent such as alcohol, preferably a mixed solvent of water and ethanol. By using a mixed solvent, the penetration of the solution into the wall of the tubular structure is improved. In detail, NF has many voids between its fine fibers, and these voids are extremely fine, so much so that water cannot pass through but air can. Water-soluble organic solvents, which have lower surface tension than water, can pass through these extremely fine voids. Therefore, when a mixed solvent is used, the hydrophilic polymer solution penetrates into the extremely fine voids of NF by capillary action, penetrating not only the inner circumferential surface of the artificial blood vessel but also the outer circumferential surface. In this way, only the fiber surface of the NF can be coated with the hydrophilic polymer over the entire tubular structure.

[0120] When a mixed solvent is used, the content of the water-soluble organic solvent in the mixed solvent is, for example, 1% by mass or more and 50% by mass or less, preferably 10% by mass or more and 40% by mass or less. Furthermore, the concentration of the hydrophilic polymer contained in the solution is, from the viewpoint of ease of liquid flow and ensuring a sufficient coating amount, for example, 0.1% by mass or more and 15% by mass or less, preferably 0.5% by mass or more and 10% by mass or less, more preferably 1% by mass or more and 5% by mass or less.

[0121] After passing a hydrophilic polymer solution through it, the tubular structure 100 is dried by standing for 0.5 hours or more, preferably 1 hour or more and up to 300 hours or less, in an environment with a temperature of 10 to 40°C and a humidity of 20 to 60%, for example, to obtain an artificial blood vessel 101.

[0122] This method, which involves passing a hydrophilic polymer solution through the lumen 120 of a tubular structure 100, allows for the formation of a thin coating only on the fiber surface of NF with a simple process. This simplifies the manufacturing process and suppresses the embedding of NF voids into the hydrophilic polymer, which is advantageous for artificial blood vessels. Furthermore, forming a coating using this method effectively suppresses the elution of the hydrophilic polymer during use of the artificial blood vessel, resulting in an artificial blood vessel with superior antithrombotic and patency properties. [Examples]

[0123] Next, we will describe specific examples of the implementations.

[0124] <Fabrication of artificial blood vessels> The artificial blood vessels of the examples and comparative examples shown in Table 1 were fabricated using the following procedure.

[0125] [Table 1]

[0126] [Example 1] Polycaprolactone (PCL, manufactured by Sigma-Aldrich, average Mn 80000) was added to a solvent mixture of DMF, THF, and MEK (in a mass ratio of 3:6:1) to achieve a resin concentration of 10% by mass in the resin solution, and the mixture was stirred to dissolve it. This resin solution was spun onto a collector using an electrospinning apparatus (manufactured in-house, Figure 5(a)) under the conditions of an applied voltage of 20kV, a syringe tip-collector distance of 200mm, and a temperature of 20-25°C to form nanofiber sheets (NF sheets) with a thickness of 25μm-30μm.

[0127] Next, the NF sheet was cut into strips 20 mm wide and 40 cm long to obtain long nanofiber ribbons (NF ribbons).

[0128] An NF ribbon was wound around a 3mm outer diameter core rod at a 60° angle to the axial direction, from one end to the other. The NF ribbon was overlapped during this winding process so that the other end of the NF ribbon was concealed. Then, another NF ribbon was wound on top of this, from the other end to the first, at a 60° angle, overlapping the NF ribbon so that one end of the previous NF ribbon was concealed. By repeatedly winding the NF ribbons alternately in opposite directions in this manner, a layer of NF ribbon was created. Afterward, the core rod was removed to obtain a tubular structure.

[0129] Next, 6 mL of polyvinyl alcohol solution (PVA solution) was passed through the lumen of the obtained tubular structure. The PVA solution was prepared by mixing a commercially available PVA aqueous solution (Kaneyonol, manufactured by Kaneyo Soap Co., Ltd.), distilled water, and ethanol in a mass ratio of 1:1:1. The PVA concentration in the commercially available PVA aqueous solution was measured to be 5% by mass.

[0130] Subsequently, the tubular structure was left to stand for more than 24 hours in an environment of 20°C and 40% humidity to obtain an artificial blood vessel with the specifications shown in Table 1.

[0131] [Example 2] An artificial blood vessel was fabricated in the same manner as in Example 1, except that the sutures were wrapped using the following procedure.

[0132] After laminating NF ribbons until the wall thickness of the tubular structure was approximately 0.15 mm, suture thread (Johnson & Johnson, Vicryl, PCL, approximately 0.4 mm in diameter) was wound from one side to the other at a 60° angle relative to the axial direction, and then wound from the other side to the first side at a 60° angle. Subsequently, NF ribbons were laminated until the final wall thickness of the tubular structure was approximately 1 mm.

[0133] [Example 3] An artificial blood vessel was fabricated in the same manner as in Example 1, except that an artificial blood vessel with the specifications shown in Table 1 was obtained using a core rod with an outer diameter of 2 mm.

[0134] [Example 4] An artificial blood vessel was fabricated in the same manner as in Example 1, except that an artificial blood vessel with the specifications shown in Table 1 was obtained using a core rod with an outer diameter of 1 mm.

[0135] [Example 5] In the apparatus shown in Figure 5(a), an artificial blood vessel was fabricated in the same manner as in Example 1, except that the collector was replaced with a core rod and a tubular structure was obtained by spinning directly onto the core rod.

[0136] [Comparative Example 1] An artificial blood vessel was fabricated in the same manner as in Example 1, except that the PVA solution was not passed through the tubular structure.

[0137] [Comparative Example 2] An artificial blood vessel was fabricated in the same manner as in Example 4, except that the PVA solution was not passed through the tubular structure.

[0138] [Comparative Example 3] An artificial blood vessel was fabricated in the same manner as in Example 5, except that the PVA solution was not passed through the tubular structure.

[0139] [Comparative Example 4] An artificial blood vessel was fabricated in the same manner as in Example 1, except that a PCL film, formed by coating resin solution 51 using an automatic coating device (AFA-Standard: Cotec Corporation, Wireless Bar Coater 30H), was wound onto a core rod to obtain a tubular structure.

[0140] <Evaluation Test> [Observation of the appearance using a digital camera] Figure 6(a) shows digital camera photographs of the tubular structures of Examples 1, 3, and 4. From the markings on the ruler placed nearby, it can be seen that tubular structures with approximately the desired inner diameter have been obtained.

[0141] Figure 6(b) is a digital camera photograph of the tubular structure of Example 2. The shape of the wound suture can be seen from the external appearance.

[0142] [Scanning Electron Microscope (SEM) Observation] Figure 7 shows an SEM image of the artificial blood vessel from Example 4.

[0143] Figures 7(a), (c), and (d) are SEM images of the radial cross-section at 40x, 250x, and 600x magnification, respectively. Figure 7(b) is an SEM image of the axial cross-section at 30x magnification.

[0144] From Figures 7(a), (c), and (d), the artificial blood vessel appears to have a layered structure consisting of approximately five layers with a thickness of about 50 to 100 μm. However, each layer itself is made up of several to tens of NF ribbons stacked on top of each other (i.e., the whole structure is made up of tens to hundreds of NF ribbons stacked on top of each other). It is thought that this cross-sectional structure was obtained because adjacent NF ribbons are in close contact with each other in the radial direction.

[0145] As described above, the ends of the NF ribbon in the width direction, i.e., the ends extending in the length direction, are arranged in a spiral pattern on the inner surface of the tubular structure made by winding the NF ribbon. In Figure 7(b), a portion of these spirally arranged ends is visible.

[0146] Furthermore, as shown in Figure 7(b), the outer layer of the tubular structure is wavy, meaning the outside of the tubular structure is somewhat bellows-like. This is thought to be due to the load applied when the core rod is pulled out of the tubular structure.

[0147] Figure 8 shows SEM images of the artificial blood vessel of Example 2. Figures 8(a) and 8(b) are SEM images of the radial cross-section at 20x and 75x magnification, respectively.

[0148] As shown in Figure 8, sutures are placed between the layers of the NF ribbon.

[0149] Figures 9(a) and 9(b) show SEM images of the inner surface of the artificial blood vessels of Comparative Example 1 and Example 1, respectively. Comparing the two, it can be seen that the voids between nanofibers observed in the artificial blood vessel of Example 1 with PVA coating are not filled in by the PVA, even in the artificial blood vessel of Comparative Example 1 without PVA coating. In other words, it is thought that the PVA coats the fiber surface of the nanofibers, and the permeability and fluid permeability of the nanofiber layer are maintained before and after the PVA coating process.

[0150] [Compression test] The artificial blood vessel was subjected to compression tests under the following test conditions.

[0151] Test equipment: KES-F3-A manufactured by Kato Tech SENS:2 Pressurized area: 2cm 2 Compression speed: 0.002 cm / s Maximum load: 50gf / cm 2 The results are shown in Table 2, Figure 10, and Table 3. Table 2 and Figure 10 show the test results for artificial blood vessels with an inner diameter of 3 mm in Examples 1 and 2 and Comparative Examples 1 and 3. Table 3 shows the test results for artificial blood vessels with an inner diameter of 1 mm in Example 4 and Comparative Example 2.

[0152] [Table 2]

[0153] [Table 3]

[0154] Tables 2 and 3 show LC, WC, and RC, respectively, representing compression linearity, compression work, and compression recovery. These values ​​were calculated based on the graph in Figure 10. WC[gf*cm / cm 2] is a value represented by the sum of the areas of region a and region b shown in Figure 11, and a larger value indicates that it is more easily compressed. LC is a value obtained by dividing the value of WC by the area of ​​triangle ABC in Figure 11, and a value closer to 1 indicates that it is less easily compressed. RC[%] is a value obtained by the following formula (1) using the compression recovery work WC' (area of ​​region b shown in Figure 11) and WC, and a value closer to 100% indicates greater recovery.

[0155] RC = (WC' / WC) × 100 ... (1) The results in Tables 2 and 3 show that the RC (resilience coefficient) of the artificial blood vessel in the example (with PVA coating) is larger than that of the comparative example (without PVA coating). This indicates that applying PVA coating to tubular structures improves shape retention and kink resistance.

[0156] [Three-point bending test] Three-point bending tests were performed on the artificial blood vessels (3 mm inner diameter) of Examples 1, 2, and 5 and Comparative Examples 1 and 3. The test conditions are shown below.

[0157] Testing equipment: Instron 5567 electromechanical universal material testing machine. Speed: 5mm / min Support span: 40mm The test was stopped when the measured intensity became constant or decreased, and the maximum load during the measurement was evaluated. The test results are shown in Table 4.

[0158] [Table 4]

[0159] The artificial blood vessels in Comparative Example 1 and Examples 1 and 2 were fabricated by winding NF ribbon around a core rod. It was found that the maximum load on the artificial blood vessels in Examples 1 and 2 with PVA coating was more than twice as large as that on the artificial blood vessel in Comparative Example 1 without PVA coating.

[0160] The artificial blood vessels in Comparative Example 3 and Example 5 were fabricated by directly spinning fibers onto a core rod. It was found that the maximum load on the artificial blood vessel in Example 5 with PVA coating was approximately 1.8 times greater than that on the artificial blood vessel in Comparative Example 3 without PVA coating.

[0161] Thus, it was found that applying a PVA coating increases the maximum load capacity of the artificial blood vessel and improves its kink resistance.

[0162] [Evaluation test of hydrophilic polymer runoff] 20 mL of distilled water was passed through the lumen of the artificial blood vessels of Example 1 and Comparative Example 4 at a flow rate of 1.5 mL / min, and the amount of PVA outflow was compared. The test results are shown in Table 5.

[0163] [Table 5]

[0164] However, in Table 5, the mass of the tubular structure is the mass of the tubular structure before PVA coating was applied. The mass of the artificial blood vessel is the value measured for the artificial blood vessel obtained by applying PVA coating to the tubular structure and leaving it to stand for 24 hours or more in an environment of 20°C and 40% humidity as described above. The difference between the mass of the artificial blood vessel and the mass of the tubular structure was defined as the amount of PVA attached. Furthermore, the mass of the artificial blood vessel after passing distilled water through it was measured after leaving the artificial blood vessel to stand for 24 hours or more in an environment of 20°C and 40% humidity after passing distilled water through it.

[0165] The amount of PVA discharged was calculated using the following formula (2).

[0166] PVA flow rate = Amount of PVA attached - (Mass of artificial blood vessel after passing distilled water through it - Mass of tubular structure) ... (2) The PVA outflow ratio [%] was then calculated using the following formula (3).

[0167] PVA outflow ratio = PVA outflow / PVA adhesion × 100 ... (3) As shown in Table 5, the amount of PVA outflow relative to the initial amount of PVA attached was 19.1% or less for samples 1-3 of Example 1, while it was 25.4% or more for samples 4-6 of Comparative Example 4, demonstrating the superior PVA retention performance of nanofiber tubular structures compared to film tubular structures.

[0168] [Contact Angle Measurement Test 1] -Manufacturing Example 1- As shown in Figure 12(a), a Teflon® sheet was wrapped around a 20 mm diameter core rod 200, and then a PCL nanofiber sheet (NF sheet) 201 with release paper was wrapped around it five times perpendicular to the longitudinal direction of the core rod 200, with the NF side facing inward. The thickness of the NF sheet 201 was approximately 35 μm and the width was approximately 6 cm.

[0169] Next, the core rod 200 and the Teflon sheet were removed to obtain a cylindrical NF sheet 202.

[0170] Then, as shown in Figure 12(b), the PVA solution used in the fabrication of the artificial blood vessel in Example 1 was sprayed into the lumen of the cylindrical sheet 202 using the atomizer 203, thereby coating the inner layer (NF surface) of the cylindrical sheet 202 with PVA.

[0171] After air-drying at room temperature, the coated cylindrical sheet 202 was cut open as shown by the dashed line in Figure 12(c). After placing a weight on the cut sheet to remove any curvature, it was cut into 6cm x 6cm squares to obtain samples, and the NF surface was subjected to a θ / 2 method contact angle test. A contact angle measuring device (Kyowa Interface Science Co., Ltd.: CA-X) was used to measure the contact angle.

[0172] -Manufacturing Example 2- A sample was prepared in the same manner as in Manufacturing Example 1, except that a PVA coating was not applied, and its NF surface was subjected to the θ / 2 method contact angle test.

[0173] -result- The contact angles were measured for five samples each from manufacturing examples 1 and 2. The results are shown in Table 6 and Figure 13. Figure 13 is a photograph taken during the contact angle measurement of sample 1.

[0174] [Table 6]

[0175] As shown in Table 6 and Figure 13, it was found that applying a PVA coating to the NF sheet improved the hydrophilicity (wettability) of the NF surface.

[0176] [Contact Angle Measurement Test 2] The artificial blood vessels of Comparative Example 1 and Example 1 were cut in half axially, and then weights were placed on them to remove the curvature. The contact angles were measured at five locations each on the inner and outer surfaces of the artificial blood vessels of Comparative Example 1 and Example 1 using a contact angle measuring device (Kyowa Interface Science Co., Ltd.: CA-X) with the θ / 2 method. The results are shown in Table 7.

[0177] [Table 7]

[0178] However, in Table 7, "nd 1" indicates that the droplet was completely absorbed by the artificial blood vessel within 1 second of contact with the artificial blood vessel surface, making measurement impossible. Also, "nd 2" indicates that the droplet was completely absorbed by the artificial blood vessel within 5 to 10 seconds of contact with the artificial blood vessel surface, making measurement impossible.

[0179] As shown in Table 7, it was found that the artificial blood vessel of Example 1 exhibited improved hydrophilicity (wettability) compared to the artificial blood vessel of Comparative Example 1. Furthermore, comparing the inner and outer surfaces of the artificial blood vessel of Example 1, it was found that the inner surface absorbed droplets faster and had higher hydrophilicity (wettability) than the outer surface. This is thought to be because, in the artificial blood vessel of Example 1, the nanofibers on the inner surface had a greater amount of PVA attached to them than the nanofibers on the outer surface.

[0180] [Platelet adsorption test] Artificial blood vessels (1 mm inner diameter) from Example 4 and Comparative Example 2 were cut to prepare grafts approximately 1 cm in length. 2 ml of concentrated human platelets collected from healthy individuals was passed through the lumen of the grafts three times. Then, the grafts were flushed with 10 ml of phosphate buffer to wash away any unadsorbed platelets. The grafts were fixed with glutaraldehyde, and their inner surface was observed using a scanning electron microscope (SEM). SEM images of the inner surface were taken at a magnification of 5000x, with one field of view (440 μm) being used. 2 The number of platelet granules adsorbed per 100 μm is counted, and the total size is 100 μm. 2 The results were converted to a per-unit value. The experiment was conducted once on each of five independent grafts, for a total of five trials. The results are shown in Table 8, Figure 14, and Figure 15.

[0181] [Table 8]

[0182] Figure 14 shows SEM images of the grafts from Example 4 and Comparative Example 2. As shown in Figure 14, it can be seen that the number of attached platelet granules in the graft from Example 4 is significantly lower than that of the graft from Comparative Example 2.

[0183] As shown in Table 8 and Figure 15, the platelet adsorption inhibition rate, expressed by the following formula (4), was calculated to be 54% based on the average number of platelet granules obtained in five experiments.

[0184] Platelet adsorption suppression rate =[(Average value of Comparative Example 2)-(Average value of Example 4)] / (Average value of Comparative Example 2)×100 ...(4) In other words, it was found that the graft of Example 4, which was PVA-coated, had a 54% reduction in the number of platelet granules adsorbed to the artificial blood vessel compared to the graft of Comparative Example 2, which was not PVA-coated, thus improving its antithrombotic properties.

[0185] [Artificial blood vessel transplantation experiment 1 (with heparin coating)] The artificial blood vessels (1 mm inner diameter) of Example 4 and Comparative Example 2 were implanted into the working abdominal aorta of rats to investigate whether they actually function as artificial blood vessels.

[0186] Prior to transplantation, grafts approximately 1 cm in length were prepared for the artificial blood vessels of Example 4 and Comparative Example 2. These grafts were hydrolyzed by alkaline hydrolysis and then immobilized with heparin by covalent bonds. Specifically, the grafts were hydrolyzed by exposing them to a 2 M sodium hydroxide aqueous solution for 30 minutes and then washed with distilled water. Subsequently, the grafts were immersed overnight at room temperature in a heparin solution with the composition shown in Table 9. The grafts were then washed with distilled water to remove excess heparin.

[0187] [Table 9]

[0188] Grafts coated with heparin using the procedure described above were transplanted into the abdominal working aorta of rats. The number of transplanted rats was 24 for each of the grafts used in Example 4 and Comparative Example 2. At 2 and 8 weeks post-transplantation, the artery distal to the anastomosis was hemectomized, and graft patency was evaluated by checking for blood flow. The results are shown in Table 10.

[0189] [Table 10]

[0190] Furthermore, autologous blood vessel-like regeneration was evaluated by immunofluorescence staining for CD31, a vascular endothelial cell marker, and alpha-smooth muscle (αSM), a smooth muscle marker. Figure 16 shows fluorescence micrographs of the radial cross-section tissue of the graft from Example 4.

[0191] As shown in Table 10, the patency rates at 2 weeks and 8 weeks were found to be improved in Example 4 compared to Comparative Example 2. Furthermore, as shown in Figure 16, regeneration of the vascular intima-like structure was observed in the lumen of the graft in Example 4. These results demonstrate that the PVA-coated artificial blood vessel (1 mm inner diameter) of Example 4 can function as an artificial blood vessel when heparin-coated.

[0192] [Artificial blood vessel transplantation experiment 2 (without heparin coating)] For the artificial blood vessel (1 mm inner diameter) of Example 4, a graft approximately 1 cm in length was prepared and transplanted into the working abdominal aorta of a rat without heparin coating for examination. Eight weeks after transplantation, the artery distal to the anastomosis was halved, and graft patency was evaluated by the presence or absence of blood flow. Digital microscope images of the graft's appearance and radial cross-section at eight weeks post-transplantation are shown in Figures 17(a) and (b), respectively.

[0193] In an evaluation of 12 rats, the patency rate at 8 weeks post-transplantation was 83.3% (10 / 12 rats), which was comparable to the results obtained in transplantation experiment 1 when heparin coating was applied. All harvested grafts showed a smooth texture in the patented cases, and no thrombi were observed in the lumen.

[0194] Next, hematoxylin-eosin staining was performed to evaluate the recellularization of the graft. Optical micrographs of the axial and radial sections of the graft are shown in Figures 17(c) and (d), respectively. As shown in Figures 17(c) and (d), cell engraftment was observed in the lumen, circumferential surface, and within the graft, with the cell layer forming on the luminal side.

[0195] Autologous blood vessel-like regeneration was evaluated by immunofluorescence staining for CD31, a vascular endothelial cell marker, and αSM, a smooth muscle marker. Fluorescence micrographs of the radial cross-section of the graft tissue are shown in Figure 18.

[0196] The results showed that the CD31(+) / αSM(+) cell layer was observed in all proximal, intermediate, and distal regions of the graft, indicating that neointimal regeneration had occurred and nearly completely covered the graft from edge to edge.

[0197] These results demonstrate that the PVA-coated artificial blood vessel (1 mm inner diameter) of Example 4 can function as an artificial blood vessel even without heparin coating. [Explanation of Symbols]

[0198] 100 Tubular structures 101 Artificial blood vessels 110 nanofiber layer 110A Nanofiber Ribbon 110B One end (end) 110C Other end (end) 111 Nanofiber layer 111A Nanofiber Sheet 112 Inner surface 114 Outer surface 120 lumen 130 sutures

Claims

1. A tubular structure made of nanofibers, The tubular structure comprises a hydrophilic polymer coating formed on the surface of the nanofibers constituting at least the inner circumferential surface, The inner diameter is between 1 mm and 5 mm. The tubular structure has a laminated structure in which multiple long nanofiber ribbons are stacked, On the inner circumferential surface of the tubular structure, the ends of the nanofiber ribbon in the width direction are arranged in a helical pattern. An artificial blood vessel characterized by the following features.

2. The ratio of the coating to the tubular structure is 2.5% by mass or more and 10% by mass or less. The artificial blood vessel according to claim 1.

3. When distilled water is passed through the artificial blood vessel, the outflow ratio of the hydrophilic polymer is 25% or less. The artificial blood vessel according to claim 1 or 2.

4. The water contact angle of the inner surface of the artificial blood vessel is 120° or less. The artificial blood vessel according to claim 1 or 2.

5. The coating is also formed on the surface of the nanofibers that constitute the outer surface of the tubular structure. The artificial blood vessel according to claim 1 or 2.

6. The water contact angle of the outer surface of the artificial blood vessel is 120° or less. The artificial blood vessel according to claim 5.

7. In the platelet adsorption test, the platelet adsorption inhibition rate, expressed as a percentage of the number of platelet granules attached to the uncoated tubular structure minus the number of platelet granules attached to the artificial blood vessel, is 30% or more. The artificial blood vessel according to claim 1 or 2.

8. The compression recovery rate obtained by compression testing for the aforementioned artificial blood vessels with an inner diameter of 2 mm to 5 mm was 63% or more. For the aforementioned artificial blood vessels with an inner diameter of 1 mm or more and less than 2 mm, the compression recovery obtained by compression testing is 41% or more. The artificial blood vessel according to claim 1 or 2.

9. The maximum load measured by the three-point bending test is 0.0055 N or greater. The artificial blood vessel according to claim 1 or 2.

10. The wall thickness of the artificial blood vessel is 0.2 mm or more and 1.2 mm or less. The artificial blood vessel according to claim 1 or 2.

11. The tubular structure has a laminated structure in which multiple nanofiber layers are stacked, The present invention further comprises sutures arranged between two adjacent nanofiber layers. The artificial blood vessel according to claim 1 or 2.

12. The hydrophilic polymer is at least one selected from the group consisting of polyvinyl alcohol, polyacrylic acid, sodium polyacrylate, polyacrylamide, polyvinylpyrrolidone, dextran, carboxymethylcellulose, and polyethylene oxide. The artificial blood vessel according to claim 1 or 2.

13. The raw material resin for the nanofiber is at least one selected from the group consisting of polycaprolactone, polylactic acid, polyglycolic acid, polylactic acid-glycolic acid, polyurethane, polyhydroxybutyric acid, and silk. The artificial blood vessel according to claim 1 or 2.

14. A tubular structure made of nanofibers, A method for manufacturing an artificial blood vessel, comprising: a coating of a hydrophilic polymer formed on the surface of the nanofibers constituting at least the inner circumferential surface of the tubular structure; The inner diameter of the artificial blood vessel is 1 mm or more and 5 mm or less. A tubular structure forming step for forming the aforementioned tubular structure, The process includes a coating forming step of passing a solution containing a hydrophilic polymer through the lumen of the tubular structure to form the coating, The tubular structure formation step is as follows: The first step involves producing a nanofiber sheet using electrospinning, A second step involves cutting the aforementioned nanofiber sheet to produce a long nanofiber ribbon, The third step involves winding the nanofiber ribbon around a core rod at a predetermined angle and then removing the core rod to form the tubular structure. A method for manufacturing artificial blood vessels, characterized by the following:

15. The third step is, Step A involves winding the first nanofiber ribbon around the core rod at a predetermined angle, from one side of the core rod to the other. The process includes step B of winding the second nanofiber ribbon around the core rod, which is wound with the first nanofiber ribbon, at a predetermined angle, from the other side of the core rod toward the one side of the core rod. The process involves repeating steps A and B to form the tubular structure. A method for producing an artificial blood vessel according to claim 14.

16. The third step includes forming a nanofiber sheet on the surface of the core rod by electrospinning before winding the nanofiber ribbon onto the core rod. A method for manufacturing an artificial blood vessel according to claim 14 or 15.

17. The thickness of the nanofiber ribbon is 5 μm or more and 100 μm or less. A method for manufacturing an artificial blood vessel according to claim 14 or 15.

18. The width of the nanofiber ribbon is 50 mm or less. The length of the nanofiber ribbon is 30 cm or more. A method for manufacturing an artificial blood vessel according to claim 14 or 15.

Images