Wound dressing material

Abstract

The present invention relates to a wound dressing material including a hydrogel nonwoven fabric having gelatin as a main component, wherein the hydrogel nonwoven fabric has a thickness retention rate of 86% or higher after six hours of collagenase treatment. The hydrogel nonwoven fabric preferably has a compressive strength retention rate of 70% or higher after six hours of collagenase treatment. Additionally, the hydrogel nonwoven fabric preferably does not include a cell growth factor. Provided thereby is a wound dressing material having improved early cell invasion and vascular invasion into the wound dressing material during use.

Description

Technical Field

[0001] The present invention relates to a wound dressing used for repairing wounds such as tissue defects and injuries of the skin and the like.

Background Art

[0002] In wound dressings used for repairing wounds such as tissue defects and injuries of the skin and the like, a hydrogel-forming component capable of providing a moist environment is used on the side in contact with the wound site. As such a wound dressing, a sponge using a hydrogel-forming component such as collagen is widely used. Further, Patent Document 1 proposes a wound dressing including a hydrogel fiber layer containing organic fibers capable of forming a hydrogel.

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0004] However, for wound healing, after treating the wound site with a wound dressing, early cell invasion and blood vessel invasion into the wound dressing are required. Conventional wound dressings such as collagen sponges and the wound dressing described in Patent Document 1 are required to improve early cell invasion and blood vessel invasion into the interior.

[0005] In order to solve the above conventional problems, the present invention provides a wound dressing in which early cell invasion and blood vessel invasion into the wound dressing during use are improved.

Means for Solving the Problems

[0006] The present invention relates to a wound dressing material containing a hydrogel nonwoven fabric mainly composed of gelatin, wherein the hydrogel nonwoven fabric has a thickness retention rate of 86% or more after being treated with collagenase for 6 hours.

Effect of the Invention

[0007] By using the wound dressing material of the present invention, early cell invasion and blood vessel invasion into the wound dressing material can be improved.

Brief Description of the Drawings

[0008]

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Mode for Carrying Out the Invention

[0009] In order to solve the above-described problems, the inventors of the present invention conducted extensive studies. As a result, by using a hydrogel nonwoven fabric mainly composed of gelatin and having a thickness retention rate of 86% or more after 6 hours of treatment with collagenase, it was found that early cell invasion and blood vessel invasion (angiogenesis) into the interior of the wound dressing material during use were improved. In particular, by using a hydrogel nonwoven fabric mainly composed of gelatin, it is possible to promote blood vessel invasion into the interior of the wound dressing material, that is, angiogenesis, even without using a cell growth factor. Angiogenesis at the wound site is induced, and it can also be applied to diabetic patients and malignant tumor patients. In addition, since the thickness retention rate after 6 hours of treatment with collagenase is 86% or more, when using this wound dressing material, for example, in the early stage of 1 to 2 weeks after transplantation of the wound dressing material, a communication hole structure that enables invasion of cells and blood vessels into the interior of the wound dressing material is maintained, and early angiogenesis can be induced.

[0010] In one or more embodiments of the present invention, the wound dressing material includes a hydrogel nonwoven fabric mainly composed of gelatin. In one or more embodiments of the present invention, "mainly composed of gelatin" means containing 90% by mass or more of gelatin. The hydrogel nonwoven fabric may contain 95% by mass or more of gelatin, or may be substantially composed of 100% by mass of gelatin. In addition to gelatin, the hydrogel nonwoven fabric may contain other components in an amount of 10% by mass or less, or 5% by mass or less, as needed. The other components may be other biocompatible polymers, crosslinking agents, drugs, plasticizers, other additives, etc.

[0011] The type and site of the animal from which the collagen serving as the raw material of the gelatin is derived are not particularly limited. The collagen may be derived from, for example, vertebrates or fish. In addition, collagen derived from various organs and tissues such as dermis, ligament, tendon, bone, and cartilage can be appropriately used. Also, the method for preparing gelatin from collagen is not particularly limited, and examples include acid treatment, alkali treatment, and enzyme treatment. The molecular weight of the gelatin is not particularly limited, and those with various molecular weights can be appropriately selected and used. Further, one type of gelatin may be used, or two or more types may be used in combination.

[0012] The gelatin is not particularly limited, but preferably has appropriate flexibility and hardness, and from the viewpoint of enhancing the handleability of the hydrogel nonwoven fabric, the jelly strength is preferably 100 g or more and 400 g or less, more preferably 150 g or more and 360 g or less. In one or more embodiments of the present invention, the jelly strength is measured in accordance with JIS K 6503:2001. The gelatin may be a commercially available product.

[0013] The other biocompatible polymers are not particularly limited, and for example, natural polymers or synthetic polymers can be used. Examples of natural polymers include proteins and polysaccharides. Examples of proteins include collagen, fibronectin, fibrinogen, laminin, fibrin, etc. Examples of polysaccharides include natural polymers such as chitosan, calcium alginate, heparan sulfate, chondroitin sulfate, hyaluronic acid, heparin, starch, gellan gum, agarose, guar gum, xanthan gum, carrageenan, pectin, locust bean gum, tamarind gum, diutan gum, etc., and derivatives of natural polymers such as carboxymethyl cellulose may also be used. Examples of synthetic polymers include non-absorbable synthetic polymers such as polyethylene glycol, polypropylene glycol, polyethylene terephthalate, polyvinyl alcohol, thermoplastic elastomer, polypropylene, polyethylene, polystyrene, polymethyl methacrylate, polycarbonate, polydimethylsiloxane, cycloolefin polymer, amorphous fluororesin, etc., and bioabsorbable polymers such as polylactic acid, polyglycolic acid, polycaprolactone, polydioxanone, etc. The above-mentioned other biocompatible polymers may be used alone or in combination of two or more.

[0014] The hydrogel nonwoven fabric has a thickness retention rate after 6-hour treatment with collagenase (hereinafter also referred to as the initial thickness retention rate) of 86% or more. The initial thickness retention rate of the hydrogel nonwoven fabric is preferably 88% or more, and more preferably 90% or more. Thereby, a communication pore structure that enables the invasion of cells and blood vessels into the wound dressing material is maintained at an early stage of 1 to 2 weeks after treatment with the wound dressing material, and early angiogenesis can be induced.

[0015] The initial thickness retention rate of the hydrogel nonwoven fabric is calculated as follows based on the thickness (Ha) before treatment with collagenase and the thickness (Hb) after 6-hour treatment with collagenase in the swollen hydrogel nonwoven fabric. The treatment with collagenase can be carried out using 2 mL of a PBS(+) solution containing 1.25 μg / mL of collagenase D per 1 mg of the hydrogel nonwoven fabric. Thickness retention rate (%) = Hb / Ha × 100

[0016] In one or more embodiments of the present invention, "swelling" means swelling to a saturated state with one or more liquids selected from the group consisting of water or a buffer solution (such as a phosphate buffer solution). For example, the hydrogel nonwoven fabric can be swollen by immersing it in one or more liquids selected from the group consisting of water or a buffer solution for about 10 minutes or more.

[0017] The hydrogel nonwoven fabric preferably has a compressive strength retention rate (hereinafter also referred to as the initial compressive strength retention rate) of 70% or more, more preferably 80% or more, and even more preferably 90% or more after 6-hour treatment with collagenase. Thereby, in the wound site, that is, the transplantation site, even when compressed by the surrounding tissue, a communication pore structure that enables the invasion of cells and blood vessels into the wound dressing material is likely to be maintained in the early stage of transplantation from 1 to 2 weeks after transplantation, and early angiogenesis is likely to be induced. In one or more embodiments of the present invention, "compressive strength" means the stress at 70% strain in the swollen hydrogel nonwoven fabric.

[0018] The initial compressive strength retention rate of the hydrogel nonwoven fabric is calculated as follows based on the stress (Fa) at 70% strain before treatment with collagenase and the stress (Fb) at 70% strain after 6-hour treatment with collagenase in the swollen hydrogel nonwoven fabric. The treatment with collagenase can be carried out using 2 mL of a PBS(+) solution containing 1.25 μg / mL of collagenase D per 1 mg of the hydrogel nonwoven fabric. Compressive strength retention rate (%) = Fb / Fa × 100

[0019] The compressive strength of the hydrogel nonwoven fabric is not particularly limited, but from the perspective of enhancing the early invasiveness of cells and blood vessels into the hydrogel nonwoven fabric, it is preferably 5000 Pa or more, more preferably 10000 Pa or more, and even more preferably 15000 Pa or more. Also, from the perspective of ease of handling during transplantation and the degradability of the hydrogel nonwoven fabric in the later stage, the compressive strength of the hydrogel nonwoven fabric is preferably 38000 Pa or less, more preferably 36000 Pa or less, and even more preferably 31000 Pa or less.

[0020] The fibers constituting the hydrogel nonwoven fabric are not particularly limited, but the average fiber diameter in the swollen state is preferably 10 μm or more and 200 μm or less, more preferably 20 μm or more and 150 μm or less, even more preferably 30 μm or more and 100 μm or less, and particularly preferably 40 μm or more and 80 μm or less. When the average fiber diameter of the fibers is within the above range, when transplanted to the wound site, cells and blood vessels are likely to invade the inside of the hydrogel nonwoven fabric at an early stage of transplantation. In one or more embodiments of the present invention, the "average fiber diameter" means the average value of the diameters of 50 fibers arbitrarily selected from the hydrogel nonwoven fabric in the swollen state.

[0021] It is preferable that the fiber intersections of the fibers constituting the hydrogel nonwoven fabric are partially welded. This partial welding is not particularly limited, but for example, as described later, it can be achieved by depositing fibers in a not completely solidified state that are blown away by a pressure fluid during the production of the hydrogel nonwoven fabric. Due to this partial welding, the hydrogel nonwoven fabric has a bridge structure, is easy to be formed into a desired shape, and has high forming stability. Also, due to the partial welding of the fiber intersections, the hydrogel nonwoven fabric does not sag even after swelling. In the hydrogel nonwoven fabric, some of the fiber intersections may be welded, or all of the fiber intersections may be welded.

[0022] The thickness of the hydrogel nonwoven fabric is not particularly limited and can be appropriately determined according to the site to be applied and the like. However, from the viewpoints of handleability and enhancing the early invasiveness of cells and blood vessels into the hydrogel nonwoven fabric, the thickness in the swollen state is preferably 0.1 mm or more, more preferably 0.2 mm or more, still more preferably 0.3 mm or more, and particularly preferably 0.4 mm or more. Also, from the viewpoints of ease of handling during transplantation and the degradability of the hydrogel nonwoven fabric in the later stage, the thickness of the hydrogel nonwoven fabric in the swollen state is preferably 5 mm or less, more preferably 3 mm or less, still more preferably 2 mm or less, and even more preferably 1 mm or less. The thickness of the hydrogel nonwoven fabric is not particularly limited, but more specifically, the thickness in the swollen state is preferably 0.1 mm or more and 5 mm or less, more preferably 0.2 mm or more and 3 mm or less, still more preferably 0.3 mm or more and 2 mm or less, even more preferably 0.4 mm or more and 2 mm or less, and even more preferably 0.4 mm or more and 1 mm or less.

[0023] The basis weight of the hydrogel nonwoven fabric is not particularly limited and can be appropriately determined according to the site to be applied and the like. For example, from the viewpoints of handleability and enhancing the early invasiveness of cells and blood vessels into the hydrogel nonwoven fabric, the basis weight is preferably 40 g / m 2 or more, more preferably 50 g / m 2 or more, and still more preferably 60 g / m 2 or more. Also, from the viewpoints of handleability and the degradability of the hydrogel nonwoven fabric in the later stage, the basis weight of the hydrogel nonwoven fabric is preferably 500 g / m 2 or less, more preferably 400 g / m 2 or less, still more preferably 350 g / m 2 or less, and even more preferably 300 g / m 2 or less. The basis weight of the hydrogel nonwoven fabric is not particularly limited, but more specifically, it is preferably 40 g / m 2 or more and 500 g / m 2 or less, more preferably 50 g / m2 400 g / m² or less is more preferable, and 2 60 g / m² or more is even more preferable. In one or more embodiments of the present invention, the basis weight of the hydrogel nonwoven fabric is measured in accordance with JIS L 1913:2010. 2 350 g / m² or less is even more preferable. 2 In one or more embodiments of the present invention, the basis weight of the hydrogel nonwoven fabric is measured in accordance with JIS L 1913:2010.

[0024] The pore size of the hydrogel nonwoven fabric is not particularly limited and can be appropriately determined according to the application site and the like. However, from the viewpoint of enhancing the early invasiveness of cells and blood vessels into the hydrogel nonwoven fabric, the pore size in the swollen state is preferably 30 μm or more, more preferably 60 μm or more, and even more preferably 100 μm or more. Also, from the viewpoint of keeping the number of fiber intersections within an appropriate range and maintaining the strength of the hydrogel, the pore size in the swollen state is preferably 700 μm or less, more preferably 600 μm or less, and even more preferably 500 μm or less. The pore size of the hydrogel nonwoven fabric is not particularly limited, but more specifically, the pore size in the swollen state is preferably 30 μm or more and 700 μm or less, more preferably 60 μm or more and 600 μm or less, and even more preferably 100 μm or more and 500 μm or less. In one or more embodiments of the present invention, the pore size of the gelatin nonwoven fabric can be calculated by the following formula (1) based on Wrotnowski's assumption.

[0025]

Equation

[0026] As described above, the hydrogel nonwoven fabric can induce angiogenesis without containing a cell growth factor by at least setting the initial thickness retention rate within a predetermined range, preferably, in addition to the initial thickness retention rate, setting the initial compression strength retention rate within a predetermined range, and more preferably, setting the fiber diameter, pore diameter, and basis weight within a predetermined range in addition to the initial thickness retention rate and the initial compression strength retention rate. Since the use of cell growth factors is contraindicated for diabetic patients and malignant tumor patients, from the viewpoint of applicability to diabetic patients and malignant tumor patients, it is preferable that the hydrogel nonwoven fabric and the wound dressing do not contain substances such as cell growth factors (also referred to as cell growth factors). Examples of cell growth factors include basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF).

[0027] The hydrogel nonwoven fabric may be coated with a cell adhesion factor, cell induction factor, cell growth factor, a substance that provides nutrition and energy to cells, a substance that suppresses or enhances the function of cells, etc., as needed, or may be immersed in a solution containing the substance to penetrate into the interior of the fibers. The cell adhesion factor is not particularly limited, and examples thereof include fibronectin. The substance that provides nutrition and energy to cells is not particularly limited, and examples thereof include ATP, pyruvic acid, and glutamine.

[0028] The hydrogel nonwoven fabric is not particularly limited. From the perspective of suppressing the generation of contaminants and preventing product contamination, a spinning solution containing gelatin is extruded from the nozzle discharge port into the air, and a pressure fluid is ejected forward from a fluid ejection port located behind the nozzle discharge port and in a non-contact state with the nozzle discharge port. The extruded spinning solution is caused to form fibers by being accompanied by the pressure fluid, and the obtained fibers are accumulated to form a nonwoven fabric. When the fibers are accumulated (deposited) after spinning, the fibers are laminated in a state containing moisture, so the fibers are integrated by welding to each other or entangling with each other. By changing the collection distance when depositing the fibers, the nonwoven fabric density can be easily changed. The collection distance is preferably, for example, 10 cm or more and 200 cm or less, more preferably 20 cm or more and 180 cm or less, and even more preferably 30 cm or more and 150 cm or less.

[0029] Figure 11 is a schematic explanatory view of a manufacturing apparatus for a hydrogel nonwoven fabric. In the nonwoven fabric manufacturing apparatus 10, a spinning solution 2 containing gelatin placed in a heating tank 1 is extruded from a nozzle discharge port 3 into the air. A predetermined pressure is applied to the heating tank 1 by a compressor 4. 12 is a heat-insulating container. Also, a pressure fluid 7 is ejected forward from a fluid ejection port 5 located behind the nozzle discharge port 3 and in a non-contact state with the nozzle discharge port 3. The fluid ejection port 5 is supplied with a pressure fluid (for example, compressed air) from a compressor 6. The distance between the fluid ejection port 5 and the nozzle discharge port 3 is preferably 5 mm or more and 30 mm or less, and more preferably 5 mm or more and 15 mm or less. The extruded spinning solution becomes gelatin fibers 8 by being accompanied by the pressure fluid 7 and is deposited as a gelatin nonwoven fabric 9 on a winding roll 11. At this time, the deposited fibers contain moisture and are not completely solidified, so the fibers in contact with each other at at least a part of the fiber intersections are welded to each other. Note that other collection means such as a net may be used instead of the winding roll.

[0030] First, gelatin alone or, if necessary, gelatin and other biocompatible polymers that can be used as the above-described other components are dissolved in a solvent, preferably water, to prepare a spinning solution. The dissolution temperature (the temperature of the solvent such as water) is preferably 20°C or higher and 90°C or lower, and more preferably 40°C or higher and 90°C or lower. If necessary, after dissolving gelatin in a solvent such as water, filtration may be performed to remove foreign substances, dust, etc. Further, if necessary, subsequent depressurization or vacuum degassing may be performed to remove dissolved air. From the viewpoint of efficiently removing gas (bubbles), the degree of vacuum during depressurization degassing is preferably 5 kPa or higher and 30 kPa or lower. Since gelatin is water-soluble, it can be spun in the form of an aqueous solution as a spinning solution, and the safety for living bodies is increased. As water, for example, pure water, distilled water, ultrapure water, etc. can be appropriately used. In addition, when using other biocompatible water-soluble polymers as other components, a spinning solution can be prepared by dissolving them in water simultaneously with gelatin.

[0031] The temperature of the spinning solution is preferably 20°C or higher and 90°C or lower, and more preferably 40°C or higher and 90°C or lower. Within the above range, gelatin can maintain a stable sol state. Also, the gelatin concentration of the gelatin aqueous solution is preferably 30% by mass or higher and 55% by mass or lower when the gelatin aqueous solution is 100% by mass. A more preferable concentration is 35% by mass or higher and 50% by mass or lower. Within the above concentration range, a stable sol state can be maintained. The viscosity of the gelatin aqueous solution (spinning solution) is preferably 500 mPa·s or higher and 3000 mPa·s or lower. If the viscosity of the gelatin aqueous solution is within the above range, stable spinning can be performed.

[0032] The spinning solution is discharged from the nozzle of a spinning machine, a pressure fluid is supplied from around the nozzle, the discharged gelatin aqueous solution is made to form fibers while being accompanied by the pressure fluid, and the obtained gelatin fibers are accumulated to form a gelatin non-woven fabric (hydrogel non-woven fabric). The discharge pressure of the nozzle is not particularly limited, and for example, it may be 0.1 MPa or higher and 1 MPa or lower.

[0033] The temperature of the pressure fluid is preferably 20°C or higher and 120°C or lower, more preferably 80°C or higher and 120°C or lower. Although it also depends on the flow rate of the pressure fluid and the temperature of the surrounding atmosphere, stable spinning can be achieved within the above temperature range. It is preferable to use air as the pressure fluid, and the pressure is preferably 0.1 MPa or higher and 1 MPa or lower. Within the above range, the spinning solution extruded into the air from the nozzle outlet can be blown away to be fiberized.

[0034] The hydrogel nonwoven fabric is preferably crosslinked. Thereby, the morphological stability and water resistance can be enhanced. The crosslinking may be chemical crosslinking using a compound such as a crosslinking agent, but from the viewpoint of biocompatibility, crosslinking using a crosslinking agent having biocompatibility and / or crosslinking without using a crosslinking agent is preferable. Examples of crosslinking without using a crosslinking agent include thermal crosslinking, radiation crosslinking such as electron beam and γ-ray, and ultraviolet crosslinking. In the case of radiation irradiation such as electron beam or γ-ray, sterilization and crosslinking can be performed simultaneously. From the viewpoint of easily obtaining a desired crosslinking effect simply, thermal crosslinking is preferable, and thermal dehydration crosslinking is more preferable. The thermal crosslinking may be performed, for example, at 100°C or higher and 180°C or lower, or may be performed at 100°C or higher and 160°C or lower. The crosslinking time may be, for example, 24 hours or longer and 96 hours or shorter. The thermal dehydration crosslinking may be performed, for example, at 100°C or higher and 180°C or lower for 24 hours or longer and 96 hours or shorter, or may be performed at 100°C or higher and 160°C or lower for 24 hours or longer and 96 hours or shorter. Further, the thermal dehydration crosslinking may be performed, for example, under a vacuum of 1 kPa or lower. Before crosslinking, it may be dried. The drying is not particularly limited, but can be performed, for example, by air drying at room temperature or freeze drying.

[0035] Within the above range, by adjusting the discharge amount of the spinning solution, nozzle diameter (inner diameter), nozzle discharge pressure, pressure of the pressure fluid, temperature of the pressure fluid, distance between the fluid injection port and the nozzle discharge port, collection distance, crosslinking conditions, etc., a hydrogel nonwoven fabric having a desired initial thickness retention rate, initial compression strength retention rate, thickness, basis weight, fiber diameter, pore diameter, etc. can be obtained. As an example, by increasing the nozzle discharge pressure to thicken the fiber diameter, the initial thickness retention rate and the initial compression strength retention rate can be increased. As an example, by shortening the distance between the fluid injection port and the nozzle discharge port to increase the fiber intersections, the initial thickness retention rate and the initial compression strength retention rate can be increased.

[0036] The hydrogel nonwoven fabric may be cut into a predetermined shape and size as needed. The hydrogel nonwoven fabric may be sterilized by ethylene oxide gas sterilization, steam (autoclave), electron beam irradiation, radiation irradiation such as γ-rays, etc., and can also be sterilized by ethanol treatment, etc. In the case of radiation irradiation such as electron beam and γ-rays, crosslinking can be carried out simultaneously with sterilization.

[0037] In one or more embodiments of the present invention, the wound dressing is used such that one surface of the hydrogel nonwoven fabric is in contact with the wound site. That is, in the wound dressing, one surface of the hydrogel nonwoven fabric becomes the wound surface side. The wound dressing may include a protective film in addition to the hydrogel nonwoven fabric. The protective film is preferably made of a waterproof material. Thereby, it is possible to prevent moisture from the outside other than the wound site from entering the hydrogel nonwoven fabric. The protective film may adhere to the hydrogel nonwoven fabric by having self-adhesiveness, or may adhere to the hydrogel nonwoven fabric via an adhesive layer. The protective film can be disposed on one or both surfaces of the hydrogel nonwoven fabric. When using the wound dressing, one protective film may be peeled off and transplanted and used such that one surface of the hydrogel nonwoven fabric is in contact with the tissue wound site.

[0038] The manufacturing process of the hydrogel nonwoven fabric and the wound dressing is preferably carried out aseptically, for example, in a clean bench or a clean room. It is possible to prevent the hydrogel nonwoven fabric and the wound dressing from being contaminated by the propagation of miscellaneous bacteria during the operation. As for the manufacturing equipment to be used, it is preferable to use those that have been sterilized by, for example, an autoclave, radiation irradiation such as electron beams or γ-rays, etc.

[0039] The wound dressing may be applied to the wound site on the skin surface or the wound site in the subcutaneous tissue.

Examples

[0040] Hereinafter, one or more embodiments of the present invention will be described more specifically using examples. Note that the present invention is not limited to the following examples. In this specification, unless otherwise specified, the operations are carried out at room temperature (20 ± 5°C).

[0041] The measurement and evaluation methods are as follows. <Thickness and thickness retention rate> In the swollen hydrogel nonwoven fabric, the thickness (Ha) before treatment with collagenase and the thickness (Hb) after treatment with collagenase were measured with a creep meter manufactured by Yamaden Co., Ltd. Also, the thickness retention rate was calculated as follows. Thickness retention rate (%) = Hb / Ha × 100 <Compressive strength> In the swollen hydrogel nonwoven fabric, the stress (Fa) at 70% strain before treatment with collagenase and the stress (Fb) at 70% strain after treatment with collagenase were measured with a creep meter manufactured by Yamaden Co., Ltd. and taken as the compressive strength. Also, the compressive strength retention rate was calculated as follows. Compressive strength retention rate (%) = Fb / Fa × 100 <Treatment with collagenase> (1) Collagenase D (Sigma-Aldrich) was dissolved in D-PBS(+) (Dulbecco's phosphate-buffered saline (D-PBS(-)(1x) from Nacalai Tesque, to which calcium chloride (100 μg / mL) and magnesium chloride (46.8 μg / mL) from Fujifilm Wako Pure Chemical Corporation were added, resulting in a collagenase D / PBS(+) solution with a concentration of 1.25 μg / mL. (2) The swollen hydrogel nonwoven fabric (dry mass of approximately 1 mg) was immersed in 2 mL of the collagenase D solution and incubated at 37 °C for 6 hours. <Average fiber diameter> The swollen hydrogel nonwoven fabric was observed with a microscope (Keyence, BZ-X700), and the fiber diameters of 50 arbitrarily selected fibers were measured respectively, and their average was calculated to obtain the average fiber diameter in the swollen state. <Areal density (mass per unit area)> The areal density of the gelatin nonwoven fabric was measured according to JIS L 1913:2010. <Pore size> The pore size of the gelatin nonwoven fabric was calculated using the following calculation formula 1 based on Wrotnowski's assumption.

Equation

[0042] (Example 1) Gelatin from Nitta Gelatin Inc. (jelly strength 262 g, raw material: alkali-treated bovine bone) was used, with a mass ratio of gelatin:water = 3:5 (gelatin concentration 37.5 mass%), and it was dissolved at a temperature of 60 °C. The viscosity of the gelatin aqueous solution at 60 °C was 960 - 970 mPa·s. This gelatin aqueous solution was used as a spinning solution, and a gelatin nonwoven fabric was produced using the manufacturing apparatus shown in Figure 11. The temperature of the spinning solution was 60 °C, the nozzle diameter (inner diameter) was 250 μm, the discharge pressure was 0.2 MPa, the nozzle height was 5 mm, the air pressure was 0.375 MPa, the air temperature was 100 °C, the distance between the fluid injection port and the nozzle discharge port was 5 mm, and the collection distance was 100 cm. The gelatin nonwoven fabric was air-dried at room temperature overnight and then heat-dehydrated and crosslinked. The crosslinking conditions were a temperature of 140 °C for 48 hours. The areal density of the obtained gelatin nonwoven fabric was approximately 65 g / m2 It was. Next, a dry gelatin nonwoven fabric was punched into a disk shape with a diameter of about 5 mm (dry mass of about 1 mg) and allowed to stand in Dulbecco's phosphate-buffered saline (D-PBS(-)(1x), Nacalai Tesque) at room temperature for 10 minutes to swell. The diameter of the swollen hydrogel nonwoven fabric was about 7 mm.

[0043] (Example 2) A gelatin nonwoven fabric was produced in the same manner as in Example 1 except that the collection distance was set to 50 cm. The gelatin nonwoven fabric was air-dried overnight at room temperature and then heat-dehydrated and crosslinked. The crosslinking conditions were a temperature of 140 °C for 48 hours. The basis weight of the obtained gelatin nonwoven fabric was about 100 g / m 2 It was. Next, a dry gelatin nonwoven fabric was punched into a disk shape with a diameter of about 4 mm (dry mass of about 1 mg) and allowed to stand in Dulbecco's phosphate-buffered saline (D-PBS(-)(1x), Nacalai Tesque) at room temperature for 10 minutes to swell. The diameter of the swollen hydrogel nonwoven fabric was about 6 mm.

[0044] (Example 3) (Production of hydrogel nonwoven fabric) A gelatin nonwoven fabric was produced in the same manner as in Example 2 except that the discharge amount of the spinning solution was increased. The gelatin nonwoven fabric was air-dried overnight at room temperature and then heat-dehydrated and crosslinked. The crosslinking conditions were a temperature of 140 °C for 48 hours. The basis weight of the obtained gelatin nonwoven fabric was about 150 g / m 2 It was. Next, a dry gelatin nonwoven fabric was punched into a disk shape with a diameter of about 3 mm, and 1.25 pieces (dry mass of about 1 mg) were allowed to stand in Dulbecco's phosphate-buffered saline (D-PBS(-)(1x), Nacalai Tesque) at room temperature for 10 minutes to swell. The diameter of the swollen hydrogel nonwoven fabric was about 5 mm.

[0045] (Example 4) A gelatin nonwoven fabric was produced in the same manner as in Example 2, except that the discharge pressure was set to 0.1 MPa and the air pressure was set to 0.275 MPa. The gelatin nonwoven fabric was air-dried overnight at room temperature and then heat-dehydrated and crosslinked. The crosslinking conditions were a temperature of 140 °C for 48 hours. The basis weight of the obtained gelatin nonwoven fabric was about 300 g / m 2 It was. Next, the dry gelatin nonwoven fabric was punched into a disk shape with a diameter of about 2 mm (dry mass of about 1 mg) and allowed to swell by standing in Dulbecco's phosphate-buffered saline (D-PBS(-)(1x), Nacalai Tesque) at room temperature for 10 minutes. The diameter of the swollen hydrogel nonwoven fabric was about 3 mm.

[0046] (Comparative Example 1) A gelatin nonwoven fabric was produced in the same manner as in Example 2, except that the discharge amount of the spinning solution was decreased. The gelatin nonwoven fabric was air-dried overnight at room temperature and then heat-dehydrated and crosslinked. The crosslinking conditions were a temperature of 140 °C for 48 hours. The basis weight of the obtained gelatin nonwoven fabric was about 50 g / m 2 It was. Next, the dry gelatin nonwoven fabric was punched into a disk shape with a diameter of about 6 mm (dry mass of about 1 mg) and allowed to swell by standing in Dulbecco's phosphate-buffered saline (D-PBS(-)(1x), Nacalai Tesque) at room temperature for 10 minutes. The diameter of the swollen hydrogel nonwoven fabric was about 9 mm.

[0047] (Comparative Example 2) The collagen sponge was punched into a disk shape with a diameter of about 8 mm (dry mass of about 1 mg) and allowed to swell by standing in Dulbecco's phosphate-buffered saline (D-PBS(-)(1x), Nacalai Tesque) at room temperature for 10 minutes. The diameter of the swollen collagen sponge was about 8 mm.

[0048] In Examples 1 to 4 and Comparative Examples 1 to 2, the thickness of the hydrogel nonwoven fabric or collagen sponge in the swollen state before and after treatment with collagenase and the stress (compressive strength) at 70% strain were measured as described above. For Comparative Example 2, since it dissolved before 6 hours had elapsed, the compressive strength after treatment could not be measured. In Examples 1 to 4 and Comparative Example 1, the basis weight (mass per unit area) and pore size of the gelatin nonwoven fabric were measured as described above. Also, in the swollen hydrogel nonwoven fabric, the average fiber diameter was measured as described above. These results are shown in Table 1 below.

[0049] [Table 1]

[0050] Figure 1 is a photograph (100 times magnification) of a scanning electron microscope of the dry hydrogel nonwoven fabric of Example 2. Figure 2 is a photograph (100 times magnification) of a scanning electron microscope of the collagen sponge of Comparative Example 2 in the dry state. As can be seen from Figure 1, in the hydrogel nonwoven fabric of Example 2, the intersections between the fibers are fused and it has a through-hole structure.

[0051] (Experimental Example 1) A disk-shaped hydrogel nonwoven fabric of Example 1, a hydrogel nonwoven fabric of Comparative Example 1, and a collagen sponge of Comparative Example 2 with a diameter of about 6 mm in the swollen state were transplanted subcutaneously on the back of normal mice (C57BL / 6J, 8 weeks old, male, obtained from Shimizu Experimental Materials Co., Ltd.). The disk-shaped hydrogel nonwoven fabric of Example 1, the hydrogel nonwoven fabric of Comparative Example 1, and the collagen sponge of Comparative Example 2 with a diameter of about 6 mm in the swollen state were sterilized with ethylene oxide gas in the dry state before transplantation. On the 10th day after transplantation, the grafts were recovered, and frozen sections with a thickness of 10 μm were prepared on the maximum cross-section in the thickness direction of the disk-shaped grafts. Using the frozen sections, HE staining and CD31 immunostaining were performed. In the range of 250 μm to the left and right from the center of the section, the total cell count (HE staining) and the number of CD31-positive cells (CD31 immunostaining) were measured. Also, the ratio of cells by depth and the ratio of CD31-positive cells in all cells were calculated. n was set to 3. The results are shown in Table 2 below. In Table 2 below, the depth means the distance from the surface in contact with the subcutaneous tissue. For CD31 immunostaining, a CD31 antibody (manufactured by Cell Signaling Technology, "Rabbit monoclonal antibody CST 77699") was used, and peroxidase color development method using DAB was performed.

[0052]

Table 2

[0053] Figure 3 is a photograph (40 times magnification) showing the results of HE staining of a cross-section (depth 200 - 400 μm) of the graft material on the 10th day after transplantation using the hydrogel nonwoven fabric of Example 1 as the graft material. Figure 4 is a photograph (40 times magnification) showing the results of CD31 immunostaining of a cross-section (depth 200 - 400 μm) of the graft material on the 10th day after transplantation using the hydrogel nonwoven fabric of Example 1 as the graft material. Figure 5 is a photograph (40 times magnification) showing the results of HE staining of a cross-section (depth 200 - 400 μm) of the graft material on the 10th day after transplantation using the collagen sponge of Comparative Example 2 as the graft material. Figure 6 is a photograph (40 times magnification) showing the results of CD31 immunostaining of a cross-section (depth 200 - 400 μm) of the graft material on the 10th day after transplantation using the collagen sponge of Comparative Example 2 as the graft material.

[0054] As can be seen from Table 2 and FIGS. 3 to 6 above, when using the hydrogel nonwoven fabric of Example 1 having a predetermined initial thickness retention rate, when using the collagen sponge of Comparative Example 2, and when using the hydrogel nonwoven fabric of Comparative Example 1 having a low initial thickness retention rate, the ratio of cells inside (depth 200 μm to deeper) was larger than that on the surface (depth 0 to 200 μm) of the graft material, and the ratio of CD31-positive cells in all cells was also higher. It was confirmed that Example 1 had a tendency to be superior in cell invasion and blood vessel invasion (angiogenesis) into the graft material at an early stage of transplantation compared to Comparative Example 1 and Comparative Example 2, and it is presumed that it can be suitably used for wound healing.

[0055] (Experimental Example 2) A disk-shaped hydrogel nonwoven fabric of Example 2 in a swollen state with a diameter of about 6 mm and a collagen sponge of Comparative Example 2 were transplanted subcutaneously on the back of normal mice (C57BL / 6J, 8 weeks old, male, obtained from Shimizu Experimental Materials Co., Ltd.). On the 7th and 14th days after transplantation, the graft materials were collected, and frozen sections with a thickness of 10 μm were prepared at the maximum cross-section in the thickness direction of the disk-shaped graft materials. Using the frozen sections, HE staining and CD31 immunostaining were performed, and the total number of cells (HE staining) and the number of CD31-positive cells (CD31 immunostaining) were measured in the range of 250 μm to the left and right from the center of the section. In addition, the ratio of cells by depth and the ratio of CD31-positive cells in all cells were calculated. n was set to 3. The results on the 7th day after transplantation are shown in Table 3 below, and the results on the 14th day after transplantation are shown in Table 4 below. In Tables 3 and 4 below, the depth means the distance from the surface in contact with the subcutaneous tissue. CD31 immunostaining was performed using a CD31 antibody (manufactured by Cell Signaling Technology, "Rabbit monoclonal antibody CST 77699") and the peroxidase color development method using DAB.

[0056]

Table 3

[0057]

Table 4

[0058] Figure 7 is a photograph (40x magnification) showing the results of HE staining of a cross-section (depth 400 - 600 μm) of the graft material on the 7th day of transplantation using the hydrogel nonwoven fabric of Example 2 as the graft material. Figure 8 is a photograph (40x magnification) showing the results of CD31 immunostaining of a cross-section (depth 400 - 600 μm) of the graft material on the 7th day of transplantation using the hydrogel nonwoven fabric of Example 2 as the graft material. Figure 9 is a photograph (40x magnification) showing the results of HE staining of a cross-section (depth 400 - 600 μm) of the graft material on the 7th day of transplantation using the collagen sponge of Comparative Example 2 as the graft material. Figure 10 is a photograph (40x magnification) showing the results of CD31 immunostaining of a cross-section (depth 400 - 600 μm) of the graft material on the 7th day of transplantation using the collagen sponge of Comparative Example 2 as the graft material.

[0059] As can be seen from Table 3 and Figures 7 to 10 above, when using the hydrogel nonwoven fabric of Example 2 having a predetermined thickness retention rate, compared with the case of using the collagen sponge of Comparative Example 2, on the 7th day of transplantation, the proportion of CD31-positive cells in all cells was higher. It was confirmed that Example 2 tended to be superior to Comparative Example 2 in terms of blood vessel invasion (angiogenesis) into the graft material at an early stage of transplantation, and it is presumed that it can be suitably used for wound healing.

[0060] As can be seen from Table 4 above, when using the hydrogel nonwoven fabric of Example 2 having a predetermined thickness retention rate, compared with the case of using the collagen sponge of Comparative Example 2, on the 14th day of transplantation, the proportion of cells inside (depth 200 μm and deeper) was larger than that on the surface (depth 0 - 200 μm) of the graft material, and the proportion of CD31-positive cells in all cells was also higher. It was confirmed that Example 2 tended to be superior to Comparative Example 2 in terms of cell invasion and blood vessel invasion (angiogenesis) into the graft material at an early stage of transplantation, and it is presumed that it can be suitably used for wound healing.

[0061] Although the present invention is not particularly limited, it preferably includes the following aspects. [1] A wound dressing material containing a hydrogel nonwoven fabric mainly composed of gelatin, The hydrogel nonwoven fabric is a wound dressing characterized in that the thickness retention rate after 6-hour treatment with collagenase is 86% or more. [2] The wound dressing according to [1], wherein the hydrogel nonwoven fabric has a compression strength retention rate of 70% or more after 6-hour treatment with collagenase. [3] The hydrogel nonwoven fabric has a swollen thickness of 0.1 mm or more and 5 mm or less, and a basis weight of 40 g / m 2 or more and 500 g / m 2 or less, and is the wound dressing according to [1] or [2]. [4] The wound dressing according to any one of [1] to [3], wherein the hydrogel nonwoven fabric has a swollen pore diameter of 30 μm or more and 700 μm or less. [5] The wound dressing according to any one of [1] to [4], wherein the fibers constituting the hydrogel nonwoven fabric have an average fiber diameter in a swollen state of 10 μm or more and 200 μm or less. [6] The wound dressing according to any one of [1] to [5], wherein the hydrogel nonwoven fabric is heat-dehydration crosslinked. [7] The wound dressing according to any one of [1] to [6], wherein the hydrogel nonwoven fabric does not contain a cell growth factor. [8] The wound dressing according to any one of [1] to [7], wherein a protective film is disposed on one or both surfaces of the hydrogel nonwoven fabric.

Explanation of Signs

[0062] 1 Heating tank 2 Spinning solution 3 Nozzle discharge port 4, 6 Compressor 5 Fluid injection port 7 Pressure fluid 8 Gelatin fiber 9 Hydrogel nonwoven fabric 10 Nonwoven fabric manufacturing apparatus 11 Take-up roll 12 Heat-insulating container

Claims

1. A wound dressing comprising a hydrogel nonwoven fabric mainly composed of gelatin, wherein the hydrogel nonwoven fabric has a thickness retention rate of 86% or more after being treated with collagenase for 6 hours, and the hydrogel nonwoven fabric has a pore diameter in a swollen state of 30 μm or more and 700 μm or less after being swollen by immersing the hydrogel nonwoven fabric in one or more liquids selected from the group consisting of water and buffer solution for 10 minutes or more. The wound dressing is characterized by this.

2. The wound dressing according to claim 1, wherein the hydrogel nonwoven fabric has a compressive strength retention rate of 70% or more after being treated with collagenase for 6 hours.

3. The hydrogel nonwoven fabric has a thickness in the swollen state of 0.1 mm or more and 5 mm or less, and a basis weight of 40 g / m 2 More than 500g / m 2 3. The wound dressing according to claim 1 or 2, wherein:

4. The wound dressing according to any one of claims 1 to 3, wherein the fibers constituting the hydrogel nonwoven fabric have an average fiber diameter in the swollen state of 10 μm or more and 200 μm or less.

5. The wound dressing according to any one of claims 1 to 4, wherein the hydrogel nonwoven fabric is heat-dehydration crosslinked.

6. The wound dressing according to any one of claims 1 to 5, wherein the hydrogel nonwoven fabric does not contain a cell growth factor.

7. The wound dressing according to any one of claims 1 to 6, wherein a protective film is disposed on one or both surfaces of the hydrogel nonwoven fabric.

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