Method for producing nerve regeneration-inducing tube
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- 1Patent claims Zastrzeżenia patentowe 1. A method of producing a nerve regeneration tube in which the outer surface of a tubular body knitted from a plurality of ultra-thin fibers containing a biodegradable polymer is coated by repeatedly applying a collagen solution, and then collagen is filled with the inner surface of said tubular body, whereby the collagen solution viscosity is obtained, which first applied to the outer surface of the tubular body from 2 cP to 800 cP, measured at 10 ° C with a B type viscometer, with a L3 spindle and a speed of 20 rpm, and in which the diameter of the ultra-thin fibers containing the biodegradable polymer is from 1 to 50 μm. 1. Sposób wytwarzania rurki wywołującej regenerację nerwu, w którym zewnętrzną powierzchnię korpusu rurowego dzianego z wielu ultracienkich włókien zawierających polimer biodegradowalny powleka się przez wielokrotne naniesienie roztworu kolagenu, a następnie wypełnia się kola5 genem wewnętrzną powierzchnię wspomnianego korpusu rurowego, przy czym uzyskuje się lepkość roztworu kolagenu, który nanosi się najpierw na zewnętrzną powierzchnię korpusu rurowego wynoszącą od 2 cP do 800 cP, zmierzoną w temperaturze 10°C lepkościomierzem typu B, z wrzecionem L3 i przy liczbie obrotów wynoszącej 20 obr./min, a ponadto w którym średnica ultracienkich włókien zawierających polimer biodegradowalny wynosi od 1 do 50 μm. 2. The method according to claim The process of claim 1, wherein the viscosity of the collagen solution applied first to the outer surface of the tubular body is between 5 cP and 200 cP. 2. Sposób według zastrz. 1, w którym uzyskuje się lepkość roztworu kolagenu nanoszonego najpierw na zewnętrzną powierzchnię korpusu rurowego wynoszącą od 5 cP do 200 cP. 3. The method according to claim 3. The method of claim 1 or 2, wherein the collagen solution with the viscosity of the collagen solution applied first to the outside of the tubular body is applied repeatedly. 3. Sposób według zastrz. 1 albo 2, w którym roztwór kolagenu o lepkości roztworu kolagenu nanoszonego najpierw na zewnętrzną stronę korpusu rurowego nanosi się wielokrotnie. 4. The method according to any of claims from 1 to 3, in which a high viscosity of the collagen solution applied later is obtained compared to the viscosity of the first applied solution. 4. Sposób według któregokolwiek z zastrz. od 1 do 3, w którym uzyskuje się wysoką lepkość roztworu kolagenu nanoszonego później w porównaniu z lepkością roztworu nanoszonego najpierw. 5. The method according to any of claims 1 to 3, in which a high viscosity of the collagen solution applied later in two or more stages is obtained compared to the viscosity of the first applied solution. 5. Sposób według któregokolwiek z zastrz. od 1 do 3, w którym uzyskuje się wysoką lepkość roztworu kolagenu nanoszonego później w dwóch lub większej liczbie etapów w porównaniu z lepkością roztworu nanoszonego najpierw. 6. The method according to claim 4. The method of claim 4 or 5 wherein the high viscosity of the collagen solution obtained after the first application is from 200 cP to 30,000 cP. 6. Sposób według zastrz. 4 albo 5 w którym wysoka lepkość roztworu kolagenu, uzyskiwana po pierwszym naniesieniu wynosi od 200 cP do 30 000 cP. 7. The method according to any of claims The method of any one of claims 1 to 6, wherein the biodegradable polymer is at least one polymer selected from the group consisting of poly (glycolic acid), poly (lactic acid) and lactic acid-caprolactone copolymer. 7. Sposób według któregokolwiek z zastrz. od 1 do 6, w którym polimer biodegradowalny stanowi co najmniej jeden polimer wybrany z grupy obejmującej poli(kwas glikolowy), poli(kwas mlekowy) oraz kopolimer kwas mlekowy-kaprolakton. 8. A tube causing nerve regeneration that can be made by the method of any one of claims from 1 to 7. 8. Rurka wywołująca regenerację nerwu, którą można wytworzyć sposobem według któregokolwiek z zastrz. od 1 do 7. EP 2 221 070 B1 compression at a speed of mm / min compression until the deformation factor is 50% leaving it alone for 10 min after removing the weight measuring the height of the tube EP 2 221 070 B1 ściskanie z szybkością mm/min ściskanie do chwili, aż współczynnik odkształcenia wyniesie 50% pozostawienie w spokoju przez 10 min po usunięciu obciążnika pomiar wysokości rurki EP 2 221 070 B1 [Fig. 4] [Fig. 3] measuring the length L, at which the collapse of the longitudinal structure arises, the collapse of the longitudinal structure EP 2 221 070 B1 [Fig. 4] [Fig. 3] pomiar długości L, przy której powstaje załamanie struktury podłużnej powstawanie załamania struktury podłużnej 50mm 50mm EP 2 221 070 B1 [Fig. 5] EP 2 221 070 B1 [Fig. 5] EP 2 221 070 B1 EP 2 221 070 B1 ODNOŚNIKI CYTOWANE W OPISIE REFERENCES CITED IN THE DESCRIPTION Niniejsza lista odnośników cytowanych przez zgłaszającego podana jest tylko dla wygody czytelnika. Nie stanowi ona części europejskiego dokumentu patentowego. Nawet mimo dużej staranności przy zestawianiu odnośników nie można wykluczyć błędów lub przeoczeń, i Europejski Urząd Patentowy zrzeka się wszelkiej odpowiedzialności w tym zakresie. This list of references cited by the applicant is for the reader's convenience only. It is not part of the European patent document. Even though great care is taken in compiling references, errors or omissions cannot be excluded and the European Patent Office disclaims all liability in this regard. Dokumenty patentowe cytowane w opisie • JP 5237139 A [0008] • WO 9822155 A [0008] • WO 99639O8 A [0008] • JP 2OOO325463 A [0008] • JP 2OO1O7O436 A [0008] • JP 2OO232O63O A [0008] • JP 2OO3O19196 A [0008] Patent documents cited in the description • JP 5237139 A [0008] • WO 9822155 A [0008] • WO 99639O8 A [0008] • JP 2OOO325463 A [0008] • JP 2OO1O7O436 A [0008] • JP 2OO232O63O A [0008] • JP 2OO3O19196 A [0008]
83 paragraphs in 2 sections, as filed
Technical field of the invention [0001] The present invention relates to a method for producing a nerve regeneration tube whereby a peripheral nerve cut or dissected as a result of an accident or surgical operation is reconnected by using nerve cell elongation. More specifically, the present invention relates to a method in which the tight adhesion of a tubular body comprising a biodegradable polymer that forms a nerve regenerative tube with collagen applied to the outer surface of the tubular body, thereby improving the initial strength, flexibility, etc. of the entire nerve regeneration tube .
Background Art [0002] There are many examples in which complete reconstruction of a peripheral nerve damaged by an accident etc. is impossible. There are also many clinical examples in which the peripheral nerve must be resected generally as a result of surgical operations. In peripheral nerve damage, the only means, apart from direct anastomosis, is autologous nerve transplantation. However, their result is not always satisfactory, but also the recovery of sensory perception and mobility is poor, and there is a consequential effect due to mismanagement. In addition, there are many patients who complain not only of the consequential effect, such as pain and insufficient sensory perception, but also of abnormal sensory perception within the affected area or, in particular, of pain.
[0003] Attempts to regenerate the nerve by connecting gaps in the peripheral nerve using a connecting tube made of artificial materials have been eagerly carried out since the early 1980s. However, all of the studies on connecting channels using non-absorbable synthetic artificial materials have been unsuccessful. In order to solve the above problem, it is necessary to consider the following issues, such as preventing the invasion of connective tissue from the outside during the regeneration of nerve fiber bundles, the fact that it is necessary to exchange substances inside and outside the canals or neogenesis of capillary blood vessels in the walls of the canal, that it is necessary a substance that acts as a scaffold suitable for the growth of Schwann cells and axon in the canal, and that after regeneration the material used degrades and is absorbed. Given these conditions, research was then conducted into an artificial tube connecting nerves by using biodegradable and absorbable material.
[0004] Regarding peripheral nerve regeneration, attempts have been made to extend the distance between stumps that can be regenerated using a silicone tube since the description of the silicone tube model in 1982. However, because the nutrients cannot penetrate the wall of the silicone tube, there is a problem that the nutrients are not sufficiently supplied to the nerve axon, making it impossible to form capillary blood vessels in the silicone and it is not satisfactory nerve regeneration is possible even when a silicone tube is used. In addition, even if there is a possibility of nerve regeneration, then
The problem arises that the silicone tube being a foreign substance must in any case be removed by another surgical operation, etc.
[0005] On the other hand, attempts have been made to regenerate the peripheral nerve using a tube containing a biodegradable polymer in place of a silicone tube. When a nerve regeneration tube containing a biodegradable polymer is used, the nerve regeneration tube is gradually broken down and absorbed in vivo as a result of hydrolysis or the action of enzymes after nerve regeneration, so there is no need to remove it by means of subsequent surgery .
[0006] With respect to a nerve regeneration tube containing a biodegradable polymer as such, there is disclosure, for example in Patent Document 1, of nerve regeneration aid material containing collagen fiber bundles coated with laminin and fibronectin. Patent document 2 discloses an artificial nerve tube comprising a tubular body that includes biodegradable and absorbable material and, in the light of the tubular body, a collagen body containing slots and a penetrating tubular body almost parallel to the axial line of the tubular body, the gap filled is a matrix gel containing collagen, laminin, etc. Patent Document 3 discloses an artificial nerve tube comprising a tubular body that includes biodegradable and absorbable material and laminin coated fiber bundles introduced into the tubular body lumen almost parallel to the tubular body axial line. Patent Document 4 discloses a nerve reconstruction support material having a structure in which the fibers containing the bioabsorbable material are coiled together. Patent document 5 discloses a carrier, such as a sponge, tube or coil, containing collagen. Patent Document 6 discloses a carrier consisting of a spongy fine matrix comprising a biodegradable or bioabsorbable material and a linear tissue-inducing path or a linear organ-inducing path. Patent Document 7 discloses a nerve regeneration tube containing a sponge, which comprises a biodegradable polymer material and a reinforcing material containing a biodegradable polymer having a longer degradation and absorption period than the polymer contained in said sponge, the inner side of which comprises a sponge.
[0007] Nerve regeneration tubes as such are usually made in such a way that the collagen is applied to the outer surface of the tubular body knitted with an ultra-thin fiber containing a biodegradable polymer, and then the collagen is filled into the inner surface of the tubular body. However, due to the fact that the tight adherence of collagen applied to the outer surface of the tubular body to the biodegradable polymer of the tubular body is weak, there is a problem related to strength, flexibility, etc. during its use.
(Patent Documents) [0008]
1. Japanese Patent Application (JP-A) No. 237139/93 released to the public
2. Publication of International Patent Application No. WO 98/22155
EP 2 221 070 B1
3. Publication of International Patent Application No. WO 99/63908
4. Publicly disclosed Japanese Patent Application (JP-A) No. 2000-325463
5. Publicly disclosed Japanese Patent Application (JP-A) No. 2001-70436
6. Publicly available Japanese Patent Application (JP-A) No. 2002-320630
7. Publicly available Japanese Patent Application (JP-A) No. 2003-19196
Description of the figures [0009]
Fig. 1 is a drawing illustrating a method of assessing pressure resistance.
Fig. 2 is a drawing illustrating a method for assessing shape recovery properties.
Fig. 3 is a drawing illustrating a method for assessing the collapse resistance of an elongated structure.
Fig. 4 is a drawing illustrating a method for assessing the resistance to peeling of a layer.
Fig. 5 shows a SEM (50x) image of the tubular body of an example.
Disclosure of the Invention
Problem to be solved by the invention [0010] The present invention has been developed taking into account the current state of the art as such, and the object of the present invention is to provide a method for producing a nerve regeneration tube in which a collagen solution is applied to the outer surface of a tubular body knitted from ultra-thin fiber containing a biodegradable polymer while collagen fills the inner surface of the tubular body, which tube causing nerve regeneration has excellent pressure resistance, shape recovery properties, resistance to collapse of the longitudinal structure, resistance to flaking of the layer, property to prevent invasion of external tissues and resistance to leakage.
Method to solve the problem [0011] In order to achieve the goal as such, the inventor of the present invention has carried out extensive search for a method by which the tight adherence of the tubular body of a knitted fiber fabric of biodegradable polymer with collagen applied to its outer surface will be strengthened and as a result it was found that the tube inducing nerve regeneration, with excellent pressure resistance, shape recovery properties, resistance to collapse of the longitudinal structure, resistance to flaking of the layer, properties to prevent the invasion of external tissues and resistance to leakage can be produced with high efficiency if a low viscosity (or concentration) of collagen solution is applied first applied to the outer surface of the tubular body, and then during subsequent application, a higher viscosity (or concentration) of collagen solution will be obtained than before, thus implementing the present invention.
[0012] Accordingly, the present invention provides a method of producing a nerve regeneration tube in which the outer surface of a tubular body knitted from a plurality of ultra-thin fibers containing a biodegradable polymer is coated by repeatedly applying collagen solution and then filled with collagen the inner surface of the above tubular body, characterized in that a viscosity of the collagen solution applied first to the outer surface of the tubular body is obtained from 2 cP to 800 cP, or preferably from 5 cP to 200 cP.
[0013] In a preferred embodiment of the method of the present invention, a high viscosity of the collagen solution applied later is obtained compared to the viscosity of the first applied solution, with high viscosity preferably being obtained in two or more stages. Alternatively, the collagen solution with the viscosity of the collagen solution, which is applied first to the outside of the tubular body, is applied repeatedly.
[0014] Furthermore, in a preferred embodiment of the method of the present invention, the biodegradable polymer is at least one polymer that is selected from the group consisting of poly (glycolic acid), poly (lactic acid) and lactic acid-caprolactone copolymer. The present invention also relates to a tube causing nerve regeneration characterized in that it is produced by the above method.
Advantages of the invention [0015] In the production method of the present invention, a low viscosity (or concentration) of collagen solution is obtained, which is applied first to the outer surface of the tubular body knitted from fibers of biodegradable polymer, so that it is now possible to provide a tube that causes nerve regeneration, in which the tubular body containing the biodegradable polymer uniformly and tightly adheres to collagen and which exhibits excellent properties in terms of pressure resistance, shape recovery properties, resistance to collapse of the longitudinal structure, resistance to peeling of the layer, properties of invasion of external tissues and resistance to leakage.
Best Mode for Carrying Out the Invention [0016] In the method of the present invention, the tube causing nerve regeneration is produced in such a way that the collagen solution is applied repeatedly to the outer surface of the tubular body knitted from a plurality of ultra-thin fibers containing a biodegradable polymer for coating and further filled collagen tube body lumen.
[0017] Examples of the biodegradable polymer forming the tubular body include poly (lactic acid), poly (glycolic acid), polycaprolactone, lactic acid-glycolic acid copolymer, lactic acid-caprolactone copolymer, glycolic acid-caprolactone copolymer, polydioxanone and trimethylene carboxylic acid glycolic acid. Considering easy availability and handling, the use of poly (glycolic acid), poly (lactic acid) or lactic acid caprolactone is preferred, and the use of poly (glycolic acid) is particularly preferred. Each of these succumbs to 4
The biodegradable polymers can be used alone, or two or more of them can be used by mixing.
[0018] In the present invention, the diameter of the ultra-thin fiber containing the biodegradable polymer is from 1 to 50 Pm. When the diameter of the fiber is too small, the spacing between the fibers is too dense, which can make it difficult for collagen to penetrate into the tubular body or that the flexibility of the tubular body decreases. On the other hand, when the fiber diameter is too large, the retained amount of collagen will become small, as a result of which it may happen that the nerve growth rate does not increase or that the strength of the tubular body is insufficient. More preferably, the diameter of the ultra-thin fiber is from 3 to 40 μm, and even more preferably from 6 to 30 μm.
[0019] When forming the tubular body, it is preferred that 5 to 60 of the ultra-thin fibers containing the biodegradable polymer and having the above fiber diameter are wound together and alternately knitted as warp and weft. When the number of coiled ultra-thin fibers is too low, it may happen that the strength of the tubular body will be insufficient or that it will be impossible to deposit enough collagen. On the other hand, when the number of coiled ultra-thin fibers is too high, it may happen that it will be impossible to prepare a very small diameter tubular body or that the flexibility of the tubular body will not be guaranteed. More preferably, the number of ultra-thin fibers is from 10 to 50, and even more preferably from 20 to 40.
[0020] In the event that the tubular body is formed by alternately knitting the bundles of ultra-thin fibers, then it is preferred that the mesh pore size is from about 5 to 300 Pm, and more preferably from 10 to 200 Pm. When the pore size of the mesh is too small, it may happen that cell and tissue growth is inhibited due to the reduction of capillary blood vessel invasion or due to a decrease in water permeability. When it is above about 300 μm, tissue invasion will become excessive, as a result of which cell and tissue growth may be inhibited.
[0021] However, it is preferable to decide that the inner diameter and outer diameter of the tubular body are compatible with the size of the nerve to be joined and, when taking into account the cost of production and time limitation, it is preferable to prepare a lot types of tubular bodies, the sizes of which vary. Although the size of the tubular body depends on the location of the nerve being regenerated and on the necessary strength, it is typical that the inner diameter is from 0.1 to 20 mm, the outer diameter is from 0.15 to 25 mm, the wall thickness is from 0.05 to 5 mm and the length is from 1.0 to 150 mm. If the wall thickness is too large, it may hinder the regeneration of biological tissues, while when it is too small, then the degradation and absorption of the tubular body occur too quickly, as a result of which the shape may not persist until the nerve regeneration is completed. In addition, when the inner diameter of the nerve to be connected is too large, there is a possibility that the nerve elongation cannot be properly carried out.
[0022] In the present invention, the outer surface of the tubular body is coated by repeatedly applying a collagen solution in a manner known to a specialist, while the inner surface
The (lumen) of the tubular body is filled by introducing collagen into it. Regarding collagen used for application to the outer surface of the tubular body and to fill the interior of the tubular body, collagen can be used, which is traditionally used as a scaffold for nerve regeneration. Examples thereof include type I collagen, type III collagen and type IV collagen and the like, and each of them may be used alone or a larger number may be used by mixing. Regarding collagen, purified collagen is preferably used in which the concentration of sodium chloride contained therein is 2.0% by weight or less, preferably 0.1 to 1.5% by weight on a dry weight basis. Collagen may also contain laminin, a proteoglycan containing heparan sulfate, entactin and growth factor. Examples of growth factors include EGF (epithelial growth factor), eFGF (fibroblast growth factor), NGF (nerve growth factor), PDGF (platelet-derived growth factor), IGF-1 (insulin-like growth factor) and TGF-β (transforming growth factor). Regarding the collagen solution, it is preferred that after each application as a solution in hydrochloric acid using a brush or a writing brush, the solution is completely dried, and then subsequent application, thus carrying out a larger number of application operations.
[0023] The most important feature of the method according to the present invention is that when the collagen solution is applied to the outer surface of the tubular body as the collagen solution for the first application, a solution with a low viscosity of from 2 to 800 cP, preferably from 5 to 800 200 cP. It is preferred that the frequency of application of this low viscosity solution is from one to ten times, preferably from one to five times. By initially applying a low-viscosity solution within the said range, the collagen solution penetrates well between the ultra-thin biodegradable polymer fibers forming the tubular body, as a result of which it is possible to significantly improve the adhesion of the biodegradable polymer to collagen and obtain a homogeneous impression. If a high-viscosity solution having a higher viscosity than the one above is applied first, the collagen solution cannot penetrate between the ultra-thin fibers, as a result of which the collagen acquires the layer's state after drying, after which there is a risk that the collagen will peel off the tubular body. When such a tube is used to induce nerve regeneration, the result is inhibition of blood vessel invasion into the tubular body or inhibition of nerve cell growth.
[0024] In the method of the present invention, it is preferred that the low-viscosity collagen solution is applied a few times first, so as to form a sealing of the gaps between the fibers forming the tubular body and the thin layer, and then a higher-viscosity collagen solution is applied to it 30,000 cP. This is due to the fact that when applying only a low-viscosity solution, it is necessary to repeat the application repeatedly to obtain a predetermined thickness of the thin layer, and thus the working capacity is bad. It is desirable that the frequency of application of this high viscosity solution is from one to fifty times, preferably from one to thirty times. If the frequency of application of the high viscosity solution is too high, this results in a reduction of the shape recovery properties and, for example, if the affected area is crushed after surgery with something, the resulting deformation of the tube will not be eliminated, which may block it nerve regeneration pathway. In addition, because collagen exhibits relatively large
Due to the rate of biodegradation, the benefit of this fact is small even when the application frequency increases too significantly.
[0025] In fact, it is preferable to obtain a higher viscosity of the collagen solution in a number of steps from the two or more following the first application of the low viscosity solution. For example, the viscosity of the collagen solution applied can be increased in three stages from 2 to 200 cP, from 200 to 3000 cP and from 3000 to 30,000 cP. In this case, the penetration between the ultra-thin fibers of the tubular body and the formation of a layer on the surface is carried out using the first low-viscosity solution, the adhesion to this layer is carried out using another medium-viscosity solution to seal the network, and the last high-viscosity solution sticks to this sealed collagen layer to improve strength, thanks to which it is possible to carry out effective coating with strong initial strength. In addition, small differences in the viscosity of collagen applied by the "step by step" method are obtained, thanks to which it is possible to improve the ability to conduct layer application operations or to reduce uneven application or to leave a given place without an applied layer.
[0026] It is preferred that the tubular body, which is coated or filled with collagen, is subjected to freezing, freeze-drying and cross-linking treatment to cross-link collagen. Preferably, freezing is carried out at a temperature of from -10 to -196 ° C for 3 to 48 hours. As a result of freezing, fine ice is formed between the collagen molecules and as a result phase separation occurs in the collagen solution to form a sponge. Then, the above frozen collagen solution is lyophilized under vacuum, preferably at a temperature from about -40 to -80 ° C and preferably for about 12 to 48 hours. As a result of freeze-drying, the fine ice between collagen particles volatilizes and at the same time the collagen sponge acquires a fine-grained structure. Examples of cross-linking methods include γ-crosslinking, ultraviolet cross-linking, electron-crosslinking, cross-linking by thermal dehydration, cross-linking with glutaraldehyde, cross-linking with epoxy moieties and cross-linking with water-soluble carbodiimide, among which cross-linking by dehydration is preferred thermal, in which the degree of crosslinking can be easily controlled, and even a cross-linking operation does not affect the living organism. Crosslinking by thermal dehydration is carried out under vacuum, for example at a temperature from about 105 to 150 ° C, more preferably from about 120 to 150 ° C, and even more preferably from about 140 ° C for about 6 to 24 hours, more preferably for about 6 to 12 hours, and even more preferably for about 12 hours. When the crosslinking temperature is too high, it is possible that the strength of the biodegradable and absorbable material will decrease, whereas if it is too low, there is a possibility that the crosslinking reaction will not occur sufficiently.
[0027] Since the tubular body containing the biodegradable polymer and collagen tightly adhere to each other in the nerve regeneration tube produced as mentioned above, initial strength and flexibility can be obtained that are not lower than the sum of the strengths of each. More specifically, in the tube causing nerve regeneration according to the present invention,
The deformation rate (pressure resistance) when compression is applied by applying a load of 100 N / m on the side in the diameter is not more than 15%, preferably 0.1 to 10%, and furthermore the degree return to the original shape (shape recovery property) for 50% deformation when similar compression is carried out so as to obtain 50% deformation of the tube (until the diameter of the tube reaches half) is not less than 60%. Pressure resistance is based on the assumption of load resistance for the tube causing nerve regeneration caused by the action of the medical device at the nerve connection and the therapy used after surgery, and generally the greater the thickness of the collagen layer, the greater the pressure resistance. However, if the tubular body and collagen do not adhere closely to each other and the layer separates, the pressure resistance will not be as high as expected. In addition, the property of returning to its original shape is based on the assumption of shape recovery after deformation due to the operation of the medical device at nerve connection (such as tweezing too strongly) or shock covering the affected area after surgery, and if the property of returning to its original shape is low, then tube deformation persists and the nerve growth path is inhibited.
[0028] Furthermore, the nerve regeneration tube according to the present invention has a limiting bending ratio (resistance to collapse of the longitudinal structure) of not less than 10% and also exhibits high resistance to peeling of the layer. The limiting bending factor indicates the extent to which bending is possible without causing the longitudinal structure to collapse and is an indicator relating to the moving region at nerve connection. If the limit bending factor is less than 10%, it cannot be used for cases where a curved nerve growth path is required, and even if used, the nerve is under stress and there is a risk of inhibiting nerve growth and causing the condition ignited due to compression of external tissues. The resistance to peeling of the layer means the resistance to flaking and cracking of coated collagen. The reason why collagen is coated on the entire outer surface of the tubular body is to prevent the invasion of external tissue into the nerve growth path (the property of preventing the invasion of external tissues) and to prevent leakage of the collagen sponge contained on the inner surface of the tubular body outside (leakage resistance), and in the case of if the collagen that forms in the coating peels off or breaks, there is a risk that the above properties cannot be guaranteed. In the nerve regeneration tube of the present invention, the tubular body and collagen tightly adhere to each other and there is no separate layer, so that high resistance to collapse of the longitudinal structure can be achieved, while there is no possibility that the effect is peeling and cracking as such.
[0029] In the nerve regeneration tube of the present invention, a large effect can also be expected in regulating the rate of degradation needed for bio-absorbability. In the event that the nerve regeneration tube formed from the biodegradable tubular body and collagen sponge and the collagen coating is embedded in the body, the strength of the collagen coating is lost because collagen degrades first. However, if the method of the present invention is used, the strength of the tubular body can be maintained for a longer time because the collagen degradation rate
In the gaps between the fibers of the tubular body is reduced. In addition, because the space between the fibers of the tubular body can be sealed for a long time, this can prevent the invasion of external tissues that creates the risk of inhibiting the growth of nerve cells. The reason why the rate of degradation is reduced is probably due to the fact that collagen adhering to the gaps between the fibers of the tubular body has a small contact surface with body fluids and external tissues.
Examples [0030] The effect of the tube causing the nerve regeneration produced by the method of the present invention will be presented below, although the present invention is not limited thereto. The evaluation of the nerve regeneration tube obtained in the examples was carried out according to the following methods.
Evaluation method (1) Pressure resistance [0031] A load of 100 N / m in diameter was applied on one side of the sample over a length of 5 mm, as shown in Fig. 1, under the following measurement conditions. The diameter of the diameter (L) was then measured in the load direction, after which the deformation factor was calculated according to the formula deformation factor = (L / Lo) x1OO (where Lo is the height in the direction of the load before applying the load). The sample was measured for the non-aging case as well as for the aging case using saline solution for one, two, three and four weeks.
Measuring conditions [0032] • Temperature: 2OO ° C; humidity: 65, O% • Tester: Tensilon (UTA-1t) • Testing speed: 1 mm / min • Load cell class: 5 kG • Number of samples: N = 3 (2) Shape recovery property [0033] The sample was compressed until until the deformation factor reached a value of 5O% in the diameter direction on one side of the 5mm sample, as shown in Figure 2, under the same measurement conditions as in the above (1) pressure resistance measurement. Immediately after compression, the weight was removed, and the sample left alone for 10 minutes. Then the diameter of the diameter (L1) was measured in the load direction, after which the degree of return to the original shape was calculated
According to the formula, the degree of recovery = [(Li-2 / L0) / (2 / Lo)] x1OO (wherein, Lo is the height of the diameter in the direction of the load before applying the load).
(3) Resistance to collapse of the longitudinal structure [0034] As shown in Figure 3, at 2O, 0 ° C and 65, 0% humidity, a 5O mm sample was bent by hand at a rate of about 1 mm / second and the length measured (L2 mm) when the longitudinal structure collapsed in the sample, after which the limit bending coefficient [1- (L2 / 5O)] x1OO was calculated. The number of samples measured was N = 3.
(4) Resistance to peeling of the layer [0035] As shown in Fig. 4, at 2O, O ° C and 65, O% humidity, side of sample
5 mm was cut with scissors and it was confirmed if there was a possibility of peeling and separation of the collagen layer on the outer surface of the sample. In addition, a photo of the outer surface of the sample was taken under an SEM microscope and it was confirmed if there was partial peeling or cracking of the layer. The number of samples measured was N = 3.
(5) Prevention of cell invasion and resistance to leakage [0036] At 25, 0 ° C and 6O, 0% humidity, with a collagen solution of 1, 0% by weight
the obtained method, which will be mentioned later, was filled the inner surface of the sample over a length of 5 mm. Then, every ten minutes with the unaided eye it was confirmed whether collagen filling the sample was leaking from this side of the sample and the time until leak was confirmed was recorded. If you consider the time to completely freeze the collagen solution, it is necessary that the leakage resistance is not less than 2 hours or, at least, not less than 1 hour.
Preparation of collagen solutions [0037] 392 g O, OO1 mol / L hydrochloric acid (pH 3) was introduced into the plastic bottle, followed by 8 g NMP collagen PS collagen (manufactured by Nippon Meat)
Packers, Inc.) and the mixture was vigorously stirred to dissolve, resulting in a collagen solution in which the final concentration of collagen was 2.0% by weight. This collagen solution was diluted with the above hydrochloric acid to obtain collagen solutions in which the final concentration of collagen was 0, 1, 0, 2, 0, 5, 0, 7 and 1, wt. in each sample.
Measurement of the viscosity of collagen solutions [0038] Each of the collagen solutions in which the collagen concentration was O, 1, O, 2, O, 5, O, 7, 1, O and 2, O wt. in each sample, stabilized at a temperature of 1 ° C using a thermostatic vessel in which cooling water circulated at a temperature of 1 ° C, then a B type viscometer (product name: Visco Basic plus, manufactured by FUNGILAB, used rotor: L3 spindle, number of revolutions of the measuring spindle: 2O rpm, number of tests: N = 3),
The measured values were read 3, 4 and 5 minutes after commissioning and their mean value was taken as the measured viscosity. The result is given in Table 1.
[Table 1]
<td>collagen concentration (wt.%)</td><td> 0,1</td><td> 0,2</td><td> 0,5</td><td> 0,7</td><td> 1,0</td><td> 2,0</td>
<td>viscosity (cP)</td><td> 17</td><td> 40</td><td> 925</td><td> 2580</td><td> 7542</td><td> 25367</td>
[0039] A bundle of fibers in which 28 ultra-thin fibers (diameter: about 15 μm) were wound containing poly (glycolic acid) was used as a warp and weft and alternately knitted to give a cylindrical tubular body with an internal diameter of 3 mm and a length of 50 mm.
Examples 1 to 8 and Comparative Examples 1 and 2 [0040] The above collagen solution was uniformly applied once to the outer surface of the above tubular body with a brush made of Teflon (registered trademark) and air dried and, after confirming that it had dried completely , subsequent applications were carried out successively. The application rates of the collagen solution applied were as described in the application method in Table 2 and the application was carried out sequentially, starting from the solution in which the collagen concentration was lower. After application of the collagen solutions, a 1.0 wt.% Collagen solution introduced into the lumen of the tubular body and frozen at -40 ° C. The frozen solution was freeze-dried, followed by thermal cross-linking under vacuum (no higher than 1 Pa) at 140 ° C for 24 hours to cross-link collagen molecules, and the tubular body thus obtained was used as the sample in each of Examples 1 to 8 and comparative example 1. The sample of comparative example 2 was the same as the other except that no collagen solution was applied.
Evaluation result [0041] Pressure resistance, property of returning to its original shape, resistance to collapse of the longitudinal structure, resistance to flaking of the coating, property of preventing cell invasion and resistance to leakage were evaluated for the samples of the above examples 1 to 8 and comparative examples 1 and 2. The results are shown in Table 2.
EP 2 221 070 B1 [Table 2]
<td>Comparative Example 2</td><td></td><td>>> N WHAT ABOUT</td><td>0 times</td><td></td><td>0 times</td><td></td><td> 48,5%</td><td> 55,2%</td><td> 56,8%</td><td> 68,9%</td><td> 72,5%</td><td> 81,3%</td><td> 23,0%</td><td>lack</td><td>lack</td><td>10 min</td><td> <</td><td>about about</td><td>ABOUT about</td><td>ABOUT</td><td> <3</td>
<td>Comparative Example 1</td><td></td><td>0 times</td><td>0 times</td><td></td><td>20 times</td><td></td><td> 16,2%</td><td> 35,5%</td><td> 52,5%</td><td> 64,5%</td><td> 66,6%</td><td> 42,6%</td><td> 3,6%</td><td>current</td><td>current</td><td>180 min</td><td>ABOUT</td><td> <3</td><td> <3</td><td> <3</td><td>ABOUT about</td>
<td>Example 8</td><td>Three times</td><td></td><td>1 time</td><td></td><td></td><td>10 times</td><td> 4,9%</td><td> 29,8%</td><td> 42,4%</td><td> 58,4%</td><td> 69,8%</td><td> 74,7%</td><td> 14,8%</td><td>lack</td><td>lack</td><td>190 min</td><td>about about</td><td>ABOUT</td><td>ABOUT</td><td>ABOUT</td><td>about about</td>
<td>Example 7</td><td>5 times</td><td></td><td></td><td>25 times</td><td></td><td></td><td> 5,2%</td><td> 30,9%</td><td> 44,1%</td><td> 60,7%</td><td> 72,5%</td><td> 75,6%</td><td> 14,0%</td><td>lack</td><td>lack</td><td>200 min</td><td>about</td><td>about</td><td>about</td><td>about</td><td>about about</td>
<td>Example 6</td><td></td><td>2 times</td><td>1 time</td><td></td><td>1 time</td><td></td><td> 6,5%</td><td> 25,8%</td><td> 37,0%</td><td> 56,2%</td><td> 64,3%</td><td> 85,1%</td><td> 20,4%</td><td>lack</td><td>lack</td><td>60 min</td><td>about</td><td>about about</td><td>about about</td><td>about</td><td>about</td>
<td>Example 5</td><td></td><td>2 times</td><td>1 time</td><td></td><td>5 times</td><td></td><td> 3,9%</td><td> 23,5%</td><td> 43,5%</td><td> 58,0%</td><td>χ0 er · ABOUT CD r-</td><td> 82,8%</td><td> 19,4%</td><td>lack</td><td>lack</td><td>90 min</td><td>about about</td><td>about about</td><td>about about</td><td>about</td><td>about</td>
<td>Example 4</td><td></td><td>2 times</td><td>1 time</td><td></td><td>10 times</td><td></td><td> 3,9%</td><td> 19,8%</td><td> 47,3%</td><td> 55,5%</td><td> 71,2%</td><td> 79,1%</td><td> 17,0%</td><td>lack</td><td>lack</td><td>130 min</td><td>about about</td><td>about</td><td>about about</td><td>about</td><td>about about</td>
<td>Example 3</td><td></td><td>2 times</td><td>1 time</td><td></td><td>20 times</td><td></td><td> 3,9%</td><td> 29,0%</td><td> 38,8%</td><td> 55,8%</td><td> 69,3%</td><td> 70,1%</td><td> 14,4%</td><td>lack</td><td>lack</td><td>190 min</td><td>about about</td><td>about</td><td>about</td><td>about</td><td>about about</td>
<td>Example 2</td><td></td><td>2 times</td><td>1 time</td><td></td><td>25 times</td><td></td><td> 2,7%</td><td> 29,0%</td><td> 40,0%</td><td> 52,7%</td><td> 61,8%</td><td> 68,5%</td><td> 11,0%</td><td>lack</td><td>lack</td><td>220 min</td><td>about about</td><td>about</td><td>about</td><td>about</td><td>about about</td>
<td>Example 1</td><td></td><td>2 times</td><td>1 time</td><td></td><td>17 times</td><td></td><td> 5,7%</td><td> 30,2%</td><td> 43,0%</td><td> 59,2%</td><td> 70,7%</td><td> 75,7%</td><td> 13,8%</td><td>lack</td><td>lack</td><td>180 min</td><td>about</td><td>about</td><td>about</td><td>about</td><td>about about</td>
<td rowspan="3">Sample No.</td><td>0.1 wt.</td><td>0.2 wt.</td><td>0.5 wt.</td><td>0.7 wt.</td><td>1.0 wt.</td><td>1.5 wt.</td><td>without aging</td><td>1 week</td><td>2 weeks</td><td>Three weeks</td><td>4 weeks</td><td rowspan="3">degree of return to original shape</td><td rowspan="3">limit bending factor</td><td rowspan="3">layer separation</td><td rowspan="3">peeling or cracking observed with SEM</td><td rowspan="3">time to leakage of fill liquid</td><td rowspan="2">pressure resistance</td><td rowspan="2">shape recovery property</td><td rowspan="2">resistance to collapse of the longitudinal structure</td><td rowspan="2">resistance to peeling of the layer</td><td rowspan="2">property of preventing cell invasion and resistance to leakage</td>
<td colspan="6" rowspan="2">ElUeZSOUBU qosods</td><td colspan="5" rowspan="2">ω / Ν00 ł Asiad eiueotejzs ^ po</td>
<td colspan="5">eueoo</td>
[0042] From the results presented in Table 2 it can be clearly seen that compared to traditional tubes, the nerve regeneration tubes produced by the method of the present invention show excellent pressure resistance, the property of returning to the original shape, resistance to collapse of the longitudinal structure , resistance to coating flaking, property of preventing cell invasion and resistance to leakage.
Industrial Applicability [0043] Due to the fact that the tube causing nerve regeneration produced by the method of the present invention exhibits the excellent properties listed above, it is characterized by excellent quality retention during storage or transport, handling during clinical use and stability as well as safety after surgery surgical, thanks to this, it finds significant application in medical procedures aimed at nerve regeneration.
EP 2 221 070 B1
Contents2
16 members in 7 offices
Priority claims6
| Document | Office | Kind | Date |
|---|---|---|---|
| 2007317462 | Japan | A | |
| 08856086 | European Patent Office (EPO) | A | |
| 2008072038 | Japan | W | |
| EP20080856086 | – | – | – |
| JP20070317462 | – | – | – |
| WO2008JP72038 | – | – | – |
Members16
| Document | Office | Kind | |
|---|---|---|---|
| WO2009072552A1 | World Intellectual Property Organization (WIPO) | A1 | |
| JP2009136575A | Japan | A | |
| EP2221070A1 | European Patent Office (EPO) | A1 | |
| US2010249811A1 | United States of America | A1 | |
| JP4596335B2 | Japan | B2 | |
| EP2221070A4 | European Patent Office (EPO) | A4 | |
| US8632844B2 | United States of America | B2 | |
| US2014099427A1 | United States of America | A1 | |
| US2014100591A1 | United States of America | A1 | |
| EP2799094A1 | European Patent Office (EPO) | A1 | |
| EP2221070B1 | European Patent Office (EPO) | B1 | |
| DK2221070T3 | Denmark | T3 | |
| PT2221070E | Portugal | E | |
| PL2221070T3This record | Poland | T3 | |
| US9687592B2 | United States of America | B2 | |
| EP2799094B1 | European Patent Office (EPO) | B1 |
Numbers
- Publication, DOCDB
- 2221070
- Publication, EPODOC
- PL2221070T
- Application
- 856086
- Application, DOCDB
- 08856086
- Application, EPODOC
- PL20080856086T
Titles2
- English
- METHOD FOR PRODUCING NERVE REGENERATION-INDUCING TUBE
- Polish
- Sposób wytwarzania rurki wywołującej regenerację nerwu
Classification
- CPC, 6
- A61B17/1128
- A61L31/044
- A61B2017/00526
- A61L27/34
- A61L27/58
- A61L2430/32
- IPC, 3
- A61B17 11
- A61B17 00
- A61L31 04