Artificial esophagus

Disclosed is an artificial esophagus suitable for human use that includes a fine fibrous collagen layer and a tubular outer surface. The invention generally provides a degradable artificial structure that helps maintain an existing esophagus or form a new esophagus. The invention has many important uses including improving esophageal reconstruction techniques.

TECHNICAL FIELD
 The present invention relates to an artificial esophagus.
 BACKGROUND ART
 Although materials of biological origin as well as artificial materials
 composed of polymer materials and so forth are used as an artificial
 esophagus for esophagus reconstruction following excision of the esophagus
 due to esophageal cancer and so forth, an artificial esophagus (Japanese
 Patent Provisional Publication No. 2-109569) has been reported in which a
 collagen coating layer is formed on the outer surface of a silicone tube
 for use as an artificial esophagus that promotes regeneration of
 esophageal epithelium at the portion where the esophagus is missing to
 regenerate a new esophagus without any artificial material remaining
 following esophagus reconstruction. However, since the collagen coating
 layer in this artificial esophagus swells and softens as a result of
 coming in contact with humor, suturing is difficult, thereby requiring
 sophisticated technique by the surgeon. In the case of suturing by a
 surgeon not having sophisticated technique, there is the risk of
 hemorrhage and leakage of air from the sutured site, and in the case of
 applying as a mediastinal esophagus, the postoperative process is not very
 good and there is the risk of causing death of the patient in which the
 artificial esophagus is applied.
 DISCLOSURE OF INVENTION
 With this in mind, there has been a need for the development of an
 artificial esophagus that is able to maintain for a desired duration the
 characteristic effect of collagen of promoting regeneration of esophageal
 epithelium at the portion where the esophagus is missing to regenerate a
 new esophagus, is degraded and absorbed by the body together with
 generation of new esophagus without leaving a foreign object in the body,
 has physical properties (strength) that do not require the surgeon who
 performs suturing to have a sophisticated technique, prevents hemorrhage
 and leakage of air from the sutured site, and allows a stent to be easily
 removed following esophagus reconstruction.
 As a result of earnest research to solve the above problems, the inventor
 of the present invention found that an artificial esophagus having a fine
 fibrous collagen layer has excellent effects, thereby leading to
 completion of the present invention.
 The present invention is characterized by being an artificial esophagus
 having a fine fibrous collagen layer on the outer surface of a tube. In
 addition, the present invention is characterized by being a method for
 producing said artificial esophagus that includes a step wherein a fine
 fibrous collagen layer is formed on the outer surface of a tube followed
 by performing crosslinking treatment.
 The artificial esophagus of the present invention is an artificial
 esophagus having a non-woven fabric-like collagen layer wherein fine
 fibers composed of collagen molecules are overlapped in multiple layers on
 the outside of a tube that fulfills the role of a lumen retaining core
 member. The thickness of this fine fibrous collagen layer is preferably
 about 2-10 mm, and particularly preferably about 5 mm. In addition, the
 tube serving as the lumen retaining core member uses a tube composed of,
 for example, medical silicone sheet (thickness: preferably about 0.5-5 mm,
 and particularly preferably 1-2 mm). Since the artificial esophagus of the
 present invention is sutured to the body with a tube serving as a lumen
 retaining core member, the inner diameter of the tube used is preferably
 about 15-30 mm, and particularly preferably 20 mm, or can be different
 depending on the particular case. The length of the tube used can also be
 different depending on the particular case. Although the artificial
 esophagus of the present invention has strength that allows suturing to be
 performed easily even with this artificial esophagus alone, in cases when
 even higher levels of strength are required, it may also have a collagen
 membrane layer on at least one side (outside) of the fine fibrous collagen
 layer. This collagen membrane differs from the fine fibrous collagen layer
 in that it is a collagen membrane having an amorphous structure in which
 collagen molecules are dispersed in the form of monomers and oligomers.
 The collagen which is the raw material of the fine fibrous collagen layer
 of the artificial esophagus of the present invention may be various types
 of conventional collagen, and preferably neutral solubilized collagen,
 acidic solubilized collagen, alkaline solubilized collagen or enzyme
 solubilized collagen. Alkaline solubilized collagen and enzyme solubilized
 collagen are the result of treating insoluble collagen with base or
 enzyme, respectively (examples of which include pepsin, trypsin,
 chymotrypsin, papain and pronase). As a result of this treatment, the
 strongly antigenic telopeptide portion of the collagen molecules is
 removed resulting in decreased antigenicity. Consequently, these types of
 collagen can be used particularly preferably. There are no particular
 restrictions on the origin of these collagens, and collagens can typically
 be used that are obtained by extraction and purification from the skin,
 bone, cartilage, tendon or organs of animals such as cows, pigs, rabbits,
 sheep and kangaroos.
 In preparing the artificial esophagus of the present invention using the
 above-mentioned collagen as a raw material, a fine fibrous collagen layer
 is formed on the outer surface of a tube used as a lumen retaining core
 member of the artificial esophagus, for example, a tube composed of
 medical silicone sheet as described above.
 The fine fibrous collagen layer can be formed on the outer surface of the
 tube preferably by the method described below. To begin with, an
 approximately 1 N hydrochloric acid solution (about pH 3) of extracted and
 purified collagen as described above (preferably about 0.5-3 wt % and
 particularly preferably about 1 wt %) is prepared, and the above tube (a
 rod-shaped core member, for example, a rod of made of Teflon, may be used)
 is immersed in the collagen hydrochloric acid solution (the lumen of said
 tube is sealed by inserting a rod-shaped object). By using such a method,
 a collagen hydrochloric acid solution layer is formed at a uniform
 thickness on the outer surface of the tube. The thickness of the collagen
 hydrochloric acid solution layer is preferably about 20-100 mm, and
 particularly preferably about 50 mm. This is then frozen preferably at
 about -15.degree. C. to 0.degree. C., and particularly preferably about
 0.degree. C., for preferably about 6-24 hours, and particularly preferably
 about 12 hours. As a result of this freezing, fine fragments of ice are
 formed between the collagen molecules dispersed in the hydrochloric acid
 solution, and this causes the collagen hydrochloric acid solution to
 separate into layers. Fine fibers are then formed as a result of the
 collagen molecules rearranging. Next, the silicone tube having the above
 frozen collagen hydrochloric acid solution layer on its outer surface is
 freeze-dried in a vacuum preferably at about -15.degree. C. to 0.degree.
 C., and particularly preferably about 0.degree. C., for preferably about
 12-48 hours, and particularly preferably about 24 hours. As a result of
 this freeze-drying, in addition to the fine ice fragments between the
 collagen molecules vaporizing, the collagen hydrochloric acid solution
 layer becomes a non-woven fabric-like collagen layer in which fine fibers
 composed of collagen molecules are overlapped in multiple layers.
 Moreover, the tube having on its outer surface this fine fibrous collagen
 layer is uniformly compressed using a pressing apparatus. In the
 artificial esophagus of the present invention, the thickness of the fine
 fibrous collagen layer after compression is preferably about 2-10 mm, and
 particularly preferably about 5 mm. Thus, the compression ratio, which is
 the ratio of the thickness of the collagen layer after compression to the
 thickness before compression, is preferably about 0.05-0.3, and
 particularly preferably about 0.1.
 Next, the tube on which is formed on its outer surface the above fine
 fibrous collagen layer is subjected to crosslinking treatment. As a result
 of performing this crosslinking treatment, the artificial esophagus of the
 present invention is adjusted so as to remain in the body for a desired
 duration after application. Although examples of crosslinking methods
 include those using gamma rays, electron beam, ultraviolet rays,
 glutaraldehyde and epoxy, as well as thermal dehydration crosslinking
 using heat, it is preferable to perform thermal dehydration crosslinking
 since it allows the degree of crosslinking to be easily controlled and
 there are no problems of a crosslinking agent having an effect on the
 body. In order to perform thermal dehydration crosslinking, the tube on
 which is formed the collagen layer obtained above is heated in a vacuum
 preferably at about 105-150.degree. C., and particularly preferably
 120-150.degree. C., for preferably about 6-24 hours, and particularly
 preferably 6-12 hours. If heated at less than 105.degree. C., a sufficient
 crosslinking reaction does not occur. On the other hand, if heating
 exceeds 150.degree. C., the collagen ends up denaturing. Furthermore, in
 the case of using a core member instead of a tube, said core member should
 be removed from the tube after crosslinking, and a silicone tube should be
 inserted into said tube lumen.
 Although the artificial esophagus of the present invention characterized by
 having a fine fibrous collagen layer on the outer surface of a tube has
 strength that allows suturing to be performed easily even with this
 artificial esophagus alone, in cases when even higher levels of strength
 are required, a collagen membrane having an amorphous structure as
 described above may be additionally formed on at least one side (outside)
 of the fine fibrous collagen layer. An artificial esophagus having a fine
 fibrous collagen layer on which is formed a collagen membrane on its
 outside can be produced in the manner described below.
 A collagen hydrochloric acid solution layer of uniform thickness is formed
 and dried (the formation of this collagen solution layer and drying
 procedure is repeated several times, and preferably about 10 times) using
 an about 1 N hydrochloric acid solution (pH of about 3) of extracted and
 purified collagen prepared in the same manner as described above (collagen
 concentration is preferably about 0.5-3 wt %, and particularly preferably
 about 1 wt %) on the outer surface of a tube (a core member may also be
 used as previously described) on which is formed on its outer surface a
 fine fibrous collagen layer prepared with the above-mentioned method. The
 thickness of the collagen hydrochloric acid solution layer is preferably
 about 5-20 mm, and particularly preferably about 5-10 mm, overall.
 Finally, a layer of a collagen membrane having an amorphous structure in
 which collagen molecules are dispersed is formed on the outside of said
 fine fibrous collagen layer. Furthermore, the previously mentioned thermal
 dehydration crosslinking is performed in this state. In addition, in the
 case of using a core member, said core member and silicone tube are
 exchanged in the same manner as previously mentioned, namely after
 crosslinking being performed. In addition, in the case of an artificial
 esophagus in which a layer of collagen membrane having an amorphous
 structure is formed on both sides of said fine fibrous collagen layer, a
 rod-shaped core member is used in place of the silicone tube during
 formation of the fine fibrous collagen layer to first produce a tube
 composed only of fine fibrous collagen, after which a method is employed
 in which said tube is immersed in a similar collagen hydrochloric acid
 solution as described above to form a collagen hydrochloric acid solution
 layer of uniform thickness on the inside and outside of the fine fibrous
 collagen layer followed by drying (the formation of the collagen solution
 layer and drying procedure are repeated several times in the same manner
 as previously described). In this case as well, the thickness of the
 collagen hydrochloric acid solution layer that is formed is the same as
 the case of forming on the outside only. In addition, thermal dehydration
 crosslinking is also performed in this state. Finally, said core member is
 removed and a silicone tube is inserted into a tube of fine fibrous
 collagen having a collagen membrane with an amorphous structure on both
 sides.
 As described above, by forming a layer of collagen membrane on at least the
 outside of a fine fibrous collagen layer, in addition to such a frayed
 state as observed in the non-woven fabric-like material on the surface of
 the fine fibrous collagen layer being covered by the amorphous structure
 collagen membrane, a portion of said amorphous structure collagen
 penetrates into said fine fibrous collagen layer, thereby further
 improving the physical properties of the artificial esophagus of the
 present invention and additionally improving suturing and retention in the
 body.
 Furthermore, it is preferable to have a trace amount of b-FGF (fibroblast
 growth factor) in the fine fibrous collagen layer and/or amorphous
 structure collagen membrane layer. In addition to enhancing the speed at
 which new esophagus is regenerated, an esophagus is regenerated that has a
 tissue structure that is more normal. Furthermore, an example of a method
 for making b-FGF contain, gelatin hydrogel microspheres containing b-FGF
 dissolved in PBS (phosphate buffered saline) may be injected into said
 layers with a syringe either immediately before application in the body or
 immediately after suturing to body tissue.

BEST MODE FOR CARRYING OUT THE INVENTION
 EXAMPLES
 As a result of immersing a Teflon rod having a length of about 9 cm and
 diameter of about 22 mm in a 1 N hydrochloric acid solution containing
 about 1 wt % enzyme solubilized collagen originating in pig skin and then
 lifting out said rod, a collagen hydrochloric acid solution layer was
 formed at a thickness of about 50 mm on the surface of the Teflon rod,
 after which this was frozen at about 0.degree. C. for about 12 hours. This
 was then freeze-dried in a vacuum at about 0.degree. C. for about 24 hours
 to form a fine fibrous collagen layer from the collagen hydrochloric acid
 solution layer. The Teflon rod on which was formed on its surface a fine
 fibrous collagen layer was compressed by using a pressing apparatus so
 that the thickness of the fine fibrous collagen layer was reduced to about
 5 mm. Next, the Teflon rod having the compressed fine fibrous collagen
 layer on its surface was again immersed in the previous collagen
 hydrochloric acid solution to form a collagen hydrochloric acid solution
 layer on the outer surface of the fine fibrous collagen layer followed by
 airdrying. This set of immersing in collagen hydrochloric acid solution
 and airdrying was repeated 10 times, and a layer of collagen membrane
 having an amorphous structure was formed at a thickness of 0.5 mm on the
 outer surface of the fine fibrous collagen layer. Further, the Teflon rod
 having both collagen layers on its outer surface was heated at 105.degree.
 C. for 12 hours in a vacuum to perform thermal hydration crosslinking
 treatment on said collagen layer. The Teflon rod was then removed, and a
 silicone tube having a length of about 9 cm, inner diameter of about 20 mm
 and thickness of about 1 mm was inserted into the lumen of the tube
 composed of both collagen layers to obtain the artificial esophagus of the
 present invention.
 A 5 cm portion in the thoracic part of esophagus of a beagle dog was
 replaced with the artificial esophagus of the present invention.
 Furthermore, immediately before applying the artificial esophagus of the
 present invention, 280 mg of gelatin hydrogel microspheres containing 100
 .mu.g of b-FGF dissolved in 1 ml of PBS were injected into the collagen
 layer.
 As a result of observing said replaced site one month after surgery, normal
 esophagus tissue was confirmed to be reconstructed at said replaced site.
 In a conventional artificial esophagus, 2-3 months are required for its
 reconstruction, and moreover, the patient in which the artificial
 esophagus is applied has the risk of dying prior to reconstruction
 depending on the skill with which the suturing procedure is performed in
 the replacement surgery. In comparison, the artificial esophagus of the
 present invention was confirmed to be significantly superior in this
 respect.
 In comparison with an artificial esophagus of the prior art, since the
 artificial esophagus of the present invention has excellent physical
 properties, and particularly excellent suturing, it does not require
 sophisticated technique on the part of the surgeon performing suturing.
 Consequently, there is hardly any occurrence of leakage of air or
 hemorrhage from the sutured site caused by a lack of suturing technical
 skill on the part of the surgeon. In addition, the collagen layer does not
 immediately dissolve, but continues to retain its shape for a desired
 duration, while promoting regeneration and epithelial formation of
 esophagus tissue. Correspondingly, body cells penetrate into the collagen
 layer and grow by using the collagen layer as a foothold thereby resulting
 in esophagus regeneration and replacement of the collagen layer that is
 gradually broken down and absorbed. Finally, the artificial esophagus
 applied in the body disappears and the biological esophagus is completely
 regenerated (an esophagus is reconstructed having a normal tissue
 structure). Consequently, the tube that has completed its role as a lumen
 retaining core member can be easily removed following esophagus
 reconstruction.
 INDUSTRIAL APPLICABILITY
 The artificial esophagus of the present invention can be preferably used
 since it retains the characteristic effect of collagen of promoting
 regeneration of esophageal epithelium at the portion where the esophagus
 is missing to regenerate a new esophagus, is degraded and absorbed by the
 body together with regeneration of new esophagus without leaving a foreign
 object in the body, has adequate strength by itself to withstand suturing
 to eliminate the occurrence of hemorrhage and leakage of air from the
 sutured site caused by the skill of the suturing technique of the surgeon,
 and allows a stent to be easily removed following esophagus
 reconstruction.