MXPA99004526A

MXPA99004526A - Tubular submucosal graft constructs

Info

Publication number: MXPA99004526A
Application number: MXPA/A/1999/004526A
Authority: MX
Inventors: F Badylak Stephen; C Hiles Michael; A Geddes Leslie; H Patel Umesh
Original assignee: Cook Biotech Inc; Fearnot Neal E; C Hiles Michael
Priority date: 1996-12-10
Filing date: 1999-05-14
Publication date: 2000-02-02

Abstract

An easy-to-produce and mechanically strong tube of an implantable submucosal tissue has been developed which is manufactured in any desired length, wall thickness, or diameter. The construct produced by the method of the invention may be used as grafts for arteries, veins, ureters, urethras, shunts, or in any application where a compliant, tissue-compatible tube is needed. The manufacture of the submucosal tissue prosthesis generally involves wrapping a first sheet of submucosal tissue (60) and a second sheet of submucosal tissue (70) around a mandrel (50), wherein the first end (74) and the second opposite end (76) of the second sheet of submucosal tissue (70) are sutured together with sutures (78). The submucosal tissue is compressed and dried on the mandrel (50) before removing the construct by pulling on a first end (54) and a second end (56) of a water permeable tape to unwind the tape and thus release the construct for eventual use.

Description

TUBULAR CONSTRUCTS OF SUBMUCOSAL GRAFT TECHNICAL FIELD This patent relates to tissue, biologic, implantable, graft constructions suitable for various medical applications and to the process for the production of such graft constructions. More specifically, the submucosal tissue is used to form tubular multilamellar constructions of varying diameter. Tissue graft constructions have applications such as arterial and venous grafts, replacements of the ureter and urethra, and as various conduits and shunts.

BACKGROUND OF THE INVENTION Researchers in surgical techniques have been working for many years to develop new techniques and materials for use as grafts, to replace or repair damaged or diseased tissue structures, particularly bones and connective tissues, such as ligaments and tendons, and to accelerate he REF, 29860 healed fractures. It is very common today, for example, for an orthopedic surgeon to harvest a patellar tendon of autogenous or allogenic origin for use as a replacement for a broken cruciform ligament. Surgical methods for such techniques are well known. In addition, it has become common for surgeons to use implantable prostheses, formed from plastic, metallic and / or ceramic materials for the reconstruction or replacement of physiological structures. Yet, despite its widespread use, surgically implanted prostheses, currently available, present many risks to the patient. Therefore, surgeons are in need of a high-tensile, non-immunogenic graft material that can be used for the surgical repair of bones, tendons, ligaments and other functional tissue structures. More recently researchers have been working to develop biological tissues for use as implants and for use in the repair of damaged or diseased tissues, since plastic or polymeric materials have drawbacks in these medical applications.

While plastics and polymers may have some desirable mechanical properties (for example, tensile strength), plastics have been found to become infected and in plastics applications plastics have been reported as inducing thrombogenesis. Tubular prostheses made from natural tissues have been widely used in recent years in the surgical repair and replacement of diseased or damaged blood vessels in human patients. Natural tissue prostheses fall into three general categories: autogenous, homologous and heterologous prostheses. Tissue prostheses of autogenous material are prepared from tissues taken from the patient's own body (for example, grafts of the saphenous vein). The use of such prostheses eliminates the possibility of rejection of the implanted prosthesis, but requires a more extensive surgical intervention and consumes more time, with the risks inherent to the patient. The homologous natural tissue prostheses are prepared from tissue taken from another human being, while the heterologous natural tissue prostheses are prepared from tissue from a different species. The use of umbilical cord vessels, homologous and heterologous, for example, vascular and urethral prostheses are described in the. US Patent Nos. 3,894,530; 3,974,526; and 3,988,782. In addition, autogenous vascular prostheses prepared from sheets of pericardial tissue have been described by Yoshio Sako, "Prevention of Dilation in Autogenous Venous and Pericardial Grafts in the Thoracic Aorta," Surgery, 30, pp. 864-948. 148-160 (1951) and by Robert G. Alien and Francis H. Cole, Jr. , "Modified Blalock Shunts Utilizing Pericardial Tube Grafts "Jour. Pediatr. Surg., 12 (3), pp. 287-294 (1977). The heterologous vascular prostheses prepared from leaves of porcine pericardial tissue have been described by Ornvold K. et al., "Structural Changes of Stabilized Porcine Pericardium after Experimental and Clinical Implantation", in Proc. Eur. Soc. For Artif. Organs, Vol. VI, Geneva, Switzerland (1979). The necessary characteristics of a tubular vascular prosthesis are biological compatibility, adequate resistance, resistance to infection, resistance to biological degradation, non-thrombogenicity and lack of aneurysm formation. As used in this application, the term "biological compatibility" means that the prosthesis is non-toxic in the environment contrary to its intended use, and is not rejected by the patient's physiological system (eg, it is non-antigenic). In addition, it is desirable that the prosthesis be capable of producing at an economical cost in a wide variety of lengths, diameters and shapes (eg, straight, curved, bifurcated), which is easily anastomosed to the patient's body and other tubular prostheses thereof. type or of a different type, and that shows dimensional stability in use. As described in U.S. Patent No. 4,902,508, vascular graft constructions comprising intestinal submucosal tissue have been previously described and used to replace damaged or diseased vascular tissues. The vascular graft constructions were prepared by inserting a glass rod of the appropriate diameter into the lumen or lumen of the submucosal tissue and manually suturing along the junction of the submucosal tissue. Vascular grafts of submucosal tissue are aseptically fabricated during surgery and typically take a surgeon about half an hour to prepare. Therefore, to avoid the expense of time in the preparation of the graft constructions during surgery, pre-sterilized, prefabricated grafts of different diameters are desirable. The preparation of a tubular prosthesis of the correct length and shape increases the ease of implantation and improves the functionality of the implant. For example, a tubular prosthesis that is too long for the intended application may be twisted after implantation, whereas the implantation of a prosthesis that is too short puts excessive strain on the anastomoses at its ends, thereby resulting in trauma to the prosthesis. the anastomoses. Thus, it could be highly desirable to provide a tubular prosthesis array that varies in size and that can be cut transversely to a desired length at any point between its ends, without substantially damaging the prosthesis in any other way. The present invention is directed to a tubular prosthesis comprising submucosal tissue and methods for the preparation of such a prosthesis. The submucosal tissue, prepared according to the present invention, has previously been described as a non-thrombogenic, biocompatible graft material, which improves the repair of damaged or diseased host tissue. Numerous studies have shown that the submucosae of warm-blooded vertebrates are capable of inducing host tissue proliferation, remodeling and regeneration of tissue structures after implantation in a number of in vitro microenvironments, including the lower urinary tract , body walls, tissues of the tendons, ligaments, bones, cardiovascular and central nervous system tissues. After implantation, cellular infiltration and rapid neovascularization is observed and the submucosal material is remodeled into the host replacement tissue with the specific structural and functional properties of the site. The submucosal tissue can be obtained from various tissue sources, harvested from animals bred for meat production, including, for example, pigs, cattle and sheep or other warm-blooded vertebrates. More particularly, the submucosa is isolated from a variety of tissue sources including the alimentary tracts, respiratory, intestinal, urinary or genital of warm-blooded vertebrates. In general, the submucosa is prepared from these tissue sources by delamination of the submucosa from the layers of the smooth muscle and mucosal layers. The preparation of intestinal submucosa is described and claimed in U.S. Patent No. 4,902,508, the description of which is expressly incorporated by reference herein. The submucosa of the urinary bladder and its preparation is described in U.S. Patent No. 5,554,389, the disclosure of which is expressly incorporated by reference herein. The stomach submucosa has also been obtained and characterized using similar tissue processing techniques. This is described in U.S. Patent Application No. 60 / 032,683 entitled TISSUE GRAFTING DERIVED FROM THE STOMACH SUBMUCOSA, filed December 10, 1996. In summary, the stomach submucosa is prepared from a stomach segment in a procedure similar to the preparation of intestinal submucosa. A segment of stomach tissue is first subjected to abrasion using a longitudinal rubbing movement to remove the outer layers (particularly the smooth muscle layers) and the luminal portions of the mucosal layers of the tunica. The resulting submucosal tissue has a thickness of about 100 to about 200 microns, and consists mainly (more than 98%) of extracellular matrix material, which stains eosinophilically (H & amp;; E) acellular.

BRIEF DESCRIPTION OF THE INVENTION In accordance with the present invention, an implantable tubular prosthesis comprising submucosal tissue, in the form of a tube, is prepared. The tubular construction comprises a first sheet of submucosal tissue wound in the form of a multi-layered tube of submucosal tissue, and a second sheet of submucosal tissue that is wrapped around the tube of submucosal tissue. The second sheet of submucosal tissue is superimposed on the submucosal tissue tube, so that a first edge is in contact with the submucosal tissue, and the second opposite edge is either sutured to the first edge or extends over the first edge and is sutured to the second leaf of the submucosal tissue.

The multi-layer tubular graft constructions of the present invention are formed to have fluid-tight joints, and can be shaped to engage the endogenous tissue that is to be replaced by the injured construct. In addition, according to the present invention, there is provided a process for producing a graft construction of implantable tissue formed in the shape of a tube in the shape having a longitudinally extending junction along the length of the graft, in where the union has been sealed to resist the movement of fluids from the lumen or light through the union to the outside of the tube. A method for the formation of the submucosal tubular constructions of the present invention comprises the steps of: A. superimposing a sheet of submucosal tissue around the circumference of a mandrel to form a tube of submucosal tissue having an overlapped region of multiple layers; B. the fixation of the submucosal tissue layers in the overlapping region, one to the other; C. the overlap of a second sheet of submucosal tissue over the submucosal tissue tube to form a second tube of submucosal tissue, where the junction of the second submucosal tissue tube is sealed by sutures; and D. the compression of the superimposed layers of submucosal tissue under conditions of dehydration. The present invention allows the construction of multilayer tubular graft constructions from sheets of submucosal tissue, wherein the walls of the formed tubular prosthesis do not contain perforations that provide a direct passageway from the lumen or lumen of the tube to the outer surface. The multi-layer tubular prosthesis has sufficient strength and durability to be used in vascular applications without leakage or failure of the tubular prosthesis.

BRIEF DESCRIPTION OF THE DRAWINGS The invention will be further described in relation to the accompanying drawing figures which show the preferred embodiments of the invention, including the specific parts and arrangements of the parts. It is intended that the drawings included as a part of this specification be illustrative of the preferred embodiments of the present invention, and should not be considered in any way as limiting the scope of the invention.

Figure 1 is a perspective view of a compression chamber with a mandrel covered with submucosal tissue, inserted into the lumen or lumen of the compression chamber.

Figure 2 is a sectional view of the compression chamber of Figure 1.

Figure 3 is a perspective view of a single strip of submucosal tissue, helically wound around a mandrel.

Figure 4 is a sectional view of the mandrel covered with submucosa, wherein one end of the mandrel has been sealed and vacuum is attracted to the open end.

Figure 5a is a perspective view of a single strip of water-permeable material wrapped around a multi-orifice mandrel.

Figure 5b is a perspective view of a multi-orifice mandrel that has been wrapped with a single strip of water permeable material, and a first sheet of submucosal tissue.

Figure 5c is a perspective view of a multi-orifice mandrel that has been wrapped with a single strip of water permeable material, a first sheet of submucosal tissue and a second sheet of submucosal tissue.

DETAILED DESCRIPTION OF THE PREFERRED MODALITIES In many medical applications a tubular prosthesis of biological, implantable tissue is desirable. The present invention provides a multi-layer, arbitrary diameter, arbitrary length, biological tissue graft construction. The product can be manipulated to suit various medical applications where a tubular construction or conduit is desired. Examples of possible applications are arterial and venous grafts, ureteral and urethral replacements, and various conduits and leads. The manufacturing process of the tubular constructions of the present invention involves the preparation of a sheet of submucosal tissue according to US Patent No. 4,902,508, and the superposition of the tissue around a mandrel of the appropriate diameter to form a tissue tube. submucosal The sheet of submucosal tissue can be wrapped around the mandrel multiple times, to form a multi-layer tube of submucosal tissue. A second sheet of submucosal tissue is then wrapped around the circumference of the tube formed of the submucosal tissue, and the end of the second leaf of the submucosal tissue is sutured to the graft construction to form a watertight seal that expands longitudinally as length of the tube. The submucosal tissue is then compressed under conditions of dehydration, and optionally heated, to produce the unitary tubular prosthesis of the present invention.

Submucosal tissue suitable for use in the formation of the present graft constructions comprises naturally associated extracellular matrix proteins, glycoproteins and other factors. More particularly, submucosal tissues for use according to the present invention include intestinal submucosa, stomach submucosa, submucosa of the urinary bladder, and uterine submucosa. The intestinal submucosal tissue is a preferred material, and more particularly the submucosa of the small intestine. Suitable intestinal submucosal tissue typically comprises the submucosa of the tunica, delaminated from the tunica muscularis and at least the luminal portion of the tunica mucosa. In one embodiment of the present invention, the intestinal submucosal tissue comprises the tunica submucosa and basilar portions of the tunica mucosa, including the muscularis mucosa lamina and the compact stratum, whose layers are known to vary in thickness and in definition depending on the vertebrate species source . The preparation of intestinal submucosal tissue for use in accordance with this invention is described in US Patent No. 4,902,508. A segment of vertebrate intestine, preferably harvested from a porcine, ovine or bovine species, but not excluding other species, is subject to abrasion using a longitudinal rubbing movement to remove the outer layers, which comprise smooth muscle tissues, and the layer more internal, for example, the portion of the luminal or luminal mucosal tunic. The submucosal tissue is rinsed with saline and optionally sterilized. As a submucosal tissue tissue graft undergoes remodeling and induces the development of endogenous tissues after implantation within a host. It has been used successfully in vascular grafts, in urinary bladder and in hernia repair, in the replacement and repair of tendons and ligaments, and dermal grafts. When used in such applications, the graft constructions appear to not only serve as a matrix for the regrowth of the tissues replaced by the graft constructs, but also promote or induce such endogenous tissue regrowth. The events common to this remodeling process include: very widespread and very rapid neovascularization, the proliferation of mesenchymal granulation cells, the biodegradation / resorption of intestinal submucosal tissue material, implanted, and the lack of rejection by the immune system. The tubular submucosal tissue graft constructions of the present invention can be sterilized using conventional sterilization techniques, including glutaraldehyde tanning, tanning with formaldehyde at acidic pH, treatment with propylene oxide or ethylene oxide, sterilization with gas plasma, gamma radiation , radiation by electron beam, sterilization with peracetic acid. Sterilization techniques that do not adversely affect the mechanical strength, structure and biotropic properties of submucosal tissue are preferred. For example, strong gamma radiation can cause loss of the resistance of the sheets of submucosal tissue. Preferred sterilization techniques include exposure of the graft to peracetic acid, gamma radiation of 1-4 megarads (more preferably 1-2.5 megarads of gamma radiation), treatment with ethylene oxide or sterilization with gas plasma; Sterilization with peracetic acid is the most preferred method of sterilization. Typically, the submucosal tissue is subjected to two or more sterilization processes. After the submucosal tissue is sterilized, for example by guimic treatment, the tissue can be wrapped in a plastic or film wrap and sterilized again using electron beam or gamma radiation sterilization techniques. The submucosal tissue can be stored in a hydrated or dehydrated state. The lyophilized or air-dried submucosal tissue can be rehydrated and used according to the present invention, without significant loss of its biotropic and mechanical properties. The sheets of submucosal tissue can be conditioned, as described in US Patent No. 5,275,826 (the disclosure of which is expressly incorporated by reference herein) to alter the viscoelastic properties of submucosal tissue. According to one embodiment, the delaminated submucosae of the muscular tunic and the luminal portion of the mucosal tunic are conditioned to have a stretch of no more than 20%. The submucosal tissue is conditioned by stretching, chemical treatment, enzymatic treatment or exposure of the tissue to other environmental factors. In one embodiment, the strips of intestinal submucosal tissue are conditioned by stretching in a longitudinal or lateral direction, so that the strips of the intestinal submucosal tissue have a stretch of no more than 20%. In one embodiment, the submucosal tissue is conditioned by stretching the graft material longitudinally to a length longer than the length of the submucosal tissue from which the graft construction was formed. A method of tissue conditioning by stretching involves the application of a given load to the submucosa for three to five cycles. Each cycle consists of applying a load to the graft tissue for five seconds, followed by a relaxation phase of ten seconds. Three to five cycles produce a stretch-conditioned graft material, which reduces deformation. The graft material does not return immediately to its original size; it remains in a "stretched" dimension. Optionally, the graft material can be preconditioned by stretching it in the lateral dimension. In one embodiment, the submucosal tissue is stretched using 50% of the predicted ultimate load. The "ultimate load" is the maximum load that can be applied to the submucosal tissue without resulting in tissue failure (for example, the point of tissue breakdown). The ultimate load can be predicted for a given strip of submucosal tissue, based on the source and thickness of the material. Consequently, a method of fabric conditioning by stretching involves the application of 50% of the last charge predicted to the submucosa, for three to ten cycles. Each cycle consists of the application of a load to the graft material for five seconds, followed by a relaxation phase of ten seconds. The resulting conditioned, submucosal tissue has a stretch or deformation of less than 30%, more typically a deformation of about 20% to about 28%. In a preferred embodiment, the conditioned submucosal tissue has a deformation or stretch of no more than 20%. The term "stretch" or "deformation" as used herein, refers to the maximum amount of elongation of the tissue before the failure or breaking of the fabric, when the fabric is stretched under an applied load. This is expressed as a percentage of the length of the fabric before loading. The conditioned submucosal strips can be used to form the tubular construction or alternatively the tubular construction can be conditioned after its formation. The tubular graft constructions of the present invention are formed as a multi-laminate construction, wherein a first sheet or sheet of submucosal tissue is formed in the shape of a tube of submucosal tissue, and a second sheet is superimposed on the tube of the submucosal tissue. The dimensions of the individual sheets of the submucosal tissue used are not critical, and the term "submucosal tissue sheet" is defined herein to include submucosal tissue from one or more vertebrate organs or sources in a wide variety of sizes and sizes. shapes. After the second sheet of submucosal tissue has been layered on the mandrel, pressure is applied to the overlapping portions to compress the submucosa against the mandrel. In preferred embodiments, the surfaces of the mandrel are water permeable. The term "water-permeable surface" as used herein, includes surfaces that are water-absorbing, microporous or macroporous. Macroporous materials include perforated plates or meshes made of plastic, metal, ceramic or wood. In a preferred embodiment, the multiple layers of the submucosal tissue are compressed under dehydration conditions. The term "dehydration conditions" is defined to include any mechanical or environmental condition that promotes or induces removal of water from submucosal tissue. To promote dehydration of the compressed submucosal tissue, at least one of the two surfaces that compress the tissue is permeable to water. The dehydration of the fabric can optionally be improved or augmented by the application of drying material, heating the fabric or blowing air through the exterior of the compression surfaces. Submucosal tissue typically has an abluminal and luminal surface. The luminal surface is the submucosal surface facing the lumen or lumen of the source organ and typically adjacent to a layer of internal mucosa in vi vo, while the abluminal surface is the submucosal surface remote from the lumen or lumen of the source organ, and typically in contact with smooth muscle tissue in vi. In one embodiment, one or more sheets of submucosal tissue are wrapped on the mandrel with the luminal surface of the submucosal tissue in contact with the surface of the mandrel. In this way, the luminal surface of the submucosal tissue sheet is facing the lumen of the tube formed of submucosal tissue. However, the submucosal tissue tube can also be formed from one or more sheets of submucosal tissue with the abluminal surface facing the lumen or lumen of the formed tubular graft construction. According to one embodiment, a tubular prosthesis is fabricated comprising a first sheet of submucosal tissue formed in the shape of a tube of submucosal tissue, and a second sheet of submucosal tissue circumferentially wrapped around and in adherent contact with the tube of submucosal tissue, wherein the joint or seam formed by the end piece of the second sheet of submucosal tissue is sutured to form a watertight seal. The submucosal tissue tube comprises the first sheet of submucosal tissue, having a first edge and a second opposite edge, formed in the shape of a tube wherein the second opposite edge of the first sheet extends over the first edge of the first. leaf, to define an overlapping region of multiple layers of submucosal tissue. As used herein, the term "overlapping region" refers to the multi-layered tube portion defined by an overlap angle (?) Which extends between the first and second edges of the first sheet of submucosal tissue formed as a tube (see Figure 5b). The layers of submucosal tissue in the overlapping region are fixed to each other using conventional techniques known to those of skill in the art. Alternatively, the layers of submucosal tissue can be fixed to each other by treating the tissue with glutaraldehyde and "vacuum pressing" the overlapping tissue layers., as described later. In one embodiment, multiple layers of submucosal tissue in the overlapped region are fixed to one another by treatment with a crosslinking agent, for example an aldehyde such as formaldehyde or more preferably glutaraldehyde. In one embodiment, the union formed in the submucosal tissue tube can be "spot welded" to ensure that the end piece does not come loose. According to this embodiment, a cotton-tipped wand Q, moistened with?% Glutaraldehyde (or another cross-linking agent or adhesive), is rubbed along the overlapped region that forms the joint or seal. The value for? it is about 0.1 to about 1.0%, more preferably about 0.5%, but there is a relationship between the width of the bond, the concentration of glutaraldehyde and the number of turns that the bursting pressure determines. In one embodiment, the complete graft construction can be immersed in a diluted glutaraldehyde solution (comprising approximately 0.1 to approximately 1.0% glutaraldehyde, and then compressed under dehydration conditions to fix the multiple layers of the submucosal tissue tube one to the other. In addition, the multiple layers of submucosal tissue in the overlapping region can be sutured to one another, and in one embodiment the layers of the overlapped region are stitched in the absence of treatment with a crosslinking agent. , the tube of submucosal tissue formed by the first sheet of submucosal tissue, is formed such that the first and second opposite edges of the first sheet of submucosal tissue are substantially parallel to each other, as shown in Figure 1 and Figure 5b In this modality, the sheet of submucosal tissue is rolled into the shape of a tube that has multiple layers. The tube of submucosal tissue comprises two layers of submucosal tissue and the multiple overlap region comprises three layers of submucosal tissue. The graft constructions of the present invention further comprise a second sheet of submucosal tissue wherein the second sheet is in adherent contact with the outer surface of the submucosal tissue tube. In one embodiment, the first and second opposite edges of the second sheet of submucosal tissue are sutured together along the length of the submucosal tissue tube, without piercing the underlying submucosal tissue tube. In an alternative embodiment, the second opposite edge of the second sheet extends over the first edge of the second sheet, and is sutured to the second sheet of submucosal tissue without piercing the underlying submucosal tissue tube. In preferred embodiments, the overlapped region of the submucosal tissue tube is displaced from the sutures formed in the second sheet of submucosal tissue (eg, the overlapped region does not contact the sutured region of the second sheet). In one embodiment, the sutures formed on the second sheet of submucosal tissue are located at 90-180 ° along the circumference of the submucosal tissue tube relative to the overlapped region, and in one embodiment the sutures are located at 180 °. along the circumference of the tube, of submucosal tissue, in relation to the overlapped region (see Figure 5c). According to one embodiment, the tubular prosthesis comprises a first sheet of submucosal tissue, which has first and second edges that are substantially parallel to one another, wound in the form of a multilayer tube having an overlapped region wherein the first and second edges remain substantially parallel to each other in the tube formed, and the layers of the overlapped region are fixed to each other with sutures, or by exposure to a crosslinking agent. The tubular construction further comprises a second sheet of submucosal tissue adhered to the outer surface of the tube formed of submucosal tissue, wherein the second sheet, which has first and second edges that are substantially parallel to each other, is wrapped circumferentially around the tube of submucosal tissue and the first and second edges are secured to each other with sutures. According to one embodiment, the tubular prosthesis of the present invention is formed by the following steps. A mandrel having a diameter that fits the preferred diameter of the final construction is selected. The mandrel is typically cylindrical in shape and in the preferred embodiments comprises a hollow tube that is permeable to water. A first sheet of submucosal tissue, having a first opposite edge and a second edge, is then superimposed on the mandrel to form a tube of submucosal tissue, wherein the second opposite edge of the first sheet of submucosal tissue extends over the first edge the first sheet of submucosal tissue to define an overlapping region of multiple layers of submucosal tissue. The layers of submucosal tissue in the overlapped region are then attached to each other to form a longitudinally extending junction along the length of the tube formed. A second sheet of submucosal tissue, having a first edge and a second opposite edge is then superimposed on the submucosal tissue tube, and the second opposite edge of the second sheet of submucosal tissue is sutured to the second superimposed sheet of submucosal tissue, along the length of the submucosal tissue tube, without perforating the tube of submucosal tissue. In one embodiment, the first and second edges of the second sheet of submucosal tissue are sutured together to form a second single layer tube that encompasses the first tube of submucosal tissue. The layers of submucosal tissue are then compressed against the mandrel under conditions of dehydration. The invention will be further described with respect to the preferred embodiments as illustrated in the figures of the drawings. With reference to Figure 1, a preferred embodiment of a mandrel 10 for wrapping sheets of biological tissue is illustrated. The mandrel 10 is a metal or hollow plastic tube that comprises holes 12 in the wall of the tube along a portion of, or, alternatively, along the entire length of the metal tube. The size of the holes in the mandrel is not critical, with the condition that the mandrel be sufficiently porous to allow dehydration of the submucosal tissue after compression of the wrapped submucosal tissue 14. In a preferred embodiment, the mandrel is a metal tube , and more preferably, the metal tube is composed of aluminum. The submucosal tissue 14 is superimposed on the mandrel 10 to form a multi-layer tube of submucosal tissue. The mandrel covered with the submucosal tissue is then inserted into the lumen space of a compression chamber 20 which is used in an embodiment to prepare the tubular constructions of the present invention. The compression chamber 20 comprises the outer shield 22, a bladder 24 and the pressure composite 26. The bladder 24 can be coupled or adhered to the inner wall of the outer sheet 22 by various techniques, for example, with an adhesive or bond hot As shown in Figure 2, the wrapped mandrel 10 is inserted into the compression chamber 20 where the inner membrane 18 of the bladder 24 contacts and compresses the submucosal tissue 14 when a fluid is distributed to the pressure gate 26 for inflate the bladder 24. The bladder 24 is inflated to the desired pressure, and the pressure is maintained until the submucosal tissue has been sufficiently dehydrated. Optionally, the compression and drying process can be increased with heating at low temperature (e.g., less than about 50 ° C) of the tissue graft construction.

In addition, air or an inert gas can be passed through (for example, nitrogen) through the lumen or light of the mandrel, as an alternative means or set to increase the drying process. The air / gas extracted through the lumen can optionally be heated to further accelerate the dehydration process. The holes 12 formed in the walls of the mandrel 10 aid in the drying process of submucosal tissue 14, however, this structure represents merely one embodiment of a mandrel, suitable for use according to the present invention (see Figure 1). The mandrel can also be formed as a solid cylinder or as a tube without holes. In one embodiment, the mandrel comprises a hollow tube with holes formed in the walls of the mandrel, and the compression of the submucosal tissue is aided by the removal of a vacuum over the lumen of the mandrel. Alternatively, compression of the submucosal tissue can be achieved by continuous winding of the mandrel containing the wrapped sheets of biological tissue, in direct contact with another surface to provide a direct compression force. Further, in a preferred embodiment, as shown in Figure 4, the application of a vacuum can provide the unique compressive force for compression of the overlapping portions of the multiple strips of submucosal tissue (vacuum pressure). In this embodiment, a mandrel 30, formed as a hollow tube having a plurality of holes 32 formed in the wall of the mandrel 30, is covered with multiple layers of submucosal tissue 34. The mandrel 30 is provided with a plug 36 for sealing the first end of the mandrel 30 and a terminal gate 40 for removing air from the lumen of the mandrel. The end gate 40 is connected to a vacuum generating source and a vacuum is drawn above the mandrel 30, thereby extracting air through the multiple layers of submucosal tissue, while compressing the tissues against each other . A non-permeable layer can be wrapped around the multiple layers of submucosal tissue (for example the mandrel can be placed inside a plastic bag 44 which is sealed with a clamp 46 in the vacuum gate) to provide a second surface that coactuates with the mandrel 30 for compressing the multiple layers of submucosal tissue 34 between the second surface and the mandrel 30. A vacuum is applied, generally in the range of 35.6 to 177.8 cm of hg. (0.49-2.46 kg / cm'1) and more preferably the vacuum applied is approximately 129.5 cm of hg (1.76 kg / cm ") Optionally a heating mantle can be placed on top of the apparatus to heat the submucosal tissue During compression of the tissue After compression of the submucosal tissue for a sufficient period of time, the compressed submucosal tissue is removed from the mandrel as a unitary construction of docile tissue.The multiple strips of submucosal tissue are typically compressed for 12 to 48 hours at room temperature, although heat may also be applied, eg, a heating mantle may be applied to the outside of the compression surfaces to raise the temperature of the compressed tissue to about 40 ° C to about 50 ° C. The overlapping portions are usually compressed by a length of time determined by the degree of dehydration of the tissue.The use of heat increases the speed dehydration and thus decreases the amount of time it takes for the overlapping portions of tissue to be compressed. Typically, the fabric is compressed for a sufficient time to produce a rigid but flexible material. Sufficient dehydration of the tissue is also indicated by an increase in the impedance of the electric current flowing through the tissue. When the impedance has increased by 100 to 200 ohms, the tissue is sufficiently dehydrated and the pressure can be released. In a preferred embodiment shown in Figure 5a, the mandrel 50 is first wrapped with a porous, removable, water-permeable strip material 52 before the sheets of submucosal tissue are superimposed on the mandrel 50.

Preferably, the removable lath material 52 comprises a water permeable material that is tear resistant, including a porous or other plastic material that does not adhere to the submucosal tissue or the mandrel. In one embodiment, the porous ribbon material 52 comprises umbilical tape. The layers of submucosal tissue are then placed in layers directly on the slat material and dried to form a unitary tubular construction. After drying of the submucosal tissue, the porous strip material 52 is unwrapped from the mandrel 50 by pulling on the first end 54 and the second end 56 of the water-permeable material 52 (see Figure 5a). Removal of the strip of water permeable material 52 leaves a space between the submucosal construction and the mandrel 50, allowing the removal of the construction from the mandrel. According to one embodiment, as shown in Figure 5a-Figure 5c, the construction forming method comprises the selection of a hollow mandrel 50, permeable to water, of appropriate diameter, and the spiral wrapping of the mandrel 50 with a strip of porous ribbon material 52, for example umbilical tape. The mandrel is preferably permeable to water and in one embodiment is provided with a plurality of holes. Then, the desired number of turns of a first sheet of submucosal tissue 60 is applied where a "turn of submucosal tissue" is defined as a piece of submucosal tissue wrapped 360 ° around the mandrel. Typically 1 or 2 turns provide a bursting resistance of approximately 1000-2000 mmHg after manufacture, as described below. The first sheet of submucosal tissue 60 is applied to the mandrel 50 by a rotary movement with the desired number of layers (typically two) and an overlapped region 62 (defined by an overlap angle (?) Extending between the first edge 64 and the second opposite edge 66 of the first sheet of submucosal tissue 60) to form a tube of submucosal tissue having an overlapping region 62 extending longitudinally, which forms the junction of the tube (Figure 5b). The overlap angle is in the range typically from about 20 ° to about 90 °, more typically from about 20 ° to about 40 °, and in one embodiment the overlap angle is about 30 °. The submucosal tissue is wrapped on the mandrel leaving the first end 54 and the second of the water permeable material 56, exposed. In one embodiment, the binding of the wrapped fabric can be "spot welded" to ensure that the second opposing edge 66 does not come loose. According to this embodiment, a cotton-tipped wand moistened with?% Glutaraldehyde (or another cross-linking agent or adhesive) is rubbed along the surfaces of the submucosal tissue portion that forms the overlapped region 62. The value for? is from about 0.1 to about 1.0%, more preferably from about 0.5%, but there is a relationship between the width of the overlapped region 62, the concentration of glutaraldehyde and the number of turns that determines the bursting pressure. The layers of submucosal tissue that form the overlapping region can also be crosslinked to ensure that the end piece does not become loose by immersing the entire tube of submucosal tissue in a solution of a crosslinking agent such as glutaraldehyde. The glutaraldehyde-treated tube of submucosal tissue can optionally be compressed under dehydration conditions before the superimposition of the second sheet of submucosal tissue onto the tube of submucosal tissue (including the use of vacuum pressing) to further bond the layers of submucosal tissue at each other. Alternatively, the union of the submucosal tissue tube can be fixed by suturing the multiple layers of the submucosal tissue tube along the overlapping region. The use of sutures negates the need to crosslink the layers of submucosal tissue of the overlapping region. According to the present invention, the first sheet of submucosal tissue can be wrapped over the mandrel in a variety of different orientations. In one embodiment, the sheets have a width equal to the length of the mandrel such that a single sheet completely covers the mandrel when it is wrapped 360 ° around the mandrel (see Figure 5b). Other wrapping techniques can be used to form the first sheet of submucosal tissue in the submucosal tissue tube, provided that there are no free spaces between the overlapping tissue junctions. In one embodiment, the submucosal tissue sheet may have a width less than the desired length of the tube formed of submucosal tissue. In this alternative embodiment, a narrow sheet of submucosal tissue 80 is wrapped around the mandrel 82 several times, wherein the sheet is at least partially overlapped as it is superimposed on the mandrel, leaving no exposed portion of the underlying mandrel (see Figure 3) . The mandrel is permeable to water and in one embodiment is provided with a plurality of holes 84. The amount of overlap in such partially overlapped sheets of submucosal tissue is in the range between 10 and 60% of the width of the individual sheet, and more preferably the overlapped portion is an overlap of 50%. In one embodiment, multiple pieces of submucosal tissue can be superimposed on the mandrel, with the proviso that at least a portion of each piece of submucosal tissue overlaps a portion of another piece of submucosal tissue wrapped around the mandrel. In a further embodiment, a narrow, long sheet of submucosal tissue can be spirally wrapped on the mandrel with an overlap, followed by a spiral wrapping in the opposite direction. This will provide 4 layers of submucosal tissue to withstand the internal pressure. In these embodiments, the seams formed by the overlapped sheets of submucosal tissue should overlap by 0.5 to 3 cm, and more preferably 1 to 2 cm. In embodiments where the first sheet of submucosal tissue used to form the submucosal tissue tube has a smaller width than the desired tube length of submucosal tissue, the tube joints will preferably be fixed by exposure to a crosslinking agent. After formation of the submucosal tissue tube, a second sheet of submucosal tissue 70 is then circumferentially wrapped around the outer surface of the tube formed of submucosal tissue 60. In one embodiment, the second sheet of submucosal tissue 70 is superimposed on the tube. of submucosal tissue 60, wrapped around the tube once, and the first edge 74 and the second opposite edge 76 of the second sheet of submucosal tissue are sutured together along the length of the tube of submucosal tissue, with sutures 78 not punctured the submucosal tissue tube 60 (see Figure 5c). Alternatively, the second sheet of submucosal tissue is superimposed on the tube of submucosal tissue, wrapped around the tube at least once, wherein the second opposite edge of the second sheet extends over the first border of submucosal tissue, and the second edge The opposite of the second leaf is sutured along the length of the submucosal tissue tube, without perforating the submucosal tissue tube. Furthermore, when the second sheet of submucosal tissue is superimposed on the tube of submucosal tissue formed from the first sheet of submucosal tissue, the second sheet can be superimposed with its abluminal surface or its luminal surface in contact with the submucosal tissue tube. Each of these combinations of overlap of the tissue-submucosal sheets from the same or from different sources of vertebrate or organ, will produce a submucosal tissue graft construction, of unitary tubular shape after compression of at least the overlapping portions, under conditions that allow tissue dehydration. After the mandrel has been wrapped with the second sheet of submucosal tissue, the submucosal tissue is compressed under conditions of dehydration. In one embodiment, the fabric is vacuum pressed, wherein one end of the mandrel is closed and the interior of the mandrel covered with submucosal tissue is connected to a vacuum pump. The vacuum causes the atmospheric pressure to compress the layers of submucosal tissue, and in those embodiments which use a crosslinking agent, which crosslink the layers of submucosal tissue one to the other, the vacuum pressing procedure causes the crosslinking agent to penetrate The full thickness of the seal or union. The vacuum sealing / drying process is completed in approximately four hours typically. After the sealing / drying process is completed, the first and second ends (54 and 56, respectively) of the porous water-permeable ribbon material 52 (e.g., umbilical tape) are fastened and pulled longitudinally. The tape is easily unrolled under the submucosal tissue tube and the submucosal tissue tube is then easily slid out of the mandrel. The result is a tube without a union that looks like a straw. In one embodiment, a tube of submucosal tissue can be prepared from one or more sheets of submucosal tissue by wrapping one or more sheets of submucosal tissue around the circumference of a mandrel, exposing the wrapped submucosal tissue to an agent of submucosal tissue. crosslinking, and comprising the wrapped tissue under dehydration conditions. The tissue sheets can be superimposed on the mandrel using a variety of wrapping techniques, provided that there are no free spaces between the seams or seams of the superimposed sheets of submucosal tissue. In one embodiment, a single sheet of submucosal tissue having a first edge portion and a second edge portion is superimposed on a mandrel, such that the first and second edge portions of the submucosal tissue sheet are superimposed to form a tube of submucosal tissue. In one embodiment, the second edge of the submucosal tissue sheet is extended beyond the first edge of the submucosal tissue sheet, wherein the first edge is substantially parallel to the second edge, to form a heterolaminar tubular construction as shown in FIG. Figure 5b. Alternatively, a simple narrow sheet of submucosal tissue can be spirally wrapped around the mandrel as shown in Figure 3. The superimposed portions of the sheet of submucosal tissue are then contacted with a crosslinking agent. In one embodiment, the complete graft construction is submerged in a crosslinking solution. A preferred crosslinking agent is glutaraldehyde, in a concentration in the range of about 0.1% to about 1% of glutaraldehyde. After the superimposed or overlapping layers of submucosal tissue have been treated with the crosslinking agent, the superimposed layers are compressed under dehydration conditions. In one embodiment, the superimposed layers of submucosal tissue are vacuum pressed, wherein one end of the mandrel is closed and the interior of the submucosal tissue is brought into contact with a vacuum pump. The vacuum causes the atmospheric pressure to compress the layers of submucosal tissue and the vacuum pressing procedure causes the crosslinking agent to penetrate the full thickness of the graft construction. As indicated by the data of Example 4, submucosal tissue tubes that have been treated with a glutaraldehyde solution of at least 1% glutaraldehyde, show significant durability under pulsatile pressure testing. In addition, a tube of submucosal tissue formed by wrapping the sheets of submucosal tissue in a tube heterolaminated and treated with glutaraldehyde and vacuum pressed, provides a tube of submucosal tissue with a smooth luminal surface. It is known that the treatment of biomaterials with glutaraldehyde promotes calcification, the poor incorporation of host tissue and the final mechanical failure of the bioprostheses. To minimize the damaging effects of glutaraldehyde treatments of the biomaterials, the tubular constructions of the present invention are exposed to relatively dilute glutaraldehyde solutions of about 0.1% to 1% glutaraldehyde. Tubular constructions treated with glutaraldehyde, formed in accordance with the present invention, by treatment with a solution comprising less than 1% glutaraldehyde can be used in various non-vascular applications such as ureter and urethral replacements and as various conduits and shunts . In addition a tube of submucosal tissue, having the joints of the submucosal tissue tube sealed by a crosslinking agent, can be attached to a second sheet of submucosal tissue to support the bonds made by glutaraldehyde. In one embodiment, the submucosal tissue tube is rehydrated and the second sheet of submucosal tissue is wrapped around the submucosal tissue tube and compressed against the submucosal tissue tube under dehydration conditions. In one embodiment, a tube of submucosal tissue is prepared from a sheet of submucosal tissue, and contacted with a solution of glutaraldehyde (wherein the concentration of glutaraldehyde is less than 1%, and more preferably less than 0.5% ), vacuum pressed and then wrapped with a second sheet of submucosal tissue that is sutured along its full length, as shown in Figure 5c. The second sheet of submucosal tissue in this modality provides sufficient support for the underlying submucosal tissue tube, such that the construction can be used in various vascular applications, including arterial and venous replacement, despite the low concentration of the crosslinking agent used for seal the submucosal tissue tube. In one embodiment, the method of preparing a submucosal tubular prosthesis comprises overlaying a first strip of water-permeable material around the circumference of a mandrel, and then superimposing a second strip of water-permeable material on top of the first strip of material permeable to water. A first sheet of submucosal tissue, which has a first edge and a second opposite edge, is then superimposed on the spirally wrapped material to form a tube of submucosal tissue, wherein the second opposite edge extends over the first edge to define a region. overlapping multiple layers, submucosal tissue. The submucosal tissue tube is then compressed against the surface of the mandrel under dehydration conditions to form a tube-like construction. The second strip of water-permeable material is then removed from the mandrel to release the tube-like construction of the mandrel. The submucosal tissue layers of the overlapped region of the tube released from submucosal tissue are then sutured together, using a continuous suture, to ensure that the tube will not unravel. The tube-like construction is then again placed on the mandrel by sliding the tube over the first strip of water permeable material covering the mandrel. A second sheet of submucosal tissue, which has a first edge and a second opposite edge, is then superimposed on the tube of submucosal tissue. The first and second opposite edges of the second sheet of submucosal tissue are sutured together along the length of the submucosal tissue tube, without perforating the submucosal tissue tube, and the multiple layers of submucosal tissue are compressed together under dehydration conditions. . The first strip of water-permeable material is then removed from the mandrel to release the tubular prosthesis from the mandrel. In preferred embodiments, the sutures of the second sheet of submucosal tissue do not overlap the sutures formed in the overlapped region of the submucosal tissue tube. The multilaminated tissue graft constructions, can be formed to have substantially isotropic properties. These substantially isotropic grafts (pseudoisotropic) are prepared from at least two sheets of intestinal submucosal tissue, delaminated from the tunica muscularis and from the luminal portion of the mucosal tunic of a warm-blooded vertebrate. Each of the intestinal submucosal tissue sheets are characterized by having a longitudinal axis that corresponds to the predominant orientation of the collagen fibers in the sheets of submucosal tissue. The method for the formation of the pseudoisotropic graft constructions comprises placing a first sheet of submucosal tissue on the mandrel, superimposing the first sheet with at least one additional sheet of submucosal tissue, so that the longitudinal axes of each individual leaf of submucosal tissue form an angle of approximately 90 ° with the longitudinal axis of the other sheet of submucosal tissue forming the heterolaminate graft. The implants or constructions of biological tissue produced according to the present invention overcome the problem of leakage of fluid around the suture holes of the sewn biological tissues. The construction of two concentric tubes of submucosal tissue, where the sutured joints of the tube of the two tubes are displaced relative to one another followed by the adherence of the two tubes of submucosal tissue together, which ensures that the suture holes are not they will leak in the formed prosthesis. In addition, the modalities that use a crosslinking treatment to seal the first tube joint clearly have no perforations formed in the wall of the tube, and thus will not leak. The building tubes produced in accordance with the present invention show an essentially unbonded tube which will not allow the fluid to escape or require extra precautions associated with the efflux of the fluid. This property is particularly important when the construction is going to be used as a vascular graft, ureter replacement or as a bypass. The construction can also be manipulated (for example, cut, folded, sutured, etc.) to suit the various medical applications where submucosal material of the present invention is regulated. Other features and aspects of this invention may be appreciated by those of skill in the art in reading and understanding this specification. Such features, aspects and variations and expected modifications are clearly within the scope of this invention.

Example 1 Preparation of Tubular Submucosal Tissue Graft Constructions In one embodiment, the method for forming the construction comprises the selection of a hollow mandrel permeable to water, of appropriate diameter, and spirally wrapping the mandrel with umbilical tape. Then the desired number of turns of submucosal tissue is applied where a "wrap or wrap of submucosal tissue" is defined as a piece of submucosal tissue wrapped 360 ° around the mandrel. Typically one or two turns provide a bursting resistance of about 1000 to 2000 mmHg after manufacture, as described below. A sheet of submucosal tissue is placed on a smooth, flat surface, with the mucosal side up. The mandrel wrapped with tape is placed on it with the longitudinal axis parallel to the longitudinal axis of submucosal tissue. A razor blade is then used to cut the submucosal tissue parallel to the axis of the mandrel to form a linear border for the submucosal tissue. The submucosal tissue is applied to the mandrel by a rotary movement with the desired number of layers (typically two) and a region of overlap (defined by an overlap angle (?) Of approximately 30 degrees which extends between the two lateral edges of the tissue. submucosal) forms a tube of submucosal tissue that has a longitudinally extending junction. After the mandrel has been completely wrapped, one end of the mandrel is closed and the inside of the mandrel covered with submucosal tissue is connected to a vacuum pump. The vacuum causes the atmospheric pressure to compress the layers of submucosal tissue and accelerate the dehydration of the submucosal tissue. The vacuum sealing / drying process is completed in approximately four hours typically. After the drying / sealing treatment, the ends of the umbilical tape are fastened and pulled longitudinally. The tape is easily unrolled under the tube of submucosal tissue which then slides out of the mandrel, easily. The result is seamless tubes that look like a straw or straw.

Other types of sealant, concentrations and wrapping techniques can be used. For example, the spiral winding of a narrow, long strip of submucosal tissue over the mandrel with an overlap is an option, followed by a spiral wrapping in the opposite direction, which is another. This will provide 4 layers of submucosal tissue to withstand the internal pressure. This double helix could be combined with the wrapping techniques shown in Figure 3. In accordance with the present invention, the submucosal tissue can be wrapped on the mandrel in a variety of different orientations. One limitation is that there should be no empty spaces between the overlapping tissue joints. In preferred embodiments, the units formed by the overlapped strips of submucosal tissue should overlap by 0.5 to 3 cm, and more preferably 1 to 2 cm.

Example 2 Spot Welding of the Submucosal Tissue Unions In one embodiment, the binding of the wrapped fabric can be "spot welded" to ensure that the end piece does not come loose. According to one embodiment, the joint is spot welded with a crosslinking agent such as glutaraldehyde. A cotton-tipped wand type Q tip, moistened with?% Glutaraldehyde (or another cross-linking agent or adhesive), is rubbed along the overlap which is the joint. The value for? it is about 0.1 to about 1.0%, more preferably about 0.5%, but there is a relationship between the width of the bond, the concentration of the glutaraldehyde and the number of turns determined by the burst pressure. Alternatively, the joint can be welded in points through the use of a laser. In yet another embodiment, the junction of the submucosal tube can be thermally welded by stitches to further seal the tissue junction in a unitary submucosa tube. There are four factors that control the quality of a thermal spot weld applied to the submucosal tissue: 1) temperature, 2) force, 3) time of force application, and 4) the shape of the thermal welding tool. A pointed tool makes a weld with a hole. A flat tip does not make a hole and a tip with a radius can make a small hole. Using a small solder iron with a calibrated temperature tip, the submucosal tissue can be fused at discrete sites to form u? point of "welding" between two pieces of submucosal tissue. When placing a specimen of submucosal tissue on a glass plate (to avoid heat shrinkage) the hot tip is applied to the submucosal tissue to determine the temperature and time of application necessary to melt the tissue. Studies have led to strips of 1 cm width of submucosal tissue overlapped by 1 cm to determine the number of thermal welds per points, required to hold two pieces of submucosal tissue together. The welds of five points in the overlap area of 1 x 1 cm produce a weld that is stronger than the force required to break a strip 1 cm wide, of simple thickness. A pointed tip produces a welded joint with a hole; therefore, the challenge is to identify the parameters that produce the strongest welding with the smallest hole. To optimize the spot welding conditions, the following experiments were conducted. Pieces 1 cm wide and 10 cm long submucosal tissue are used and 50 specimens will be manufactured and tested with three of the four variables (temperature, time, strength and shape of the tip) that are kept constant and a variety . In these first studies, the pointed tip will be used. Subsequently the studies will be repeated with the tip of radius of 0.5 mm, then with a flat tip of diameter of 1 mm. The result will be the formula for the strongest welding with the smallest hole, which is measured with a microscope. The resistance to breakage of the selected specimens will also be determined, using the MTS machine. b) Animal Model The weanling rat model will be used to investigate the host response to submucosal strips welded by stitches. The anesthesia will be induced and maintained with metafano administered by means of a facial mask. The ventral abdomen will be clamped and prepared for aseptic surgery. Longitudinal incisions will be made in the skin in each abdominal quadrant. Then the bilateral subcutaneous bags will be created in the subcutis of each rat by blunt dissection. A 1 cm2 test specimen will be placed subcutaneously with each bag and secured in position with a 5-0 polypropylene suture to the underlying fascia. The incisions in the skin will be closed with a simple interrupted suture pattern with 5-0 polypropylene. Twenty-four rats will be used in the study. After the elapsed time, euthanasia will be performed with intracardiac potassium chloride (at 1, 2, 4 and 8 weeks after implantation).

Morphological Analysis The samples removed for morphological evaluation will be fixed in Trump's fixative for 24 hours, then placed in a phosphate buffer. Specimens for light microscopy will be embedded in paraffin and sectioned at 2-3 μm. The sections will be stained with hematoxylin and eosin (H &E) for complete morphology and VonKossa staining for evaluation of calcification.

Example 3 Submucosal Tissue Tube Manufacturing and Testing The goal of this study is to produce grafts with an outside diameter of 5.0 mm. A hollow mandrel with multiple orifices of 4 mm diameter is spirally wrapped with non-lapped umbilical tape. Then two and a half turns of submucosal tissue are applied to form a tube of submucosal tissue, and the overlapped portion is fixed with glutaraldehyde in one group and with sutures in another group. A second sheet of submucosal tissue is then wrapped around the tube of submucosal tissue and the opposite ends of the second sheet of submucosal tissue are sutured together. After wrapping, one end of the mandrel is connected to a vacuum pump and the other end of the mandrel is closed. The resulting vacuum in the mandrel causes the atmospheric pressure to press the layers of submucosal tissue together and firmly, and extracts the moisture from the submucosal tissue, which requires approximately 24 hours. After drying of the submucosal tissue layers, the ends of the umbilical tape are fastened and pulled longitudinally. The tape is unrolled uniformly under the tube of submucosal tissue, which then slides out of the mandrel, easily.

Static Test Before the burst test, each submucosal tissue graft will be soaked in 0.9% saline at 37 ° C for 24 hours. The purpose of this procedure is to determine the durability of the joints.

To determine the burst strength / one end of the tubular submucosal tissue graft, it is mounted to an accessory which will be used to apply air pressure. The other end of the tubular submucosal tissue graft is closed with a suture and increasing air pressure is applied. A continuous record of the pressure versus time allows the exact identification of the bursting pressure. The objective is a burst pressure of 1000 mmHg or more. A successful manufacturing technique is one that produces a burst (at any pressure), without delamination. The submucosal tubular tissue graft that passes this static test will proceed to the pulsatile test.

Pulsatile Test The critical period for a tubular submucosal tissue graft is the first few weeks after the implant, when it is in the initial stage of remodeling and is exposed to warm blood with a static (diastolic) and pulsatile pressure. During this time it is essential to know if the graft will retain its resistance during early remodeling. Consequently, the pulsatile pressure test will be carried out with the graft constructions in saline solution at 37 ° C. A pulsatile pressure of 200/150 mmHg will be used with a frequency of 1 / second. The test will continue for 2 weeks after which the graft will be tested in static burst.

Example 4 Pulmonative Test of Submucosal Tissue Tubes To test the durability of tubular constructions derived from the submucosa, fabricated from sheets of submucosal tissue according to the present invention, a pulsatile test method was developed. A pulsating pressure pump was mounted which supplies pulses of adjustable intensity pressure at 4.5 Hz. The working fluid for the system is sterile water. The pressure is supplied to the tubular graft constructions of submucosal tissue formed, which are attached to the outlet of the pump. The tubes are immersed in water at 37 ° C to simulate physiological conditions. Typically, the pulsatile test system is run at pressures of 150/50 mmHg or 400/200 mmHg. A drop in the applied pressure is indicative of tissue graft failure.

Test Results of Tubes Derived from the Submucosa Submucosal Tissue Tubes Treated with Glutaraldehyde Submucosal tissue of the small intestine, hydrated (2.5 turns), wrapped around a hollow mandrel with multiple orifices, and immersed in a diluted solution of glutaraldehyde (GA) and dried under vacuum as described above. These GA-treated tubes of submucosal tissue were then tested on a pulsatile test machine. The grafts that were treated with > of 1% solution of GA (w / w) showed significant durability under pulsatile pressure (400/200), surviving more than 5.5 million pulsatile cycles. Tubes treated with GA solution to < of 1% (w / w) were significantly less durable, lasting less than four hundred cycles under similar conditions. Consequently, treatment of the submucosal tissue tubes with 1% (w / w) or more of GA solution provided a tube of submucosal tissue having the desired durability at pulsatile pressure.

Submucosal Tissue Tubes Sutured by Hand, Not Treated The tissue of the small intestine submucosa, hydrated, was wrapped around a hollow mandrel, with multiple orifices, dried under vacuum, the union was sutured and then wrapped with an additional layer of submucosal tissue from the small intestine, and sutured by hand longitudinally as shown in Figures 5b and 5c. These tubes, hand-sutured, untreated, were then tested on the pulsatile test magnet. The results of these tests showed that the hand-stitched tubes survived more than 6 million pulsatile cycles, the test being intentionally discontinued and the durability limit not yet evaluated.

It is noted that in relation to this date, the best method known to the applicant to carry out the aforementioned invention is that which is clear from the present description of the invention.

Having described the invention as above, property is claimed as contained in the following:

Claims

1. A unitary multi-layer submucosal tissue prosthesis, characterized in that it comprises: a first sheet of submucosal tissue, having a first edge and a second opposite edge, formed in the shape of a tube of submucosal tissue, wherein the second opposite edge of the first sheet extends over the first edge of the first sheet, to define an overlapped region of multiple layers of submucosal tissue, wherein the layers of submucosal tissue in the overlapped region are fixed one to the other; and a second sheet of submucosal tissue, having a first edge and a second opposite edge, wherein the second sheet is in adherent contact with the tube of submucosal tissue and the first edge and the second opposite edge of the second sheet are sutured together along the length of the submucosal tissue tube, without perforating the underlying submucosal tissue tube.

2. The prosthesis according to claim 1, characterized in that the tube of submucosal tissue is formed from the first sheet of submucosal tissue such that the first and second opposite edges of the first sheet of submucosal tissue are substantially parallel to one another.

3. The prosthesis according to claim 2, characterized in that the tube of submucosal tissue comprises two layers of submucosal tissue and the multiple overlapped region comprises three layers of submucosal tissue.

4. The prosthesis according to claim 1, characterized in that the multiple layers of submucosal tissue in the overlap region are fixed to each other by treatment with a crosslinking agent.

5. The prosthesis according to claim 4, characterized in that the tube of submucosal tissue and the second sheet of submucosal tissue are fused together by compressing the submucosal tissue under conditions which lead to dehydration of the tissue.

6. The prosthesis according to claim 4, characterized in that the crosslinking agent is glutaraldehyde.

7. The prosthesis according to claim 1, characterized in that the multiple layers of submucosal tissue in the overlapped region are fixed to each other by sutures, and the overlapped region is displaced from the sutures formed in the second sheet of submucosal tissue.

8. The prosthesis according to claim 7, characterized by the tube of submucosal tissue and the second sheets of submucosal tissue are fused together by compressing the submucosal tissue under conditions that lead to dehydration of the tissue.

9. The prosthesis according to claim 1, characterized in that the submucosal tissue comprises submucosa of delaminated intestinal tissue of the abluminal muscle layers and at least the luminal portion of the mucosal tunic of a warm-blooded vertebrate.

10. The prosthesis according to claim 1, characterized in that the tube of submucosal tissue is formed with the abluminal surface of the sheet of submucosal tissue on the outside of the tube.

11. A submucosal, multilayer, unitary tissue prosthesis, characterized in that it comprises: a first sheet of submucosal tissue, having a first edge and a second opposite edge, formed in the shape of a tube of submucosal tissue, wherein the second edge opposite of the first sheet extends over the first edge of the first sheet, to define an overlapping region of multiple layers of submucosal tissue, wherein the layers of submucosal tissue in the overlapped region are fixed to each other; and a second sheet of submucosal tissue, having a first edge and a second opposite edge, wherein the second sheet is in adherent contact with the tube of s-ubmucosal tissue and the second opposite edge of the second sheet extends over the first edge of the second leaf and the second sheet of submucosal tissue is sutured, without perforating the underlying submucosal tissue tube.

12. The prosthesis according to claim 11, characterized in that the tube of submucosal tissue is formed from the first sheet of submucosal tissue such that the first and second opposite edges of the first sheet of submucosal tissue are substantially parallel to one another.

13. The prosthesis according to claim 11, characterized in that the multiple layers of submucosal tissue in the overlapped region are fixed to each other by treatment with a crosslinking agent.

14. The prosthesis according to claim 13, characterized in that the tube of submucosal tissue and the second sheet of submucosal tissue are fused together by compressing the submucosal tissue under conditions which lead to dehydration of the tissue.

15. The prosthesis according to claim 13, characterized in that the crosslinking agent is glutaraldehyde.

16. The prosthesis according to claim 11, characterized in that the multiple layers of submucosal tissue in the overlapped region are fixed to each other by sutures, and the overlapped region is displaced from the sutures formed in the second sheet of submucosal tissue.

17. The prosthesis according to claim 11, characterized in that the tube of submucosal tissue and the second sheets of submucosal tissue are fused together by compressing the submucosal tissue under conditions that lead to dehydration of the tissue.

18. The prosthesis according to claim 11, characterized in that the submucosal tissue comprises submucosa of delaminated intestinal tissue of the abluminal muscle layers and at least the luminal portion of the mucosal tunic of a warm-blooded vertebrate.

19. A method for preparing a tubular prosthesis from sheets of submucosal tissue, characterized in that the method comprises: selecting a mandrel having a predetermined diameter; the superimposition of a first sheet of submucosal tissue, having a first edge and a second opposite edge, on the mandrel to form a tube of submucosal tissue, wherein the second opposite edge of the first sheet of submucosal tissue extends over the first edge of the first sheet of submucosal tissue, to define an overlapping region of multiple layers of submucosal tissue; the fixation of the layers of submucosal tissue in the overlapping region, one to the other; the superposition of a second sheet of submucosal tissue, having a first edge and a second opposite edge, on the tube of submucosal tissue; the suture of the second opposite edge of the second sheet of submucosal tissue to the second overlapped sheet of submucosal tissue, along the length of the submucosal tissue tube, without perforating the tube of submucosal tissue; and the compression of the submucosal tissue tube and the second sheet of submucosal tissue against the mandrel, under dehydration conditions.

20. The method according to claim 19, further characterized in that it comprises the step of superimposing a strip of water permeable material on the mandrel, before the first sheet of submucosal tissue is superimposed on the mandrel, where after the formation of The tubular prosthesis, the strip of water-permeable material is removed from the mandrel to aid in the release of the tubular prosthesis from the mandrel.

21. The method according to claim 20, characterized in that the first and second opposite edges of the second sheet of submucosal tissue are sutured together along the length of the tube of submucosal tissue, without piercing the tube of submucosal tissue.

22. The method according to claim 20, characterized in that the second opposite edge of the second sheet extends over the first edge of the second sheet of submucosal tissue, and the second opposite edge of the second sheet is sutured along the length of the submucosal tissue tube, without perforating the submucosal tissue tube.

23. A method for preparing a tubular prosthesis from sheets of submucosal tissue, characterized by the porgue method comprises: selecting a mandrel having a predetermined diameter; the superimposition of a first sheet of submucosal tissue, which has a first edge and a second opposite edge, on the strip of water permeable material to form a tube of submucosal tissue, wherein the second opposite edge of the first sheet of tissue submucosal extends over the first edge of the first sheet of submucosal tissue to define an overlapping region of multiple layers of submucosal tissue, contacting the tube of submucosal tissue with a solution containing a cross-linking agent, the superposition of a second sheet of submucosal tissue, which has a first edge and a second opposite edge on the tube of submucosal tissue, the suture of the first and second opposite edges of the second sheet of submucosal tissue, together along the length of the tube of submucosal tissue , without perforating the tube of submucosal tissue, and the compression of the tube of submucosal tissue and the second sheet of submucosal tissue against the mandrel, conditions of dehydration.

24. The method according to claim 23, further characterized by comprising the step of superimposing a strip of water permeable material on the mandrel, before the first sheet of submucosal tissue is superimposed on the mandrel, where after the formation of The tubular prosthesis, the strip of water-permeable material is removed from the mandrel to aid in the release of the tubular prosthesis from the mandrel.

25. The method according to claim 24, further characterized by comprising the step of compressing the submucosal tissue tube against the mandrel under conditions that lead to dehydration of the tissue after the tube of submucosal tissue is brought into contact with the agent of submucosal tissue. cross-linking, and before the second sheet of submucosal tissue overlaps the submucosal tissue tube.

26. The method according to claim 24, characterized in that the crosslinking solution comprises from about 0.1 to about 1.0% glutaraldehyde.

27. The method according to claim 24, characterized in that the mandrel is a hollow porous tube having a luminal space, and the step of compressing the biological tissue is carried out by extracting a vacuum on the lumen of the mandrel.

28. The method according to claim 24, characterized in that the tube of submucosal tissue and the second sheet of submucosal tissue are heated during the compression step of the submucosal tissue tube and the second sheet of submucosal tissue.

29. The method according to claim 24, characterized by the step of compressing the tube of submucosal tissue and the second sheet of submucosal tissue is carried out by means of a compression chamber.

30. A method for preparing a tubular prosthesis from sheets of submucosal tissue, characterized by the porgue method comprises: selecting a mandrel having a predetermined diameter; overlapping a first sheet of submucosal tissue, having a first edge and a second opposite edge, on the water-permeable material to form a tube of submucosal tissue, wherein the second opposite edge of the first sheet of submucosal tissue extends on the first edge of the first sheet of submucosal tissue to define an overlapped region of multiple layers of submucosal tissue; the compression of the submucosal tissue tube against the mandrel, under conditions of dehydration; the suture of the layers of the overlapped region, together; the superposition of a second sheet of submucosal tissue, which has a first lateral edge and a second opposite edge, on the tube of submucosal tissue; the suture of the first and second opposite edges of the second sheet of submucosal tissue, together along the length of the tube, without perforating the tube of submucosal tissue; and the compression of the submucosal tissue tube and the second sheet of submucosal tissue against the mandrel under dehydration conditions.

31. The method according to claim 30, further characterized by comprising the step of superimposing a strip of water permeable material on the mandrel, before the first sheet of submucosal tissue is superimposed on the mandrel, where after the formation of The tubular prosthesis, the strip of water-permeable material is removed from the mandrel to aid in the release of the tubular prosthesis from the mandrel.

32. The method according to claim 31, characterized in that the mandrel is a porous hollow tube having a luminal space, and the step of compressing the biological tissue is carried out by imposing a vacuum on the lumen of the mandrel.

33. The method according to claim 31, characterized in that the tube of submucosal tissue and the second sheet of submucosal tissue are heated during the compression step of the submucosal tissue tube and the second sheet of submucosal tissue.

34. The method according to claim 31, characterized by the compression step of the submucosal tissue tube and the second sheet of submucosal tissue is achieved by a compression chamber.

35. A method for the formation of a unitary heterolaminar tubular construction from a sheet of submucosal tissue, characterized by the porgue method comprises: superimposing a sheet of submucosal tissue having a first edge portion and a second edge portion on a mandrel , wherein the first and second edge portions of the submucosal tissue sheet overlap to form a tube of submucosal tissue; the contact of the first and second edge portions overlapped with a crosslinking agent; and the compression of the first and second overlapping edge portions, under dehydration conditions.

36. The method according to claim 35, characterized in that the first and second overlapping portions are compressed by vacuum pressure.

37. The method according to claim 36, characterized in that the crosslinking agent is glutaraldehyde.