Method for producing preserved collagen connective tissue, collagen connective tissue, its use and kit for tissue implantation

Abstract

The present invention relates to the preservation of collagen connective tissue, which achieves structural integrity of collagen fibers and makes the tissue suitable for use in implants without causing immune and / or inflammatory rejection, comprising the steps of trimming the collagen connective tissue in a buffer solution, washing the connective tissue with a buffer solution, stabilizing the tissue in an ethanol solution, treating the tissue with a polyethylene glycol solution, washing and storing the tissue in ethanol, sterilizing the tissue with an ethanol and hydrogen peroxide solution, and storing the tissue in an ethanol and indomethacin solution for transport. A preferred use of the present invention is in surgical interventions to correct various abnormalities of the human body, where implants are particularly needed.

Description

[Technical field]

[0001] The present invention focuses on a method for producing collagen connective tissue, the tissue thus produced, and its uses. In particular, the present invention refers to the preservation of collagen connective tissue to provide structural integrity of collagen fibers, making said tissue suitable for use in implants, free of immune and / or inflammatory rejection. The present invention is preferably utilized in surgical interventions to correct various abnormalities of the human body, where grafts are particularly needed. [Background technology]

[0002] In the area of ​​grafts or implants, there are several examples in the state of the art of using collagen connective tissue. For example, US Pat. No. 5,399,633 discloses a method and method of manufacturing for treating collagen-containing biomaterials to reduce or mitigate calcification and improve the lifespan of biomaterials that can be used as implantable devices. US Pat. No. 5,499,633 discloses a biocompatible composite implant and a method of forming the same, which comprises a framework of cross-linked collagen tissue processed without the use of glutaraldehyde and elongated members that are held in contact with each other throughout the length of the framework.

[0003] Nevertheless, there remains a need in the medical field for improved methods for processing collagen connective tissue from animal donor sources so that the collagen fibers can be implanted in a recipient without any immune and / or inflammatory rejection reaction, which is the main object of the present invention.

[0004] Although the present invention is not limited to use in reconstructive surgical procedures in humans, two applications of conventional grafts in surgical procedures are described below: the first is (i) the correction of penile curvature, and the second is (ii) facial reconstruction applications in humans.

[0005] (i) Penile curvature: As is well known in the medical field, the penis, the male genitalia, exhibits some degree of curvature in most men when in an erect state. Desvios of less than 20 degrees on the shaft of the penis are considered normal and generally do not adversely affect, for example, penetration and therefore do not cause problems in the male's sex life. Desvios of more than 20 degrees, however, are considered pathological and affect at least 10% of men worldwide. Depending on the degree of curvature, penetration may become difficult or even impossible. Exaggerated penile curvature may be of congenital origin or may be caused by Peyronie's disease. Although the condition is benign and does not involve a health risk to the patient, it may cause serious problems in the patient's sex life and psychology.

[0006] The penis is an organ that has the ability to grow in size and become erect when a man is sexually aroused. This is possible due to its anatomical structure, which is formed in the center by a bundle of tissue called the corpus spongiosum, inside which the urethra passes, and on the sides by two bundles of tissue called the corpus cavernosum. These tissues have an internal space that fills with blood when a man is sexually aroused, making them grow in size and harder, allowing penetration and thus sexual intercourse. These tissues are coated from the outside by a malleable tissue (tecido complacente) called the tunica albuginea.

[0007] Peyronie's disease is most common in men over 40 years old, and is generally caused by microtrauma in the tunica albuginea, which may occur during the course of the patient's sexual life.In the healing process of these microtraumas, fibrous tissue may develop in the tunica albuginea in the form of plaques or nodules, which reduces the compliance of blood vessels in the fibrotic area.In the fibrotic area, this compliance is lost, so that when the penis is erect, the corpus cavernosum cannot fully expand, and the penis curves.

[0008] Over the years, surgeons who specialize in treating this problem have attempted to develop various techniques in an attempt to ameliorate the problem, however, most have had limited success.

[0009] One of the techniques used today is the so-called "plicatura", which consists in surgically reducing the side not affected by fibrosis, but this procedure has an undesirable effect for most men, since it reduces the size of the penis.

[0010] Another common technique consists of surgically removing the fibrous tissue and replacing it with some type of tissue graft, which can be (a) of the autograft type, in which tissue from one part of the body is transplanted to tissue from another part of the body of the same individual, (b) of the allogeneic, homogeneous or homogeneous graft type, in which the transplant is made with tissue from the body of another animal of the same species, but with a different genotype than the individual, or (c) of the xenograft type, in which the transplant is made between animals of different species.

[0011] If successful, these techniques are preferred by most patients, since they do not reduce the size of the penis. However, the tissue utilized for the graft must have some special properties, which are difficult to find in xenogeneic tissues, which are mainly available commercially. Among these properties, the main ones are that the tissue utilized for the graft must have a conformability similar to that of the tunica albuginea, otherwise the problem will not be solved; the tissue utilized for the graft must be biocompatible and must not cause allergic, inflammatory or rejection reactions; the tissue utilized for the graft must not undergo calcification; the tissue utilized for the graft must be able to allow the proliferation of the patient's cells on it and promote biological integration; and the tissue utilized for the graft must not sustain shrinkage after surgery, otherwise the problem will not be solved.

[0012] All of these properties are absent in commercially available xenografts, which at least in part justify the low success rates achieved today with this type of procedure.

[0013] One of the most commonly available types of xenografts on the market is glutaraldehyde-preserved bovine pericardial grafts. This type of graft has been utilized with relative success in various types of surgery, primarily vascular surgery. However, attempts to use it in surgery to correct penile curvature have been unsuccessful. The primary reasons for the lack of success of this type of graft relate to the fact that glutaraldehyde-preserved bovine pericardial grafts are: - Not having the flexibility and flexibility necessary to accommodate the changes in size that occur in the penis; - Usually retracts after surgery and shortens the components in the repaired area, leading to failure of the procedure; - the patient's cells cannot grow on its surface and it behaves like a foreign body to the body; Although biocompatible, it is prone to inflammatory reactions and calcification, which is associated with tissue hardening and subsequent failure of the procedure.

[0014] One exemplary attempt to solve the above-cited problems is U.S. Patent No. 5,399,633, the entirety of which is incorporated herein by reference, which discloses a method for treating collagen connective tissue having a collagen fiber structure of donor animal origin such that after transplantation of said tissue into a recipient, said tissue contains anti-inflammatory agents and is tolerated by the recipient without immune rejection.

[0015] The method disclosed by US Pat. No. 5,399,633 is based on the use of a glutaraldehyde-free preservation solution containing physiological saline, a stabilizing solution, a solution consisting of polyglycol, salt, a phosphate buffer solution and an oxidizing agent. Summary of the Invention

[0016] However, this solution is still not sufficient to achieve a tissue that is adequately preserved to be acceptable to the human body, because it has been found to be ineffective and insufficient to achieve a tissue that is adequately preserved to be acceptable to the human body, for example, as tissue for tunica albuginea in correcting penile curvature.

[0017] Thus, as discussed above, there remains a need, and this is the primary object of the present invention, for improved methods for processing collagen connective tissue from animal donor sources so that the collagen fibers can be transplanted into a recipient without any immune and / or inflammatory rejection response.

[0018] (ii) Facial reconstruction: Various oral and maxillofacial reconstruction techniques currently practiced include mucosal bone marrow patches, bone replacement with grafts, guided bone regeneration and guided tissue regeneration using bone substitutes and barriers such as resorbable and non-resorbable membranes.

[0019] Although there is no consensus regarding the ideal material for the graft, the choice is guided by several important criteria: the material must be easy to obtain, available in sufficient quantities, compatible, and able to resist infection and resorption. In addition, it must be inert, resistant, easy to sterilize, and not interfere with the function of other structures.

[0020] The scientific literature has demonstrated that autologous cartilage has been generally successful due to its low metabolism and lack of side effects, however most autologous grafts suffer from limitations such as shedding, difficulty in handling, difficulty in delineating the graft, rate of resorption and shrinkage, in addition to donor site morbidity.

[0021] For the use of biomaterials to be successful, they must be biologically acceptable, i.e., they must not cause local or systemic damage and must not be toxic, carcinogenic, or radioactive. Furthermore, an ideal biomaterial must be manufacturable, sterilizable, and stable during its application or implantation. Studies have reported that the use of bioabsorbable materials has advantages over non-absorbable materials, because they avoid problems of migration, extrusion, and delayed infection.

[0022] The most commonly used biomaterials in BMF surgery are absorbable collagen membranes. Despite their good performance, they may show some limitations in certain cases, such as smaller than expected thickness, resistance to tension, volume and size, etc. In such cases, the use of autologous myocardial membrane patches is often adopted.

[0023] It is therefore desirable to provide a thicker biomaterial, with greater size and resistance to tension, that can be advantageously utilized in facial reconstruction to avoid the problems mentioned above and provide results not yet achieved using other materials from the prior art. [Prior art documents] [Patent documents]

[0024] [Patent Document 1] U.S. Pat. No. 9,205,172 [Patent Document 2] Brazilian Patent No. PI0814523-7 [Patent Document 3] U.S. Patent No. 7,008,763 DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0025] The method for producing preserved collagenous connective tissue from an animal donor source according to the present invention comprises: a) trimming collagen connective tissue in a buffer solution after removal from a donor; b) washing the connective tissue with the same buffer solution; c) stabilizing the tissue with a 50% ethanol solution; d) treating the tissue with a solution of polyethylene glycol; e) washing and storing the tissue in 50% ethanol; f) 50% ethanol solution and 1.5% hydrogen peroxide (H 2 O 2 sterilizing the tissue with g) storing the tissue in a 50% ethanol solution and indomethacin; Includes.

[0026] More specifically, the buffer solution of the present invention is sodium phosphate monobasic (NaH 2 PO 4 ) 0.1% and dibasic sodium phosphate (Na 2 HPO 4 The preferred washing solution is a mixture of phosphate buffer solution consisting of 0.6% phosphate (NaCl) and 0.9% sodium chloride (NaCl) solution, the pH of which should be physiological, preferably in the range of 7.3 to 7.5, and the temperature in the range of 5° C. to 15° C. More preferably, about 500 ml of buffer solution is used to wash each piece of collagen-connected tissue, and about 56 grams / liter of buffer solution is used.

[0027] Stabilization of collagen connective tissue should be carried out using ethanol 50% at temperatures ranging from 2 ° C to 10 ° C.

[0028] The stabilized tissue was then soaked in a 5% aqueous solution of 6% polyethylene glycol, 32% sodium chloride (NaCl), 13% phosphate buffer solution, and hydrogen peroxide (H 2 O 2 ) Treat with a solution of polyethylene glycol containing 2 volume % of the mixture and control the temperature within the range of 2°C to 8°C.

[0029] The tissue thus stabilized is then washed and stored in a solution of 50% ethanol at a temperature of about 25° C., followed by a 50% solution of ethanol and hydrogen peroxide (H 2 O 2 ) Sterilize with a 1.5% solution.

[0030] Finally, the collagen connective tissue is stored ready for use for up to two years in a solution of 50% by volume ethanol and approximately 0.05% by volume indomethacin.

[0031] The washing steps with a buffer solution, stabilization with a 50% ethanol solution and washing and storage of the collagen-bound tissue with a 50% ethanol solution are advantageously repeated 3 to 6 times, preferably the washing step of the tissue is repeated 5 to 6 times, the stabilization is repeated 3 times and the washing and storage steps are repeated 3 to 5 times.

[0032] Furthermore, in the steps of stabilization, treatment and sterilization of the collagen connective tissue, the immersion time varies from 24 to 96 hours, with the stabilization step being preferably performed for 24 hours per iteration, the treatment step being performed for 96 hours, and the sterilization step being performed for 24 hours. EXAMPLES

[0033] (Application of the present invention) Penile curvature: The following example describes a testing program evaluating graft implants comprising the aldehyde-free bovine pericardium of the present invention in a penile curvature correction procedure and reports subjective patient satisfaction ratings of the correction.

[0034] A total of 15 patients underwent surgery, of which 13 had Peyronie's disease and 2 had congenital penile curvature. The surgeries were performed at the Department of Urology, Faculty of Medical Sciences, Pedro Ernessto University Hospital, State University of Rio de Janeiro, between April 2019 and April 2020. The ages of the patients ranged from 18 to 72 years. Of the patients with Peyronie's disease, three had previously received an associated semi-rigid penile prosthetic implant.

[0035] Of those surveyed, 11 had already completed the 12-month postoperative monitoring period as per the study protocol, and seven of them were re-evaluated with regard to satisfaction with surgery, correction of curvature, presence of surgical complications, erectile quality, and penile length.

[0036] A scale of 1 to 5 was used for the subjective assessment of the individual's satisfaction with the correction of the curvature. Four patients (57%) stated that they were satisfied (a scale of 4 / 5) and three patients (43%) stated that they were very satisfied (a scale of 5 / 5). Satisfactory correction of the penile curvature was obtained in 89% of cases, and only one patient who presented with complex penile curvature in two axes still had significant penile curvature in one of the axes (approximately 60 degrees). Nevertheless, this patient reported no difficulties with penile intercourse. In the seven patients reevaluated so far, no significant complications according to the Clavien Dindo scale were identified. One of the surveyed (11%) showed poor erectile quality after the procedure, as assessed by the International Index of Erectile Function (IIEF). Six of the seven patients evaluated (86%) had an increase in penile length, and this increase in length varied from 0.7 cm to 3.0 cm. One patient had a decrease in penile length of 0.5 cm.

[0037] In a preliminary safety analysis with an estimated 46% of the total sample, it can be concluded that patients were subjectively satisfied with an acceptable success rate in correcting the penile curvature and without complications that sometimes prevent the resumption of studies of aldehyde-free pericardial graft implants in patients with penile curvature due to Peyronie's disease or congenital curvatures.

[0038] Beyond the excellent results obtained with the method for producing preserved collagen connective tissue from an animal donor source according to the invention, it has become possible to note unexpected features in addition to the reduced anti-inflammatory response, such as: - allowing the growth of the patient's own structural cells and promoting biological integration in such a way that the graft begins to show functional and biological characteristics similar to the transplanted tissue, such as vascularization, innervation and incorporation of collagen cells and elastin, to the extent that after a period of time the transplanted tissue can no longer be distinguished from the native tissue; The tissue exhibits elasticity, flexibility and malleability, even allowing for graft growth if necessary to accommodate the patient's ongoing anatomical changes; Unlike state-of-the-art implants treated with glutaraldehyde, the implants produced by the method of the present invention do not regress after surgery. It is.

[0039] Facial reconstruction: A study of facial reconstruction with grafts was carried out, using absorbable collagen biomaterial and aldehyde-free compounds produced, preserved and stabilized according to the method described by the present invention. Nine patients aged 18 years or older underwent facial reconstruction surgery at the Santa Casa de Misericordia Hospital School of Dentistry, Pocos de Caldas (MG) between March and November 2021. The study was submitted to and approved by the Ethics Committee of the institution. All patients approved and signed informed consent.

[0040] The implant investigated was aldehyde-free bovine pericardium manufactured by Labcor Laboratorios Ltda (Contagene, MG) and produced by the method described in this invention. The product was produced by a non-aldehyde technique.

[0041] Case 1: The first study involving preserved non-aldehydic collagen connective tissue was performed on a 36-year-old female patient with a history of ankylosis in the left (L) temporomandibular joint (ATM) associated with dentofacial deformity. The patient had three surgical procedures, including one procedure to correct mandibular distraction and the other for the release of the L ATM by implantation of a silicone prosthesis and osteotomy of the head of the L mandible. The patient had scarring from previous surgery, severe restriction of mouth opening, impossibility of oral hygiene with multiple caries, advanced periodontal disease, pinched teeth in the mandible, and cyst-like lesions in the mandible and maxilla. The patient also had an infection with an abscess on the L side, where the prosthesis was exposed.

[0042] The patient underwent surgery, under general anesthesia, for the removal of all teeth in the mandible and maxilla, and the installation of maxillary and mandibular implants in the All-on-4 system. After the removal of the maxillary and mandibular lesions and the installation of the implants, the empty spaces were filled with biomaterial. An Al Kayat access was made from the outside. The prosthesis was extracted, the tissue derived from the chronic inflammatory process was removed, and the articular bone surface was exposed. An aldehyde-free bovine pericardial graft was installed as if it were a temporal patch, placed between the remaining head of the mandible and the glenoid fossa, and fixed under tension with absorbable threads.

[0043] A preauricular incision was made, partially following the fistula present in this area. Plane expansion required utmost care due to the presence of a silicone appliance interposed between the tissues from the L-temporomandibular joint to the ear area, as well as changes in the local anatomy and local tissues due to the persistence of an old infection / inflammatory process. Accessing the joint through the "channel" left by the reaction to the silicone appliance, we found partly fibrous and partly granulomatous tissue in the cavity of the former L-ATM. This material acted as a barrier between the remaining part of the head of the L-mandible and the glenoid cavity (mandibular fossa).

[0044] Removal of the brace required mechanical cleaning of the site, exposing the articular bone surface or its remains. To avoid local inflammation and / or healing processes, and the resulting loss of motion on that side, we considered using a left temporalis fascia patch. However, the local fibrosis and properties altered by chronic infection processes made it impossible to create a satisfactory patch. For this reason, an aldehyde-free bovine pericardial graft was used, due to properties similar to patches of this nature, resistance and ability to conform to the area, as well as being fastened by local sutures.

[0045] An aldehyde-free bovine pericardial graft was used for the interposition between the bone surfaces and for the closure of the fistula. The graft was sutured to the adjacent tissue under constant tension and therefore remained immobile. Postoperative tomography showed a satisfactory position of the graft, and one week after the surgical procedure, the patient showed an improvement in the previously limited opening and closing of the mouth. The patient did not show any inflammatory reaction or rejection to the graft, even after six months of monitoring, thus demonstrating the full feasibility of using preserved non-aldehyde collagen connective tissue grafts for facial reconstruction.

[0046] Case 2: The second study with preserved non-aldehyde collagen connective tissue was performed on a 69-year-old female patient with a history of multiple facial fractures over at least 20 years, the presence of a silicone prosthesis dislodged from the floor of the orbit, and an infectious process in the L orbit with a fistula in the medial part of the L lower eyelid. The prosthesis created pressure on the globe, resulting in diplopia and "blurred vision". The correction was performed under general anesthesia through the L infraorbital access. After the prosthesis was withdrawn, the bone gap at the lower edge of the L orbit was adjusted using a titanium miniplaque of about 1.5 mm and fixed with two screws of about 1.5 mm in diameter. A preserved non-aldehyde collagen connective tissue graft was received in the floor of the orbit to occupy the space resulting from the absence of the prosthesis (which occupies the area and preserves the eye) and to correct the orbital deformity. The graft was fixed to the plaque and adjacent tissue, and the fistula was obliterated.

[0047] The patient already had a silicone brace on the L face, which caused a fistula in the inner area of ​​the lower eyelid, with a part of the brace coming out and compressing the eyeball. In addition to the pain and diplopia reported by the patient, a chronic infection / inflammatory process caused ongoing secretions in the L eye. The option was then taken to perform an L infraorbital incision with a compression line under the eyelid, resulting in a wider access, preserving the eyelid and allowing removal of the brace. After removal of the brace, sloughing of the L eyeball was noted, which, in addition to the bone space at the lower edge of the L orbit, had invaded the area of ​​the maxillary sinus, which allowed said sloughing. Careful tissue stripping exposed the bone remnants from the floor of the L orbit, also restricting the space that originally belonged to the L maxillary sinus. As decades have passed since the facial trauma and the overall bone structure has changed, a titanium plaque of about 1.5 mm was used to redo the lower edge.

[0048] Even with the restoration of the lower orbital rim, it was below the right (R) orbital rim. It was then necessary to interpose material on the floor of the orbit to elevate the eye and allow closure of the fistula and reconstruction of the eyelid. In cases where it was not possible to use a local patch and because of the need to avoid alloplastic material from previous chronic processes, non-aldehydic collagen connective tissue was used as a graft. This graft was sutured to the adjacent tissue and fixed to the plaque with absorbable threads. The graft was folded to give volume and elevate the L eye. Part of the graft was sutured to the internal tissue of the L eyelid, closing the fistula and partially reconstructing it. After 6 months of monitoring, the patient reported improvement in diplopia, correction of the fistula, and repositioning of the eye.

[0049] Case 3: A third study involving preserved non-aldehyde collagen connective tissue was performed on a 76-year-old female patient with periodontal damage in the area of ​​teeth 43, 44, and 45. Previously, the patient had undergone healing and biomaterial grafting as demonstrated by imaging studies. Examination also revealed a fistula in the area of ​​tooth 45 on the buccal side, and an enlargement on the lingual side in the area of ​​teeth 43, 44, and 45. On palpation, the areas showed a sclerotic appearance.

[0050] The surgical approach was buccal, preserving the mental foramen and its structures. After removing the above mentioned lesions and teeth, a lesion to the base was noticed that had a sclerotic appearance and was closely associated with the salivary duct in the area. The second lesion was carefully removed without damaging the duct, preparing the area for the graft.

[0051] The preserved collagen connective tissue was sutured to the tongue tissue isolating the canal. A bone graft was made with the biomaterial and the preserved collagen connective tissue was folded over the graft to act as a barrier. The preserved collagen connective tissue was sutured under tension to the graft.

[0052] The lesion was diagnosed as an ameloblastoma and was considered to be amenable to reconstruction with guided bone regeneration (GBR) to allow visualization and removal of the lesion. Preserved collagenous connective tissue was used due to the large extent of the repair performed, which required an extensive barrier that is not commercially available. The choice of preserved collagenous connective tissue was also based on the possibility of handling the membrane and fixing it to the local tissue. A slight exposure of the preserved collagenous connective tissue was observed, but in the postoperative period this did not in any way impair the results.

[0053] Case 4: A fourth study with preserved non-aldehyde collagen connective tissue was performed on a 36-year-old male patient with two missing mandibular central incisors. During surgery for jaw implantation, the preserved collagen connective tissue was utilized as a barrier for bone grafts and as a connective tissue substitute to improve gingival recesses in the mandibular incisors, ensuring the possibility of future rehabilitation with implants.

[0054] Medical literature indicates the use of guided bone regeneration (GBR) and connective grafts due to the patient's thin gingival profile. Therefore, this case required a bone graft, a barrier for the graft, and a connective tissue graft. Due to these requirements, as well as the extent of the defect, preserved collagen connective tissue was selected to act as a connective tissue barrier and replacement and improve the gingival profile.

[0055] The bone bed was exposed and prepared for the graft, and after implantation of the bone graft, the preserved collagen connective tissue was sutured to the buccal periosteum, folded over the bone graft and sutured to the tissue in the lingual area. The results two months after the procedure were completely satisfactory. In the six-month monitoring period, the alveolar margin and gingival profile were completely restored.

[0056] Case 5: The fifth study, involving preserved non-aldehyde collagen connective tissue, was performed on a 32-year-old female patient who had lost 44 teeth and developed buccal dehiscence, requiring rehabilitation with implants.

[0057] Because the patient objected to removing connective tissue from the second surgical site, preserved collagen connective tissue from the buccal side was used to obtain a gingival profile. The preserved collagen connective tissue was used as a tissue graft and secured by sutures. Complete correction was observed one month after the procedure.

[0058] Case 6: The sixth study, involving preserved non-aldehyde collagen connective tissue, was performed on a 62-year-old female patient with 34 root fractured teeth and loss of cheekbone. After removal of the residual root, an implant was placed in the site. Bone loss was offset by the biomaterial graft, and the preserved collagen connective tissue acted as a barrier to prevent gingival recession.

[0059] An immediate loaded implant was placed in the area of ​​the L mandibular premolar. Due to buccal bone loss, the option of guided bone regeneration (GBR) was chosen. However, the soft tissue was insufficient to cover the bone graft, as the fracture of the root and a chronic inflammatory process in the area had caused gingival recession. The option of preserved collagen connective tissue was chosen, as this acts as a barrier for bone regeneration and allows the acquisition of gingival tissue. The result 2 months after the prosthesis was placed on the implant was good.

[0060] Case 7: The seventh study with preserved non-aldehyde collagen connective tissue was performed on a 37-year-old female patient, a smoker, with poor oral hygiene, destruction of various crowns of her remaining teeth, and periodontal disease. She had previously undergone extraction of multiple teeth in both the mandible and maxilla for the placement of cone morse-type dental implants with the intention of rehabilitation with protocol-type prostheses in both dental arches.

[0061] The loss of tissue reproduction in the maxilla to remove periodontal disease-related damage prevented the closure of the patch. A loss of volume in the maxilla was also noted. The option of using preserved collagen connective tissue (instead of bone graft) was adopted to obtain tissue in the vicinity of the implant, avoid possible recesses, and provide oral volume. The preserved collagen connective tissue was divided into a right fragment and a separate left fragment, which were adapted to the prosthetic components (mini-pillars). The preserved collagen connective tissue was therefore perforated at the points corresponding to the mini-pillars and adjusted at its base, at the junction between the mini-pillars and the implant. The preserved collagen connective tissue was placed over the maxillary zygomatic tissue as if it were periosteum. The sutures were made of non-absorbable filaments and were removed on the 15th day.

[0062] Another reason for choosing preserved collagen connective tissue was the need to remove areas surrounding the patient's teeth due to periodontal disease and smoking, which resulted in insufficient patch coverage adjacent to the implant. Preserved collagen connective tissue also allowed for the acquisition of maxillary buccal tissue, improving the acute profile of the prosthesis and upper lip posture.

[0063] Case 8: The eighth study, involving preserved non-aldehyde collagen connective tissue, was performed on a 59-year-old male patient with a mandibular lesion suggestive of a mandibular cyst formed after tooth extraction in order to use a completely removable prosthesis.

[0064] The bone site of R revealed an exposed inferior alveolar plexus with a lesion anterior to R. After surgical removal of the lesion, a bone graft implant containing biomaterial was created. The preserved collagen connective tissue was used as a barrier in GBR and sutured tongue-and-cheek to be pulled over the bone graft. Diagnosis: ceratocistos. Imaging after the surgical procedure showed good resorption and recovery of the patient. The 6-month result showed that the lesion had completely healed.

[0065] Case 9: The ninth study involving preserved non-aldehyde collagen connective tissue was performed on a 44-year-old male patient with complications from orthopedic surgery. The patient had been referred by his orthodontist to have element 47 extracted, which had been compromised by a postoperative fracture from a first operation. However, as a result of the infectious process and complications of the first surgical procedure, the anchoring plaque was exposed in the area, which was removed in a second orthopedic operation.

[0066] Extraction of the tooth presented its challenges as the tissue in the area was not elastic and there was severe fibrosis from a previous infectious process. Preserved collagen connective tissue was used in the tooth removal to ensure that there would not be a worsening of plaque exposure and therefore a major tissue reaction resulting in bone exposure after extraction. Preserved collagen connective tissue was used to retrieve the remaining bone after extraction. The graft was sutured sublingually and doubled over the exposed bone and sutured again buccally, keeping tension over the covered area.

[0067] In this case, suture dehiscence was expected as the tissue lost its elasticity, which did occur, exposing areas of preserved collagen connective tissue, which was maintained by rinsing with 0.12% chlorhexidine.

[0068] As seen above, preserved collagen connective tissue and stabilized collagen connective tissue are preserved non-aldehyde biomaterials that exhibit broad versatility, deliver results beyond short-term and long-term expectations, and avoid the extraction of autologous patches in some situations, and thus can be utilized in a variety of restorative facial applications.

[0069] These unexpected properties demonstrate that the preserved collagen connective tissue from an animal donor source according to the present invention is an excellent choice for use as a graft in various types of surgery, such as cardiac surgery, ophthalmic surgery, neurosurgery, and surgery to correct penile curvature as reported above, among other applications in which these unexpected properties may be exhibited, since said connective tissue produced according to the present invention has all the characteristics that are considered to be ideal for these purposes.

[0070] Furthermore, the present invention also relates to collagen connective tissue preserved by the above method, its use in restorative surgery using collagen connective tissue in any region or organ of the human body, and to stabilized surgical kits comprising the preserved collagen connective tissue.

Claims

1. 1. A method for producing preserved collagen connective tissue, comprising: a) trimming the collagen connective tissue after removal from the donor in a buffer solution, the buffer solution being a phosphate buffer solution and a 0.9% solution of sodium chloride (NaCl), the temperature being in the range of 5°C to 15°C, and the pH being in the range of 7.3 to 7.5; b) washing the connective tissue with the buffer solution described in step a), the volume of the buffer solution being 500 ml per piece of pericardium; c) stabilizing the tissue with a 50% ethanol solution, the tissue stabilization temperature being in the range of 2° C. to 10° C.; d) treating the tissue with a polyethylene glycol solution and hydrogen peroxide, the tissue treatment temperature being in the range of 2°C to 8°C; e) washing and storing the tissue in 50% ethanol, the temperature of the 50% ethanol solution being approximately 25° C.; f) 50% ethanol solution and 1.5% hydrogen peroxide (H ２ O ２ sterilizing the tissue with g) storing the tissue for shipment in a 50% ethanol solution and indomethacin, the concentration of indomethacin being about 0.05% by volume; 23. A method for producing preserved collagen connective tissue, comprising:

2. The phosphate buffer solution is sodium phosphate monobasic (NaH ２ P.O. ４ ) 0.1% by volume and dibasic sodium phosphate (Na ２ H.P.O. ４ 2. The method for producing preserved collagen connective tissue according to claim 1, characterized in that the mixture consists of 0.6% by volume of collagen.

3. The polyethylene glycol solution of step d) is composed of 6% by volume of polyethylene glycol, 32% by volume of sodium chloride (NaCl), 13% by volume of phosphate buffer solution, and hydrogen peroxide (H ２ O ２ 2. The method for producing preserved collagen connective tissue according to claim 1, characterized in that the mixture is 2% by volume of collagen.

4. 2. A method for producing a preserved collagen connective tissue according to claim 1, characterized in that the steps of washing, stabilizing and cleaning the connective tissue are repeated 3 to 6 times, preferably 5 to 6 times for step b), 3 times for step c) and 3 to 5 times for step e).

5. 2. A method for producing a preserved collagenous connective tissue according to claim 1, characterized in that the steps of treatment, stabilization and sterilization of the connective tissue are carried out with an immersion time of 24-96 hours, preferably 24 hours for step c), 96 hours for step d) and 24 hours for step f) for each repetition.

6. A collagen connective tissue, comprising collagen connective tissue extracted from a donor and preserved and stabilized according to the method of any one of claims 1 to 5.

7. 7. A kit for producing preserved collagen connective tissue, comprising the collagen connective tissue of claim 6 and a storage solution comprising a mixture of 50% by volume of ethanol and about 0.05% by volume of indomethacin.

8. Use of the collagen connective tissue according to claim 6, characterized in that it is intended for application to any area or organ of the human body.

9. Use of collagen connective tissue according to claim 8, characterized in that it is intended for application as a graft in cardiac surgery, ophthalmic surgery, neurosurgery and surgery for correcting parts of the human body.

10. 10. Use of collagen connective tissue according to claim 9, characterized in that it is intended for the correction of penile curvature.

11. Use of the collagen connective tissue according to claim 9, characterized in that it is for facial reconstruction purposes.

12. Use of collagen connective tissue according to claim 11, characterized in that it is intended to repair ankylosis in the temporomandibular joint (TMJ), to repair dentofacial deformities, to repair deformities resulting from facial fractures, and / or to suture and reconstruct tongue tissue as a form of barrier and supplement.

13. 13. Use of collagen connective tissue according to claim 12, characterized in that the reconstruction is carried out by guided bone regeneration (GBR).

14. Use of collagen connective tissue according to claim 11, characterized in that it is for facial bone lining.

15. Use of collagen connective tissue according to claim 11, characterized in that it is for obtaining cheek tissue of the jaw.