MXPA97004454A - Tissue engineering - Google Patents

Abstract

A method for the augmentation or reconstruction of breast tissue, comprising implanting an effective amount of dissociated cells selected from the group consisting of mesenchymal cells, myocytes, chondrocytes, adipocytes, fibromyoblasts, ectodermal cells and nerve cells, to form a tissue of the

Description

FABRIC ENGINEERING OF THE BREAST BACKGROUND OF THE INVENTION The present invention is found, generally in the field of breast tissue reconstruction and augmentation. The breasts, or mammary glands, are modified sweat glands that attach to the internal muscle of the anterior chest wall through a layer of connective tissue. Internally, each mammary gland consists of 15 to 25 lobes, separated by dense connective tissue formed, mainly by fibroblasts and several bundles of collagen fibers, and adipose tissue that contains adipose cells (fat) that are held together by the reticular and collagen fibers. Within each lobe there is a lactiferous duct that branches extensively. At the ends of the smaller branches are glandular epithelial cells (alveolar cells) that synthesize and secrete milk into the canal system. The ducts, which are composed of a cuboidal epithelium and single column, and the alveolar cells are inside a loose connective tissue containing collagen fibers and fibroblasts, lymphocytes and plasma cells that secrete in unoglobulin A in milk, which it confers passive immunity to the newborn. Just outside the alveolar cells and the epithelial duct are the myoepithelial cells that respond to a neural and hormonal stimulus when contracting and expelling the milk. Each lactiferous duct opens up on a breast surface through the skin covering the nipple. Breast surgery can be classified broadly, as cosmetic and therapeutic. Cosmetic surgery includes augmentation, for example, using implants; reduction; and reconstruction. Therapeutic surgery, which is the primary treatment for most cancers, includes radical surgery that includes the removal of all of the tissue anterior to the chest wall and lymph nodes and veins that extend into the head and neck; lumpectomy, which involves only a small portion of the breast and laser surgery for the destruction of small tissue regions. Reconstructive surgery and the use of implants is often combined with radical breast surgery. The mastectomy involves the removal of the breast, both in the pectoralis major and minor, and the lymph nodes. Each year more than 250,000 breast reconstruction procedures are performed. Women who suffer from breast cancer, congenital defects or damage resulting from trauma have very few alternatives for reconstruction. Breast reconstruction is often used at the time or shortly after a mastectomy for cancer. Reconstruction procedures often include moving parts of the vascularized skin with the underlying connective and adipose tissue of a region of the body, such as the buttocks or the abdominal region, to the breast region. The surgery also uses breast implants for reconstruction. There are numerous surgical methods of breast reconstruction, including tissue expansion followed by silicone implantation, latissimus dorsi skin flaps, transverse abdominal pedicle myocutaneous flap (TRAM according to its international abbreviation), free TRAM flap and flap free glute Frequently, complete reconstruction requires numerous additional procedures to mastectomy and primary reconstruction. The procedures include tissue dilator exchanged for permanent implants, reconstruction of the nipple, reconstruction revision and mastopexy / reduction. Unfortunately, silicone prostheses, which are used for reconstruction and augmentations, have caused numerous medical complications. It would be desirable to have an alternative material for implantation. Even with the methods of reconstructive surgery that are currently used, it is extremely difficult to achieve a tissue that looks and feels normal, particularly when an extensive removal of associated muscle tissue is done.

Therefore, it is an object of the present invention to provide methods and compositions for the reconstruction and augmentation of breast tissue. Yet, it is another object of the present invention to provide methods and materials to provide a breast structure that is made of tissue, not foreign material such as silicone, and which has the appearance of normal tissue.

SUMMARY OF THE INVENTION The methods and compositions are described herein for the reconstruction and augmentation of breast tissue. The dissociated cells, preferably muscle cells, are implanted in combination with a suitable biodegradable polymer matrix to form a new tissue. There are two forms of matrices that can be used: a polymeric hydrogel formed of a material such as eg alginate having cells suspended therein, and a fibrous matrix having an intertitial space between about 100 and 300 microns. Preferably, polymeric materials are those that degrade in about one or two months, such as for example copolymers of polylactic acid-glycolic acid. These matrices can be sown before implantation or implanted, allowing vascularization, then seeded with cells. In a preferred embodiment, the structures of the cell matrix are implanted in combination with tissue dilator devices. While the matrix of the cell or cells that proliferate and form new tissue is implanted, the size of the dilator decreases, until it can be removed and the desired reconstruction or augmentation is obtained. Preferred cell types are muscle cells, although other types of mesenchymal cells, fibroblasts, chondrocytes and adipocytes can be used. Cells that are obtained from tissue, such as the lips, can be used for specialized applications such as the formation of a tissue of the nipple type. Other materials, such as for example bioactive molecules that improve the vascularization of the implanted tissue and / or which inhibit the growth of fibrotic tissue, can be implanted with the matrix to improve the development of a more normal tissue. The structures of the cell matrix can be implanted at the time of surgery to remove cancerous tissue from the breast, during a subsequent reconstruction surgery, or for a period of time, for example, weekly, if a series of injections are used. of cell-hydrogel suspensions to create the new tissue.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 is a schematic of the procedure of implanting dissociated cells in a polymeric matrix in the breast to augment breast tissue. Figure 2 is a schematic of a fibrous plaque implanted in the breast tissue with supports to provide support of surrounding tissue and skin and which allows the new tissue to form from the support followed by the injection of the cell-hydrogel suspension. Figures 3A, 3B and 3C are diagrams of the series of cell-hydrogel suspension injections followed by the implantation of a tissue dilator, with the tissue dilator decreasing in size each time the suspension is injected. Figure 3A is the tissue dilator maximally expanded; Figure 3B shows the fluid that is removed from the dilator to create a space between the adjoining tissue and the dilator, in which the cell-polymer suspension is injected; and Figure 3C shows the dilator deflated to the maximum, with the new tissue that is formed in the space concealed by the majority of the expanded tissue dilator as in Figure 1. Detailed description of the invention 1. Cells to be implanted The cells that are they are to be dissociated using standard techniques such as digestion with collagenase, trypsin or other protease solutions. The preferred cell types are mesenchymal cells, especially (muscle trunk cells), chondrocytes, adipocytes, fibromyoblast and ectodermal cells, including flexible and skin cells. In some cases, one may also wish to include nerve cells. The cells can be normal or genetically structured to provide additional or normal function. Preferably, the cells are cells derived from themselves, obtained by biopsy and expanded in culture, although cells from close relatives or from other donors of the same species can be used with appropriate immunosuppression. Immunologically inert cells, such as embryonic cells, stem cells, and genetically engineered cells can also be used to avoid the need for immunosuppression. The methods and drugs for immunosuppression are known to those skilled in the art of transplantation. A preferred compound is cyclosporin used in the recommended doses. In the preferred embodiment, smooth muscle or skeletal cells are obtained by biopsy and expanded in culture for subsequent implantation. Skeletal cells such as smooth can be easily obtained through a biopsy in any part of the body, for example, main muscle biopsies can be easily obtained from an arm, forearm or lower extremities, and smooth muscles can be obtained from a area adjacent to the subcutaneous tissue throughout the body. To obtain some type of muscle tissue, the area where the biopsy will be performed can be locally anesthetized with a small amount of lidocaine injected subcutaneously. Alternatively, a small patch of lidocaine gelatin can be applied to the area where the biopsy is to be performed and let it act for a period of 5 to 20 minutes, before obtaining a sample for the biopsy. The biopsy can be easily obtained with the use of a needle for biopsy, a fast-acting needle with which the procedure is performed in an extremely simple and almost painless manner. With the addition of an anesthetic agent, the procedure will not be painful at all. This small nucleus of biopsy either of the major muscles or of the smooth muscles can then be transferred to a medium consisting of a buffered saline phosphate substance. Then, the sample of the biopsy is transferred to a laboratory where the muscle can be grown using the technique explained, where the muscle is divided into several pieces that adhere to the culture dish, and the serum containing the muscle is added. medium. Alternatively, the muscle biopsy can be enzymatically digested with agents such as trypsin and dispersed cells in a culture dish with any of the means used routinely. After the cells expand within the culture dish, the cells can be easily passaged using the normal technique until an adequate number of cells is achieved.

II. Device fabrication Three main s of matrices can be used to create new tissue or augmentation tissues. The term "bioerodible," or "biodegradable," as used herein, refers to materials that are degraded enzymatically or chemically in vivo in a simpler chemical species.

Hydrogel polymer solutions In one embodiment, polymers that can form ionic hydrogels that are malleable are used to support the cells. Injection the suspension of the cells in a polymer solution can be carried out to improve the reproducibility of the cell seeded through a device, to protect the cells from shearing or pressure forces that induce necrosis, or to help in the definition of a spatial placement of the delivered cell. The injectable polymer can be used to deliver the cells and promote the formation of new tissues without the use of any matrix. In a preferred embodiment, the hydrogel is produced by the crosslinking of the ionic salt of a polymer with ions, whose strength increases both by increasing the concentrations of ions and the polymer. The polymer solution is mixed with the cells to be implanted to form a suspension, which is then injected directly into the patient before the solution hardens. The suspension subsequently hardens after a short period of time, due to the presence in vivo of physiological concentrations of ions such as calcium in the case where the polymer is a polysaccharide such as alginate.

Polymers The polymeric material that is mixed with the cells for the implant in the body must form a hydrogel. The hydrogel is defined as a substance formed when the organic polymer (natural or synthetic) is crosslinked via covalent, ionic or hydrogenic bonds to create an open grid structure that traps water molecules to form a gel. Examples of the materials that can be used to form hydrogel include polysaccharides such as for example alginate, polyphosphazenes, and polyacrylates such as for example hydroxyethyl methacrylate (HEMA according to its international abbreviation), which is ionically crosslinked, or polymer blocks such as for example Pluronics ® or Tetronics®, polyethylene oxide-polypropylene glycol block copolymers that are crosslinked by temperature or pH, respectively. Other materials include proteins such as fibrin, polymers such as polyvinylpyrrolidone, hyaluronic acid and collagen. In general, these polymers are at least partially soluble in aqueous solutions, such as for example water, buffered salt solutions, or aqueous solutions of alcohol, having charged side groups, or a monovalent ionic salt thereof. Examples of polymers with acidic side groups that can be reacted with cations are poly (phosphazenes), poly (acrylic acids), poly (methacrylic acids), copolymers of acrylic acid and methacrylic acid, poly (vinyl acetate), and sulfonate polymers , such as, for example, sulfonated polystyrene. Copolymers having acidic side groups formed by the reaction of acrylic or methacrylic acid and vinyl ether monomers or polymers can also be used. Examples of acidic groups are carboxylic acid groups, sulfonic acid group, halogenated alcohol groups (preferably fluorinated), phenolic OH groups, and acidic OH groups.

Examples of polymers with basic side groups that can be reacted with anions are poly (vinyl amines), poly (vinyl pyridine), poly (vinyl imidazole) and some substituted polyphosphates. The ammonium or quaternary salt of the polymers can also be formed from spinal nitrogens or imino crown groups. Examples of basic side groups are amino and imino groups. Alginate can be ionically crosslinked with divalent cations, in water, at room temperature, to form a hydrogel matrix. Due to these conditions, alginate has been the most commonly used polymer for the encapsulation of the hybridoma cell, as described, for example in U.S. Patent No. 4,352,883 issued to Lim. In the Lim process, an aqueous solution containing the biological materials to be encapsulated in a water-soluble polymer solution is suspended, the suspension is formed into droplets that are formed into microcapsules by contact with multivalent cations, then the surface of the microcapsules are crosslinked with polyamino acids to form a semipermeable membrane surrounding the encapsulated materials. Polyphosphazenes are polymers with structure in the form of vertebral column consisting of nitrogen and phosphorus separated alternating simple link and double bonds.

Each phosphorus atom is linked covalently to the side chains ("R"). The unit repeated in polyphosphazenes has a general structure: R - (- P - N -) n R where n is an integrated The polyphosphazene suitable for crosslinking has a majority of side chain groups which are acidic and capable of forming salt bridges with di- or trivalent cations. Preferred examples of the laterale acidic groups are carboxylic acid groups and sulfonyl acid groups. Hydrolytically stable polyphosphazenes are formed from monomers having carboxylic acid side groups that are crosslinked by divalent or trivalent cations such as Ca 2+ or Al 3+. The polymers can be synthesized which are degraded by hydrolysis by incorporating imidazole-containing monomers, amino acid esters, or glycerol side groups. For example, polyanionic poly [bis (carboxylatophenoxy)] (PCPP by its international abbreviation), which is crosslinked with multivalent cations dissolved in an aqueous medium at room temperature or below to form hydrogel matrices, can be synthesized. Bioerodible polyphosphazene has at least two different types of side chains, acidic side groups capable of forming salt bridges with multivalent cations, and side groups which are hydrolyzed under in vivo conditions, for example imidazole groups, amino acid esters, glycerol and glucosyl. The term bioerodible or biodegradable, according to their use herein, refers to a polymer that dissolves or degrades in a period that is acceptable in the desired application (usually in vivo therapy), less than about 5 years and more preferably less than one year, once exposed to a physiological solution of pH 6-8 having a temperature between about 25 ° C and 38 ° C. The hydrolysis of the side chain results in the erosion of the polymer. Examples of acid hydrolyzed side chains imidizole and substituted amino esters in which the group is attached to the phosphorus atom via an amino linkage (polyphosphate polymers in which both R groups which are linked in this manner, are known as polyaminophosphazenes) . For polyimidazolaphosphazene, some of the nR "groups in the backbone of polyphosphazene are imidazole rings, attached to the phosphorus in the backbone through the nitrogen ring atom.Other" R "groups can be organic wastes that do not participate in the hydrolysis, such as for example methyl phenoxy groups or other groups which are presented in the scientific document of Allcock et al., Macromolecule (Macromolecule) 10: 824-830 (1977) .The methods for the synthesis and analysis of various types of polyphosphazenes are described by Allcock, HR, and collaborators in Inorg. Chem. (Inorganic Chemistry) 11, 2584 (1072); Allcock, and collaborators in Macromolecules (Macromolecules) 16, 715 (1983); Allcock, and collaborators in Macromolecules (Macromolecules) 19, 1508 (1983); Allcock, and collaborators in Bio aterials (Biomaterials) 21, 1980 (1988); Allcock, and collaborators in Inorg. Chem. (Inorganic Chemistry) 21 (2), 515-521 (1982); Allcock, and collaborators in Macromolecules (Macromolecules) 22.75 (1989); U.S. Patent Nos. 4,440,921; 4,495,174 and 4,880,662 granted to Allcock et al; U.S. Patent No. 4,946,938 issued to Magill et al .; and Grolleman et al. J. Controlled Reléase (Controlled Release of J.) 3, 143 (1986), the teachings of which are specifically incorporated herein by reference. The methods for the synthesis of other polymers that were described above are known to those skilled in the art. See, for example, Concise Encyclopedia of Polymer Science and Polymeric Amines and Ammonium Salts (Polymer amines and ammonium salts), E '. Goethals, editor (Pergamen Press, Elmsford, NY 1980). Many polymers, such as poly (acrylic acid), are commercially available. The water-soluble polymer with charged side groups is crosslinked by reacting the polymer with an aqueous solution containing multivalent ions of opposite charge, either multivalent cations if the polymer has acidic side groups or multivalent anions if the polymer has basic side groups . The preferred cations for the crosslinking of the polymers with the acidic side groups to form a hydrogel are bivalent and trivalent cations such as copper, calcium, aluminum, magnesium, strontium, barium and tin, although organic functional cations can also be used. -, tri- or tetra such as, for example, alkylammonium salts, such as R3N + - \ / \ / \ / - + NR3. The aqueous solutions of the salts of these cations are added to the polymers to form hydrogels and highly swollen, smooth membranes. The greater concentration of cations, or the greater valence, the greater the degree of crosslinking of the polymer. Concentrations from as low as 0.005 M have been shown to crosslink with the polymer. The highest concentrations are limited by the solubility of the salt. Preferred anions for crosslinking the polymers to form a hydrogel are divalent and trivalent anions such as for example dicarboxylic acids with low molecular weight, for example, terephthalic acid, sulfate ions and carbonate ions. The aqueous solutions of the salts of these anions are added to the polymers to form highly swollen, soft hydrogels and membranes, as described in connection with the cations. A variety of polycations can be used to give complexity and, therefore, to stabilize the polymer hydrogel in a semipermeable surface membrane. Examples of the materials that can be used include polymers having basic respective groups such as for example amines or imine groups, having a preferred molecular weight between 3,000 and 100,000, such as polyethylene imine and polylysine. These are commercially available. A polycation is poly (L-lysine), examples of synthetic polyamines are: polyethyleneimine, poly (vinylamine) and poly (allylamine). These are also natural polycations such as polysaccharides, chitosan. Polyanions that can be used to form a semipermeable membrane by reaction with the basic surface groups in the polymer hydrogel include polymers and copolymers of acrylic acid, methacrylic acid, and other acrylic acid derivatives, polymers with crown S03H groups such as for example sulfonated polystyrene, and polystyrene with carboxylic acid groups.

Method for making cell suspensions The polymer is dissolved in an aqueous solution, preferably a 0.1 M potassium phosphate solution, a physiological pH, at a concentration that forms a polymer hydrogel, for example, for alginate, of between 0.5 up to 2% by weight, preferably 1%, alginate. The isolated cells are suspended in the polymer solution at a concentration between 1 and 50 million cells per ml, more preferably between 10 and 20 million cells per ml.

Polymer matrix Matrix configuration For the construction, successful implantation and function of an organ, matrices must have a sufficient surface area and be exposed to nutrients in such a way that cell growth and differentiation and differentiation can occur before the regrowth of blood vessels followed by implantation. The time required for the successful implantation and the growth of the cells within the matrix are considerably reduced if the area in which the matrix is implanted is pre-vascularized. After the implant, the configuration must be allowed by diffusion of nutrients and waste of products and for a continuous regrowth of the blood vessels as the proliferation of the cell occurs. The organization of the tissue can be regulated by the microstructure of the matrix. Specific pore sizes and structures can be used to control the pattern and extend the regrowth of fibrovascular tissue from the host, as well as the organization of the implanted cells. The geometric and chemical surface of the matrix can be regulated to control adhesion, organization and function of the implanted cells or host cells. In a preferred embodiment, the matrix is formed of polymers having a fibros structure having sufficient intertidal space to allow free diffusion of nutrients and gases to cells attached to the surface of the matrix. The space or separation is typically on average from 100 to 300 microns, although closer separations can be used if the matrix is implanted, the blood vessels are allowed to infiltrate the matrix, then the cells are seeded into the matrix. According to the form of use herein, "fibrous" includes one or more interlacing fibers, multiple fibers in a woven or non-woven mesh and a device such as a sponge. The cells can be implanted after being sown in the matrix or they can also be injected into the already implanted matrix at the desired site. The latter has the advantage that the matrix can be used to pre-vascularize the site. In this case, the design and construction of the structure is of vital importance. The matrix must be flexible, non-toxic, with injectable pores formed for vascular regrowth. The pores should allow the vascular regrowth and the injection of the cells such as the muscle cells without damaging the cells or the patient. Generally, these are interconnected pores in the average of between approximately 100 to 300 microns. The matrix should be formed to maximize the surface area, to allow an adequate diffusion of nutrients and growth factors for the cells and allow the re-growth of the new blood vessels and the connective tissue. Currently, a porous structure that is resistant to compression for the implant, prevascularization, followed by seeding is preferred. In the embodiment where the matrix is prevascularized, it may be desirable to incorporate elements in the matrix to disperse the cells throughout the matrix, for example, using catheters that can be removed after sowing. The totality, or the external part, of the configuration of the matrix depends on the tissue that is reconstructed or increased. In most cases, the cell-matrix structure will be similar to the silicone implants used so far, which are essentially disks that deform due to the force of gravity to take the shape of a tear. The shape can also be obtained using supports, as will be described below, to impart strength to the mechanical forces and thereby produce the desired shape. The shape of the matrix per se will not have a disc shape, but it will appear to have a disk shape when the cells to be implanted are sown, or they will create the edge of a disc or tear shape after implantation. Polymers Both synthetic and natural polymers can be used to form the matrix, although synthetic polymers are preferred for reproducibility and controlled kinetic release. Synthetic polymers that can be used include bioerodible polymers such as poly (lactide) (PLA), poly (glycolic acid) (PGA), poly (lactide-co-glycolide) (PLGA), poly (caprolactone), polycarbonates, polyamides , polyanhydrides, polyamino acids, polyoxytes, polyacetals, polycyanoacrylates and degradable polyurethanes and non-erodible polymers such as polyacrylates, ethylene-vinyl acetate polymers and other cellulose acetates substituted by acyl and derivatives thereof, non-erodible polyurethanes, polystyrenes, polyvinyl, polyvinyl fluoride, poly (vinyl imidazole), chlorosulfonated polyolefins, polyethylene oxide, polyvinyl alcohol, Teflon® and nylon. Although non-degradable materials can be used to form the matrix or a portion of the matrix, they are not preferred. The preferred non-degradable materials for the implant of the matrix that is pre-vascularized prior to the implantation of the dissociated cells is a polyvinyl alcohol sponge, or alkylation and acylation thereof, including esters. The non-absorbent polyvinyl alcohol sponge is commercially available as Ivalon®, from Unipoint Industries. Methods for making this material are described in US Pat. Nos. 2,609,347 issued to ilson.; 2,653,917 granted to Ha mon, 2, 659,935 granted to Hammon; 2,664,366 granted to Wilson; 2,664,367 granted to Wilson and 2,846,407 granted to Wilson, the teachings of these are incorporated herein by reference. These materials are all commercially available. Examples of natural polymers include proteins such as for example albumin, collagen, synthetic polyamino acids and proamines, and polysaccharides such as for example algin, heparin, and other naturally biodegradable polymers of sugar units. The PLA, PGA and PLA / PGA copolymers are particularly useful for forming the biodegradable matrices. PLA polymers are usually prepared from cyclic esters of lactic acids. Both L (+) and D (+) forms of lactic acid can be used to prepare the PLA polymers, as well as the optically inactive DL lactic acid mixture of lactic acids D (-) and L (+). The methods for preparing polylactides are well documented in the patent literature. The following U.S. Patents, the teachings of which are incorporated herein by reference, describe in detail the appropriate polylactics, their properties and their preparation: 1,995,970 issued to Dorough; 2,703,316 granted to Schneider; 2,758,987 granted to Salzberg; 2,951,828 granted to Zeile; 2,676,945 granted to Higgins; and 2,683,136; 3,531,561 granted to Trehu. PGA is the homopolymers of glycolic acid (hydroxyacetic acid). In the conversion of glycolic acid to poly (glycolic acid), the glycolic acid is initially reacted therewith to form a cyclic ester glycolide, which in the presence of heat and a catalyst is converted to a linear chain polymer of high molecular weight. PGA polymers and their properties are described in detail in "Cyanamid Research Develops World's First Synthetic Absorbable Suture," Chemistry and Industry, 905 (1970). Erosion of the matrix is related to the molecular weights of PLA, PGA or PLA / PGA. The higher molecular weights, the average weight of molecular weights of 90,000 or greater, result in polymer matrices that retain their structural integrity for longer periods of time; while lower molecular weights, average molecular weights of 30,000 or less, result in both slower release matrices and shorter lives. A preferred material is poly (lactide-co-glycolide) (50:50), which degrades in about six weeks after implantation (between one and two months). All polymers for use in the matrix must cover the mechanical and chemical parameters necessary to provide adequate support for the cells with subsequent growth and proliferation. Polymers can be characterized, with respect to mechanical properties such as tensile strength using an Instron tester, for the molecular weight of a polymer by permeation chromatography (GPC), the glass transition temperature by differential scanning calorimetry (DSC according to its international abbreviation) and the link structure using infrared spectroscopy (IR), with respect to toxicology by means of the initial analysis tests involving the Ames assays and the in vitro teratogenicity assays, and the studies of Implants in animals for immunogenicity, inflammation, release and degradation studies. Polymer coatings In some embodiments, the binding of cells to polymers is improved by coating the polymers with compounds such as base membrane, agar, agarose, gelatin, gum arabic, collagen types 1, II. , III, IV and V, fibronectin, laminin, glycosaminoglycans, polyvinyl alcohol, mixtures thereof, and other hydrophilic binding materials and peptides known to those skilled in the cell culture art. A preferred material for coating the polymer matrix is polyvinyl alcohol or collagen. Stands In some embodiments it may be desirable to create an additional structure using devices provided for support, referred to herein as "mullions". These can be biodegradable or non-degradable polymers that are inserted to give a more defined shape than that obtained using the cell matrices, especially the suspensions of hydrogel cells. An analogy can be made with a corset, with the uprights acting as "props" to push the tissue around and the skin up and away from the implanted cells. In a preferred embodiment, the posts are implanted before or at the same time of the implantation of the cell matrix structure. The uprights are formed of a polymeric material of the same type that can be used to form the matrix, as mentioned above, having sufficient strength to withstand the necessary mechanical forces. Tissue Dilators Alternatively, or additionally, tissue dilators can be used to create additional space for the implantation of the cell matrix structures. Tissue dilators are commercially available and are routinely used for skin expansion, for example, before plastic surgery, as presented by Cohen J. Dermatol. Surg. Oncol. 19: 614-615, Bennet and Hirt J. Der atol. Surg. Oncol. 19: 1066-1073 (1993), Hammond et al., Plastic and Reconstructive Surgery 92 (2): 255-259 (1993), Walton and Bro n, Annals of Plastic Surgery (Annals of Surgery Plastics) 30 (2), 105-110 (February 1993), Kenna et al., Annals of Plastic Surgery 32, 346-349, the teachings of these are incorporated herein by reference. When the skin tightens for long periods of time, from weeks to months, it responds with a significant stretch. These are associated with metabolic activity and tissue growth. The generally accepted definition of tissue dilator is a device that resides beneath the surface of the skin that is used to stretch the skin. A spherical tissue dilator is a multidimensional dilator, typically applied by volumetric expansion of a subcutaneous space with an inflatable device. Alternatively, multiple materials can be implanted and the device shrunk or replaced by removing one or more of the bolus materials. The use of tissue dilators in breast reconstruction is well understood (see, for example, Hammon et al., 1993). Several different types of dilators have been designed or anatomically oriented to give a more natural contour to the reconstructed breast. The devices are commercially available, for example, from McGhan Medical Corporation, Santa Barbara, CA, Dow Corning-Wright, Arlington, TN and Mentor Corporation, Goleta, GA. It is important to remove the pressure from the implanted cells which can kill the cells. For example, in a preferred embodiment that is described in more detail below, the suspension of hydrogel cells is injected into the area where the tissue will be created. The space for the injection of the cell-polymer suspension is created by the implantation of a tissue dilator before injecting the cell-hydrogel suspension. The tissue dilator is inflated or expanded through the implantation of a desired number of modules, to maximize the space and skin required for tissue formation. As shown in detail in Figures 3A, 3B and 3C, each time the cell matrix is injected, the tissue dilator deflates or a module is removed to leave a space of an amount equivalent to the volume of the matrix of the cell. injected cells. Once the space is filled essentially with new tissue or suspension of the matrix cell, the tissue dilator is removed, using, in most cases, a local anesthetic and a minor incision. Additives to polymer matrices In some embodiments it may be desired to add bioactive molecules to the cells. A variety of bioactive molecules can be provided using the matrices described herein. These are generically mentioned in the present as "factors" or "bioactive factors". In the preferred embodiment, the bioactive factors are the growth factors, angiogenic factors, compounds that selectively inhibit tissue regrowth of fibroblasts as anti-inflammatories and compounds that selectively inhibit the growth and proliferation of transformed (cancerous) cells. These factors can be used to control the growth and function of the implanted cells, the regrowth of the blood vessels in tissue formation, and / or the arrangement and organization of fibrous tissue around the implant. Examples of growth factors include heparin binding growth factor (hbgf by its international abbreviation), alpha or beta growth transforming factor (TGFβ), fibroblast growth factor (FGF according to its international abbreviation), growth factor epidermal (TGF), vascular endothelial growth factor (VEGF), some of which are also antiogenic factors. Other factors include hormones such as insulin, glucagon and estrogen. In some embodiments it may be desirable to incorporate factors such as for example nerve growth factor (NGF) or muscle morphogenic factor (MMP). Steroidal anti-inflammatory drugs can be used to decrease inflammation in the implanted matrix, thereby decreasing the amount of fibroblast tissue that grows in the matrix.

When selective chemotherapeutic agents are available that do not inhibit the growth of normal cells, such as chemotherapeutic agents destined for antibodies, they can be incorporated into the matrix and used to inhibit any residual cancer cells that remain after mastectomy. . These factors are known to those skilled in the art and are commercially available or described in the literature. In vivo doses are calculated based on in vivo studies in cell culture; an effective dose is the dose that increases cell proliferation or its survival compared to controls, as will be described in more detail in the following examples. Preferably, the bioactive factors are incorporated between 1 and 30% by weight, although the factors can be incorporated at a percentage by weight between 0.01 and 95 percent by weight. Bioactive molecules can be incorporated into the matrix and released for a time by diffusion and / or degradation of the matrix, they can be suspended with the suspension of the cell, they can be incorporated into microspheres that are suspended with the cells or are attached to or incorporated within the matrix, or some combination of these. The microspheres would typically be formed from materials similar to those that form the matrix, selected for their release properties rather than for their structural properties. The release properties can also be determined by the physical characteristics and size of the microspheres. The desired microspheres and methods for their use in tissue generation are described in US Serial No. 08 / 358,235 by David J. Mooney, Robert S. Langer and Joseph P. Vacanti, entitled "Localized Delivery of Factors Enhacing Survival of Transplanted Cells "(Localized Delivery of Factors to Improve Survival of Transplanted Cells), co-filed with the United States Patent and Trademark Office on December 16, 1994, the teachings of which are incorporated herein . III. Methods for Implantation As discussed generally above, there are three methods that can be used to create new breast tissue. These can be used alone or in various combinations. Variations include the place of the cell, which may be hydrogel solution or a solid fibrous matrix, before implantation or introduced in series form after implantation to allow prevascularization of the matrix. The shape of the tissue structure can be regulated using a tissue dilator to create the desired space for tissue formation, and then in series form the dilator tissue is deflated while the cells of interest are provided in the newly created space . These allow the formed tissue to be predefined, and allow the serial introduction of the cells to form the new tissue. Alternatively, the preformed matrix can be implanted, allowing vascularization, then sowing with the dissociated cells that form the new tissue, preferably as the matrix degrades. The selection of an appropriate system depends on. degree of magnification required and determines whether the entire injection can be carried out at one time, or alternatively, performed in sequence form, to allow the formation of tissue with adequate vascularization before subsequent injections are applied. The selection of the cell type can be used to vary the texture of the implanted material, as well as its appearance. For example, cartilages can be used, if a more rigid implant is desired. In some embodiments it may be desirable to create softer tissue, for example, using adipocytes or other smooth tissue components. Figure 1 is a schematic of the process for the implantation of the dissociated cells 10 in a polymeric matrix 12 in the breast 14 to increase the breast tissue. The cells attached to the matrix 12, which originally have a disc shape but are deformed into tears when implanted.

As vascularization occurs and the matrix degrades, new tissue forms. Figure 2 is a schematic of a fibrous plate 20 implanted in the breast tissue 22 with uprights 24 to provide support for the tissue around and to the skin and allows the new tissue to be formed within the support after injection of the cell suspension hydrogel (not shown) Figures 3A, 3B and 3C are diagrams of the injections series of the hydrogel-cell suspension after implantation of the tissue dilator (Figure 3A), with the tissue dilator decreased in size each time the suspension is injected (Figure 3B), so that the new tissue is formed in the space left as the dilator decreases in its volume (Figure 3C).

Claims (18)

1. Rei iications 1. A method for breast tissue augmentation or reconstruction, comprising implanting an effective amount of dissociated cells selected from the group consisting of mesenchymal cells, myocytes, chondrocytes, adipocytes, fibromyoblasts, ectodermal cells and nerve cells, for form a breast tissue 2. A method according to claim 1, wherein the cells are smooth or major muscle cells. 3. The method according to claim 1, further comprising implanting the cells in combination with a matrix. 4. The method according to claim 3, wherein the matrix is a biocompatible and biodegradable hydrogel. 5. The method according to claim 3, wherein the matrix is implanted, vascularization is allowed and then sown with the cells. 6. The method according to claim 3, wherein the matrix is a fibrous and polymeric matrix. The method according to claim 3, wherein the matrix is formed from a biodegradable polymer. 8. The method according to claim 3, further comprising implanting uprights with the matrix supporting the surrounding tissue. 9. The method according to claim 1, further comprising implanting bioactive molecules selected from the group of molecules that improve vascularization, cell survival, proliferation or differentiation, inhibit growth in fibrotic tissue, inhibit growth of cancer cells and inhibits inflammation. 10. The method according to claim 1, further comprising implanting a tissue dilator before implanting the cells in combination with the matrix, then implanting the cells in combination with the matrix each time the dilator decreases in size. 11. A composition for augmenting or reconstructing breast tissue, comprising dissociated cells selected from the group consisting of mesenchymal cells, myocytes, chondrocytes, adipocytes, fibromyoblasts, ectodermal cells and nerve cells in combination with a fibrous and polymer matrix, wherein The combination of the cells and the matrix is effective for the augmentation or reconstruction of breast tissue. 12. The composition according to claim 11, wherein the cells are smooth or major muscle cells. The composition according to claim 11, wherein the fibrous matrix has a disc or tear configuration. The composition according to claim 11, further comprising stiles that support the surrounding tissue. 15. The composition according to claim 11, which further comprises bioactive molecules selected from the group of molecules that improve vascularization, cell survival, proliferation or differentiation, inhibit the growth into the fibrotic tissue, inhibit the growth of cancer cells and inhibit inflammation. 16. The composition according to claim 11, further comprising a tissue dilator. The composition according to claim 11, wherein the matrix is suitable for implantation and vascularization before seeding with the cells, characterized by means that disperse the cells throughout the matrix after implantation, and which are resistant to compression. 18. The composition according to claim 11, wherein the matrix is formed from biodegradable polymers. Summary of the Invention Methods and compositions are described herein for the reconstruction or augmentation of breast tissue. The dissociated cells, preferably muscle cells, are implanted in combination with a suitable biodegradable polymer matrix to form new tissue. There are two forms of matrices that can be used: a polymeric hydrogel formed of a material such as, for example, alginate that has cells suspended in it, and a fibrous matrix having an interstitial space between about 100 to 300 microns. Preferred polymeric materials are those that degrade in about one to two months, such as, for example, polylactic acid-glycolic acid polymers. The matrices can be sown before implantation or implanted, to allow vascularization, then sowed with the cells. In a preferred embodiment, the structures of the cell matrix are implanted in combination with the tissue expanding devices. As the cell matrix is implanted or the cells proliferate and form the tissue, the size of the expander expands, until it can be removed and the desired reconstruction or augmentation is obtained. The preferred cell types are muscle cells, although other cell types can be used such as mesenchymal cells, fibriblastos, chondrocytes and adipocytes. The cells obtained from the tissue, such as that of the lips, can be used for special applications such as, for example, the formation of the tissue of the nipple type. Other materials, such as, for example, bioactive molecules that improve the vascularization of the implanted tissue and / or inhibit the growth of fibrotic tissue, can be implanted with the nuance to improve the development of more tissue. In the drawing: BIODEGRADABLE POLYMER STRUCTURE THAT IS IMPLEMENTED BREAST CELLS THAT ARE REPRODUCED BETWEEN THEY ARE PLACED ON THE IMPLANT WHERE THE VASCULAR TISSUE GROWTH IS PERFORMED, A NEW TISSUE OF THE BREAST IS FORMED AND THE STRUCTURE IS EVENTUALLY DEGRADED