A double interpenetrating polymeric matrix hydrogel containing progenitor cells and biomolecules, for neuronal tissue repair

Abstract

Double interpenetrating polymeric matrix hydrogel comprising: - an acrylic acid polymer or carbomer, - a natural polymer selected from agarose, chitosan, dextran, a polysaccharide or a glycosaminoglycan, - a cross-linking agent selected from glycerol, propylene glycol, ethylene glycol, triethylamine or a mixture thereof, - added chemical moieties by click chemistry said double interpenetrating polymeric matrix hydrogel able to contain progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, placed on its external surface or hosted within a tridimensional lattice formed by said polymeric matrix, wherein said double interpenetrating polymeric matrix hydrogel is used for, integrally or partially, regenerating or reconstructing damaged tissues, particularly in acute and sub-acute spinal cord injury repair.

Description

[0001] DESCRIPTION “A DOUBLE INTERPENETRATING POLYMERIC MATRIX HYDROGEL CONTAINING PROGENITOR CELLS AND BIOMOLECULES, FOR NEURONAL TISSUE REPAIR” *.*.*.*.* Technical field of the invention The present invention relates to a double interpenetrating polymeric matrix hydrogel able to contain progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, inparticular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, evenmore preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via en- richment, most preferably autologous cells obtained via enrichment, said double interpenetrating polymeric matrix hy- drogel to be used for neuronal tissue repair, such as acute and subacute spinal cord injury (SCI) repair, chronic or post traumatic brain injury (TBI) repair, or peripheral nerve injury repair. State of the art When treating pathologies, that lead to the degeneration of a specific organ, the only possible therapeutic alternative is often represented by organ transplantation. Where possible, however, it is associated with a high grade of risk for the patient, since complications are likely to occur, such as the transmission of pathogens from donor to recipient or the rejection of the organ itself, along with the chronic shortage of available organs. Moreover, such an approach cannot be applied to all the body areas susceptible to degenerative pathologies. Actually, at present, transplant therapies may not be applied to many degenerative pathologies, in particular those of the central nervous system and sometimes also the myocardium. The above reasons are leading the research to be addressed towards the development of support matrixes, which allow to carry cells, allowing and / or increasing the same to survive and integrate with the target tissue, in order to giverise to a new functional tissue, histologically integrated with the adjacent structures.Since most cells of the central nervous system, for example neurons, do not, or poorly, proliferate nor renew, attention has been addressed to therapies based on replacement or regeneration of the damaged tissue, using isolated and re- grafted cells. Caron, I. et al., [“A new three dimensional biomimetic hydrogel to deliver factors secreted by human mesenchymal stem cells in spinal cord injury” Biomaterials 75, p. 135-147] describes a three-dimensional, agarose / carbomer / poly- ethylene glycol-based hydrogel for stem cell therapy of spinal cord injuries. According to Caron, extracellular matrix (ECM) and RGD are incorporated into the hydrogel for the attachment and maintenance of stem cells. However, Caron does not disclose that the acrylic acid polymer or carbomer is functionalized by click chemistry. WO 2009 / 144569 A2 describes a double interpenetrating polymer matrix hydrogel made of an acrylic acid polymer, a natural polymer, a polysaccharide or glycosaminoglycan and a cross-linking agent, which contains cells, e.g. stem cells, on its surface or in its interior. WO 95 / 05204 A1discloses a porous material made of one or more hydrophilic polymers as a wound dressing or pad material. The polymer solution used to produce the porous material can also comprise pharmaceutical active ingredi- ents. RGD peptides are also mentioned as an example of medication.In particular, the achievement of a particularly effective treatment of acute and subacute spinal cord injury seems to bea problem to which a solution has been sought for a long time. Summary of the invention Surprisingly and unexpectedly, the Applicant has developed a technology capable of allowing the in situ regeneration of the spinal cord favoring the generation of an anti-inflammatory and pro-regenerative environment. Said technology consists of a double interpenetrating polymeric matrix hydrogel, according to the present invention, that is applicable,

[0002] P70149PC00 in particular, to the external part of the spinal dura mater, say epidurally, a tightly woven tubular membrane that envel- ops the spinal cord along its entire length, allowing to keep in situ out of the spinal cord the regenerative cells placed on the external surface or hosted within the tridimensional lattice formed by said double interpenetrating polymeric matrix hydrogel, which is, in particular, intimately structured to specifically entrap and keep cells alive for a long timeframe. Accordingly, as a particularly effective treatment of acute and subacute spinal cord injury, the applicant has developed a double interpenetrating polymeric matrix hydrogel, according to anyone of the embodiments of the present invention, capable of allowing the in situ regeneration of the spinal cord or neuronal tissue repair, favoring the generation of an anti-inflammatory and pro-regenerative environment.Said double interpenetrating polymeric matrix hydrogel, according to anyone of the embodiments of the present inven-tion, is applicable, in particular, to the external part of the spinal dura mater, a tightly woven tubular membrane that its partially covering the circumference of the spinal cord, allowing to retain and keep in situ the regenerative progenitor cells of the spinal cord, such as progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment in par- ticular in a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, placed on the external surface or hosted within the tridimensional lattice formed by said double interpenetrating polymeric matrix hydrogel. Preferably said tridimensional lattice specifically tuned to allow cells entrapment and survival over time. The combination of “hydrogels and cells” shows medical properties because, in view of the nature of the hydrogel, said combination achieves the goal of keeping a mixture of stem cells and immunocompetent cells alive for many days, atleast 2 weeks and maintaining the stem cells in their stemness, and all capable of playing their role as secreting cellsto create an anti-inflammatory and pro-regenerative environment in the lesion site. It is an object of the present invention a double interpenetrating polymeric matrix hydrogel technology comprising: - an acrylic acid polymer or carbomer or acrylic polymer functionalized using click chemistry - a natural polymer selected from agarose, chitosan, dextran, cellulose, pullulan, starch, other polysaccharide or a glycosaminoglycan, - a cross-linking agent selected from glycerol, propylene glycol, ethylene glycol, triethylamine or a mixture thereof, - added chemical moieties by click chemistry said double interpenetrating polymeric matrix hydrogel is able to contain progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macro- phages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment in particular in such a way to ensure a sufficient number of cells in order tolet them spontaneously generate extracellular matrix inside the hydrogel, placed on its external surface or hosted withina tridimensional lattice formed by said polymeric matrix, preferably said tridimensional lattice specifically tuned to allow cells entrapment and survival over time; said double interpenetrating polymeric matrix hydrogel to be used in human or veterinary surgical or wound treatment, in, integrally or partially, regenerating or reconstructing damaged tissues, wherein, preferably said tissue is selected from the group comprising spinal cord tissue, brain tissue, or peripheral nerves, or for neuronal tissue repair, specifically in acute and subacute spinal cord injury repair, chronic or TBI repair, or peripheral nerve injury repair.

[0003] P70149PC00 Preferred embodiments of double interpenetrating polymeric matrix hydrogel according to the present invention are defined in embodiments from 2 to 18. The double interpenetrating polymeric matrix hydrogel object of the present invention is suitable to be used for prepar- ing a stem cell implant for, integrally or partially, regenerating or reconstructing damaged tissues. In particular wherein said tissue is selected from the group comprising, for example spinal cord tissue, brain tissue, or peripheral nerves. In particular, the double interpenetrating polymeric matrix hydrogel according to any preferred embodiments of the present invention is suitable to be used for acute and sub-acute spinal cord injury repair. More in general the double interpenetrating polymeric matrix hydrogel according to any preferred embodiments of the present invention according to anyone of the embodiments 1-18, or obtained with any method according to anyone ofthe embodiments 19 – 24 and 31 - 32 or claims 1-10, as herewith disclosed, containing progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesen- chymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, is suitable for use in in human or veterinary surgical or wound treatment, in, integrally or partially, regenerating or reconstructing damaged tissues, wherein, pref- erably said tissue is selected from the group comprising spinal cord tissue, brain tissue, or peripheral nerves, or for neuronal tissue repair, specifically in acute and subacute spinal cord injury repair, chronic or TBI repair, or peripheral nerve injury repair. It is a further object of the present invention a method for preparing a double interpenetrating polymeric matrix hydrogel as herewith disclosed, said method comprising: the preparation of a mixture comprising (% by weight) from 0,01% to 5% of acrylic or polymeric carbomer, from 0,01%to 5% of natural polymer, from 0,1% to 40% of cross-linking agent and optionally from 0,01% to 5% of elasticizing agentin an aqueous medium, at a temperature between 20°C and 90°C, for a time ranging from 10 seconds to 240 minutes, and eventually a subsequent cooling down to a temperature ranging from 37°C and room temperature; the use of acrylic or polymeric carbomers functionalized with chemical moieties trough click chemistry to produce hy- drogels. Preferred embodiments of the method for preparing double interpenetrating polymeric matrix hydrogel according to the present invention are defined in the embodiments from 19 to 24 and 31 to 32. List of figures Particular embodiments of the invention are described in detail herein below, as a way of example and not limited to, with reference to the attached figure, wherein: Fig.1: it is an essential representation of a human surgical application of the double interpenetrating polymeric matrix hydrogel according to the present invention wherein: A fat-tissue-derived stem cells (1) are loaded to interpenetrating polymeric matrix hydrogel according to the present invention in a form of freeze-dried hydrogel (2); B hydrogel swollen (3) and loaded with non-embryo-derived stem cells on its external surface or hosted within a tridimensional lattice formed by said polymeric matrix (4); C application (5) of the hydrogel loading stem cells nearby (6) the lesion zone (7),where laminectomy has been performed (6); D hydrogel loading the non-embryo-derived stem cells in epidural position(8) wherein the cellular healing factors cross over (9) the lesion zone (7). Detailed description of the invention A double interpenetrating polymeric matrix, according to an embodiment of the invention, comprises: - an acrylic acid polymer or carbomer, - a natural polymer selected from agarose, chitosan, dextran, a polysaccharide or a glycosaminoglycan, - a cross-linking agent selected from glycerol, propylene glycol, ethylene glycol, triethylamine or a mixture thereof,

[0004] P70149PC00 - added chemical moieties by click chemistry said double interpenetrating polymeric matrix hydrogel which is able to contain progenitor cells, such as non-embryo- derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or ob- tained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchy- mal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, placed on its external surface orhosted within a tridimensional lattice formed by said polymeric matrix, preferably said tridimensional lattice specificallytuned to allow cells entrapment and survival over time; said double interpenetrating polymeric matrix hydrogel to be used in human or veterinary surgical or wound treatment, in, integrally or partially, regenerating or reconstructing dam- aged tissues, wherein, preferably said tissue is selected from the group comprising spinal cord tissue, brain tissue, or peripheral nerves, or for neuronal tissue repair, particularly in acute and subacute spinal cord injury repair, chronic or TBI repair, or peripheral nerve injury repair. Said object is achieved through a double interpenetrating polymeric matrix hydrogel comprising an acrylic acid polymer or carbomer, a natural polymer selected among agarose, chitosan, dextran, a polysaccharide or a glycosaminoglycan, a cross-linking agent selected among glycerol, propylene glycol, ethylene glycol, triethylamine or a mixture thereof and, eventually, an elasticizing agent selected among alginic acid and salts thereof, any kind of collagen or derivatives thereof, fibrin, fibrinogen and hyaluronic acid or a derivative thereof, such as, for example, gelatine, collagen, chitin, chitosan, laminin, polyethylene glycol (PEG), methacrylates, cellulose, nanocellulose, or a mixture of said agents, said double interpenetrating polymeric matrix hydrogel is able to contain progenitor cells, such as non-embryo-derived stem cells, immune competent cells, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, APCs, preferably non-embryo-derived stem cells, more preferably non-embryo-derived stem cells and immune com-petent cells placed on its external surface or hosted within the tridimensional polymeric matrix, that is within the tridi-mensional lattice of the matrix itself. The polymeric matrix may comprise, preferably, (% by weight) from 0,01% to 5% of acrylic acid polymer or carbomer, from 0,01% to 5% of natural polymer, from 0,1% to 40% of cross-linking agent, and, eventually, from 0,01% to 5% of elasticizing agent. According to a preferred aspect, the double interpenetrating polymeric matrix hydrogel comprises Carbopol, as acrylic polymer, agarose, as natural polymer, and a mixture of glycerol, propylene glycol and triethylamine, as cross-linking agent and in another particularly preferred aspect, the double interpenetrating polymeric matrix hydrogel further com- prises gelatine or PEG, as elasticizing agent. Further, the double interpenetrating polymeric matrix hydrogel can comprise nutrients and / or growth factors for pro- genitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or viaenrichment, most preferably autologous cells obtained via enrichment.Such nutrients and / or growth factors for cells may be selected among: saline buffer, B27 (GIBCO®, Invitrogen, USA), Insulin-Transferrin-Selenium-G Supplement, D–(+)Glucose, L–Glutamine, Penicillin-Streptomycin, Sodium pyruvate, MEM (Minimum Essential Medium) Vitamins 100X, MEM Aminoacids 50X, MEM Non-Aminoacids 100X, NEAA (Non- essential Amino Acids), HEPES (4-2-hydroxyethil-1-piperazinil-ethanesulfonic acid) Buffer, bicarbonate, carnitine (Al- car), D-vitamin, L-arginine. According to a particularly preferred aspect, the double interpenetrating polymeric matrix hydrogel can further comprise

[0005] P70149PC00 one or more drugs and / or factors and / or active substances, capable of modulating the action of the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells , more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via en- richment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, and / or keeping them viable and / or stimulating their metabolism and / or stimulating their growth and development and / or stimulatingtheir differentiation and / or stimulating the interaction and mutual interconnectivity, because progenitor cells, such asnon-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells , more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, most preferably autologous cells obtained via enrichment, maintain their nature or stemness. Another aspect of the present invention relates to the use of said double interpenetrating polymeric matrix hydrogel for preparing a cell implant for regenerating or reconstructing or replacing damaged or dead or removed or non-functioning, even partially, spinal cord tissue. Indeed, through the present invention, the spinal cord tissue in the zone of said implant can be improved and cured by supplying progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrich- ment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells , more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained fromfat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrich- ment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, and controlling their proliferation, infiltration and integration with the spinal cord tissue surrounding the implant zone. Another aim of the present invention is to provide a means for the spinal cord tissue regeneration for patients suffering from acute and sub-acute spinal cord injury. Another object of the present invention is to provide a means for regenerating spinal cord tissues, with such character- istics that it can be easily positioned in the target zone, in particular on to the external spinal dura matter by means of traditional surgical or mini-invasive techniques and that it can be easily adapted to the dimensions of the implant site. Another aspect of the present invention is a method to prepare said double interpenetrating polymeric matrix hydrogel, which comprises the preparation of an aqueous dispersion comprising (% by weight) from 0,01% to 5% of acrylic acid polymer or carbomer, from 0,01% to 5% of natural polymer, from 0,1% to 40% of cross-linking agent, and, optionally, from 0,01% to 5% of elasticizing agent, at a temperature of 20-90°C and for a time ranging from 10 seconds to 240 minutes, as a function of the desired cross-linking grade and of the polymers used, and the possible subsequent coolingdown to a temperature ranging from 37°C and room temperature. To said polymer mixture a base can be added,capable of neutralizing the acid, which develops during the reaction, preferably selected among triethylamine and am- monia. Further, to the polymer mixture suitable salts or saline solutions or buffer solutions, directed to control the pH may be optionally added. According to a preferred aspect, the added buffer solution is PBS (phosphate buffered saline), according to a further particularly preferred aspect, to the polymer mixture, both the base triethylamine and the buffer solution PBS and NaOH

[0006] P70149PC00 are added. The weight ratios between the several components can be varied in the above defined ranges, thereby enabling to modulate the dimensions of the polymeric lattice mesh, according to the cell type to be carried in order to make the matrix more suitable to each specific population. Preferably the polymers and all the reagents used meet the purity and sterility standards required for medical, biological, biomedical and pharmaceutical applications. It is another object of the present invention to provide a new polymer mixture with the proper chemical-physical char- acteristics, so that progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autolo- gous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferablynon-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cellsand immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, can be mixed therein, guaranteeing the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen pre- senting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, survival and proliferation. Therefore, it is another aspect of the present invention a method for making a bi-dimensional or tridimensional bio- hybrid device, which can be used for housing and conveying progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigenpresenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells , more pref-erably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them sponta- neously generate extracellular matrix inside the hydrogel, for living tissue regeneration within the human body, i.e. non- embryo-derived stem cells, for the regeneration of soft spinal cord tissues. a further basic characteristic is that such device degrades with a kinetics predetermined and compatible with the integration between conveyed non-embryo- derived stem cells and target spinal cord tissue. Another aspect of the present invention relates to a method for the preparation of a device based on said double interpenetrating polymeric matrix hydrogel containing progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen pre- senting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells , more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtainedvia enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneouslygenerate extracellular matrix inside the hydrogel. Said progenitor cells, such as non-embryo-derived stem cells, im- mune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more pref- erably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells

[0007] P70149PC00 obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them sponta- neously generate extracellular matrix inside the hydrogel, can be drawn from a fresh tissue or kept in a culture, away from the support (for example a Petri dish or other), eventually washed and subsequently added to the polymer mixture, cross-linking agent and op-tonally elasticizing agent during the gelling process, preferably during the cooling step. In order to keep the highest number of non-embryo-derived stem cells trapped in the double interpenetrating polymeric matrix hydrogel matrix in a viable status and in growing conditions, it is preferable to add a suitable culture medium to the aqueous dispersion of polymers, before gelling. Alternatively, progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macro-phages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stemcells , more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, trapped in the polymeric matrix can be sup- plied with nutrients and factors necessary for their growth, contacting or immersing the double interpenetrating poly- meric matrix hydrogel in a proper culture medium. Moreover, during the gel preparation, one or more drugs and / or active substances and / or specific factors and / or other substances can be added, which are capable of modulating the specific action of progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progeni- tors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non- embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside thehydrogel, also according to the injured spinal cord tissue to be regenerated, and that are gradually released from thepolymeric matrix once the implant has been introduced outside spinal cord tissue, say epidurally. Such drugs and / or active substances and / or specific factors and / or other substances, added during the preparation of double interpenetrating polymeric matrix hydrogel, are capable of keeping the progenitor cells, such as non-embryo- derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or ob- tained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchy- mal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, viable and / or stimulating their metabolism and / or stimulating their growth and development and / or stimulating their differentiation and stimulating the interaction and mutual interconnectivity. Moreover, the aqueous polymeric mixture can be replaced with a culture medium, suitable for keeping the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those ob-tained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors,and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells , more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, in viable and growing conditions.

[0008] P70149PC00 According to an embodiment, during the preparation of the double interpenetrating polymeric matrix hydrogel, said aqueous mixture or said suitable culture medium can contain, additionally, the above-described nutrients and / or growth factors. Another aspect of the present invention is the use of a double interpenetrating polymeric matrix hydrogel, having a high capacity to swell, to embed a mixture, mainly based on water, and a culture medium, eventually added with factors and / or other active substances, said mixture being suitable to provide the hosting non-embryo-derived stem cells with the necessary nutrients, so that they can survive and proliferate within the double interpenetrating polymeric matrix hydrogel. Another aspect of the present invention is the use of a suitable culture medium, directed towards an improvement ofthe viable conditions of progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such asautologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, once encapsulated in the gel. Such medium can be used directly for making the gel with or without adding a buffer saline solution. In an embodiment of the present invention, the double interpenetrating polymeric matrix hydrogel has suitable chemical- physical characteristics to be produced in the form of a thin film by one of the methods known in the state of the art such as, by way of example and not comprehensive, spindle spin-coating with subsequent cooling, spindle dip-coating with subsequent cooling or sol-gel. The double interpenetrating polymeric matrix hydrogel thus obtained has a thickness in the order of the dimensions of the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment,endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs),preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in par- ticular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, to be hosted (a few tenths of a millimetre to several micrometres). Such a double interpenetrating polymeric matrix hydrogel can be also realized in the form of a thin film assembly, arranged on multiple overlapping layers and suitable for supporting and hosting different progenitor cells, such as non- embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesen- chymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, populations, in order to obtain apoly-functional advanced structure (Multi Layer Systems, MLS).In a preferred embodiment of the invention, the double interpenetrating polymeric matrix hydrogel is obtained from a mixture containing Carbopol, agarose, glycerol, and propylene glycol, in presence of triethylamine, dispersed in a proper aqueous medium, to which a nutrient mixture is added, such as, for example, glucose, essential amino-acids, proteins, oligo-elements, mineral salts, vitamins and bactericidal antibiotics. Such a double interpenetrating polymeric matrix hydrogel containing progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial

[0009] P70149PC00 progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells , more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, can be positioned, conveniently, within wells suitable for the cell culture. The encapsulated progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, inparticular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, evenmore preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via en- richment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, in the tridimen- sional matrix remain viable and keep on growing and aggregating, as it could be seen during the experiments, when the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuro- progenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-de- rived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from me- dullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydro- gel, morphology, viability and proliferation within the above described double interpenetrating polymeric matrix hydrogel were assessed. In another preferred embodiment of the invention, the double interpenetrating polymeric matrix hydrogel is obtainedfrom a mixture containing Carbopol, agarose, glycerol and propylene glycol, in presence of triethylamine, dispersed ina proper aqueous medium. Such a double interpenetrating polymeric matrix hydrogel is within a hollow circular support and rotated to such a speed that a thin and regular film may be obtained once cooled. Subsequently, on this double interpenetrating polymeric matrix hydrogel film, once removed from the support and positioned in a micro-electronic device, a liquid dispersion, containing non-embryo-derived stem cells, is laid. Likewise, within the mixture, progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those ob- tained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, can be directly positioned and therefore a thin film containing the same non-embryo-derived stem cells can be produced by means of the described method. A further aspect of the invention relates to the use of a double interpenetrating polymeric matrix hydrogel as describedabove, in any embodiment thereof, for preparing a progenitor cells, such as non-embryo-derived stem cells, immunecompetent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen pre- senting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously

[0010] P70149PC00 generate extracellular matrix inside the hydrogel. implant for, integrally or partially, regenerating or reconstructing or supporting self-regeneration of spinal cord tissue. According to a further aspect, such a double interpenetrating polymeric matrix hydrogel, as described above, in any embodiment thereof, can be used for preparing an implant of progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more pref- erably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cellsobtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them sponta-neously generate extracellular matrix inside the hydrogel, for, integrally or partially, regenerating or reconstructing dam- aged spinal cord due to acute or sub-acute injury. In case of mixtures of agarose and carbomer, for example, the use of cross-linking agents and acidity buffers results not only to be fundamental for controlling the gelling, but also for obtaining a suitable matrix for hosting the specific selected progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo- derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from me- dullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydro- gel, and for conveying them, keeping them viable and, thereby, guaranteeing their survival and therapeutically efficacy once implanted in situ. It is to be pointed out that the more efficient the polymeric matrix, obtained in a gel form, is when enabling the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologouscells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progeni-tors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non- embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, support, viability and proliferation therein, the more the gel structure matrix conformation fits that of the ex- tracellular matrix, expected by the cells. The double interpenetrating polymeric matrix hydrogel object of the present invention is suitable to be used for prepar- ing an implant comprising progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in sucha way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside thehydrogel, for, integrally or partially, regenerating or reconstructing damaged tissues. In particular wherein said tissue is spinal cord tissue. In particular, the double interpenetrating polymeric matrix hydrogel according to any preferred embodiments of the present invention is suitable to be used for acute and sub-acute spinal cord injury repair. More in general the double interpenetrating polymeric matrix hydrogel according to any preferred embodiments of the present invention, such as those according to anyone of the embodiments 1-18, or obtained with any method according

[0011] P70149PC00 to anyone of the embodiments 19 – 24 and 31 - 32 or claims 1-11, as herewith disclosed, is suitable for use in human or veterinary surgical or wound treatment, in, integrally or partially, regenerating or reconstructing damaged tissues, wherein, preferably said tissue is selected from the group comprising spinal cord tissue, brain tissue, or peripheral nerves, or for neuronal tissue repair, specifically in acute and subacute spinal cord injury repair, chronic or TBI repair, or peripheral nerve injury repair. As far as one or more drugs and / or factors and / or active substances, according to the present invention, capable of modulating the action of the progenitor cells, and / or keeping them viable and / or stimulating their metabolism and / or stimulating their growth and development and / or stimulating their differentiation and / or stimulating the interaction and mutual interconnectivity, are concerned, said one or more drugs and / or factors and / or active substances are preferablyselected among the following three main categories:1. Immunomodulating factors 2. Trophic Factor Supporting molecules 3. Angiogenic factors Such as those listed in the following table: . That are, per se, biologically known compounds. In the state of the art, Caron teaches (see “Introduction”) that ECM is a crucial element for maintaining the adhesion / at- tachment of the stem cells (hMSCs) as well as for trophic supply and survival and also maintains and preserves their stem cell ability. For this reason, the RGD-containing polymer matrix hydrogel according to Caron is also coated with a three-dimensional ECM (“deposition”). According to the claimed invention, however, the viability and stem cell capability of the (stem) cells contained in the polymer matrix hydrogel is achieved also without ECM coating. The hydrogel according to the invention as claimed compared to Caron’s teachings in the provision of RGD (or other cell adhesion promoters), namely by incorporation into the polymer matrix hydrogel using click chemistry (i.e. function- alization), ensures the (vital) adhesion of the cells also without the need for an ECM coating.Moreover, Caron does not encourage the skilled person to provide for functionalization of the polymer matrix hydrogelwith RGD using click chemistry. Furthermore, Caron also teaches away from the invention, since Caron prompts the skilled person to provide extracellular matrix for the preservation of viable cells by coating the hydrogel with it. Moreover, performing comparative examples among hydrogels disclosed in Caron's publication and the hydrogels ac- cording to the present invention, the latter showed improved mechanical and rheological properties. In the first comparative test, at the same time of 10 minutes, 30 minutes, and 24 hours the swelling ratio in % was 3000 - 3500, 4000 - 4500, and 3000 - 3500 for the hydrogel according to the present invention, preferably the hydrogel

[0012] P70149PC00 double lyophilized, respectively, compared with the swelling ratio in % of 500, 1000 and 1500 for the hydrogel disclosed in Caron. In the second comparative test, at the same time of 10 minutes, 30 minutes, and 24 hours the swelling ratio in % was 3500 - 4000, 4500 - 5000, and 3000 - 3500 for the hydrogel according to the present invention, preferably the hydrogel double lyophilized, respectively, compared with the swelling ratio in % of 1000, 1000 and 1500 for the hydrogel dis- closed in Caron. As far as the viability of the progenitor cells entrapped in the Hydrogel according to the present invention is concerned, said viability is maintained at 100% for at least 14 days, while the hydrogel disclosed in Caron ensures, at 14 days, only the 50% survival of the entrapped progenitor cells.PREFERRED EMBODIMENTSAmong the preferred embodiments of the double interpenetrating polymeric matrix hydrogel according to the present invention there are the following embodiments: 1. A double interpenetrating polymeric matrix hydrogel comprising: - an acrylic acid polymer or carbomer, - a natural polymer selected from agarose, chitosan, dextran, a polysaccharide or a glycosaminoglycan, - a cross-linking agent selected from glycerol, propylene glycol, ethylene glycol, triethylamine or a mixture thereof, - added chemical moieties by click chemistry. 2. A double interpenetrating polymeric matrix hydrogel according to embodiment 1, wherein the added chemical moie- ties are selected from the group comprising RGD or GFOGER sequence or gelatine. 3. A double interpenetrating polymeric matrix hydrogel according to embodiments 1 and 2 further containing progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those ob- tained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells,even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or viaenrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, placed on its external surface or hosted within a tridimensional lattice formed by said polymeric matrix, preferably said tridimensional lattice specifically tuned to allow cells entrapment and survival over time. 4. A double interpenetrating polymeric matrix hydrogel according to embodiment 3, wherein the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, maintain their stem nature or stemness. 5. A double interpenetrating polymeric matrix hydrogel according to anyone of embodiments 3-4, wherein the non-embryo-derived stem cells are mesenchymal stem cells.6. A double interpenetrating polymeric matrix hydrogel according to anyone of the embodiments 3-5, further comprising immune competent cells. 7. A double interpenetrating polymeric matrix hydrogel according to embodiment 6, wherein the immune competent cells are autologous cells. 8. A double interpenetrating polymeric matrix hydrogel according to embodiment 7, wherein the autologous cells are autologous cells obtained from fat tissue.

[0013] P70149PC00 9. A double interpenetrating polymeric matrix hydrogel according to embodiment 7, wherein the autologous cells are autologous cells obtained from medullary tissue, also via enrichment, also in such a number to allow them to sponta- neously generate extracellular matrix inside the hydrogel. 10. A double interpenetrating polymeric matrix hydrogel according to anyone of embodiments 1-9, wherein said poly- meric matrix further comprises an elasticizing agent selected from alginic acid and salts thereof, collagen or derivatives thereof, fibrin, fibrinogen, hyaluronic acid or derivatives thereof, laminin, gelatine, polyethylene glycol (PEG) or a mix- ture of said agents. 11. A double interpenetrating polymeric matrix hydrogel according to embodiment 10, wherein said polymeric matrix comprises from 0,01% to 5% of acrylic acid polymer or carbomer, from 0,01% to 5% of natural polymer, from 0,1% to40% of cross-linking agent, and eventually from 0,01% to 5% of elasticizing agent, wherein said % are by weight.12. A double interpenetrating polymeric matrix hydrogel according to embodiment 11, wherein said acrylic polymer is Carbopol, said natural polymer is agarose, said cross-linking agent is a mixture of glycerol, propylene glycol and tri- ethylamine and said elasticizing agent is gelatine. 13. A double interpenetrating polymeric matrix hydrogel according to embodiment 11, wherein said acrylic polymer is Carbopol, said natural polymer is agarose, said cross-linking agent is a mixture of glycerol, propylene glycol and tri- ethylamine. 14. A double interpenetrating polymeric matrix hydrogel according to anyone of the embodiments 1-13, further com- prising nutrients and / or growth factors for progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrich- ment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo- derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrich- ment.15. A double interpenetrating polymeric matrix hydrogel according embodiment 14, wherein said nutrients and / orgrowth factors for progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autolo- gous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, are selected from the group consisting of: saline buffer, B27 (GIBCO®, Invitrogen, USA), Insulin-Transferrin-Selenium-G Supplement, D–(+)Glucose, L–Glutamine, Penicillin-Streptomycin, Sodium pyruvate, MEM Vitamins 100X, MEM Aminoacids 50X, MEM Non-Essential Aminoacids 100X, NEAA, HEPES Buffer, bicarbonate, carnitin (Alcar), D-vitamin, L-arginine. 16. A double interpenetrating polymeric matrix hydrogel according to anyone of embodiments 1-15, further comprising one or more drugs and / or factors and / or active substances capable of modulating the action of the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, inparticular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, evenmore preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via en- richment, most preferably autologous cells obtained via enrichment, and / or keeping them viable and / or stimulating their metabolism and / or stimulating their growth and development and / or stimulating their differentiation and / or stimu- lating the interaction and mutual interconnectivity. 17. A double interpenetrating polymeric matrix hydrogel according to any of the previous embodiments, in the form of a thin film suitable for supporting progenitor cells, such as non-embryo-derived stem cells, immune competent cells,

[0014] P70149PC00 such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, for the use in acute and subacute spinal cord injury. 18. A double interpenetrating polymeric matrix hydrogel according to any of the previous embodiments, in the form of a thin film assembly, arranged on multiple overlapping layers suitable for supporting and hosting different progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those ob-tained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors,and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, populations in order to obtain a multifunctional advanced structure (Multi Layer Systems). 19. A method for preparing a double interpenetrating polymeric matrix hydrogel according to any of the previous em- bodiments, which comprises the preparation of a mixture comprising (% by weight) from 0,01% to 5% of acrylic or carbomeric polymer, from 0,01% to 5% of natural polymer, from 0,1% to 40% of cross-linking agent and optionally from 0,01% to 5% of elasticizing agent in an aqueous medium, at a temperature between 20°C and 90°C, for a time ranging from 10 seconds to 240 minutes, and eventually a subsequent cooling down to a temperature ranging from 37°C and room temperature. 20. Method according to embodiment 19, further comprising the addition of a base capable of neutralizing the acid which develops during the reaction and optionally the addition of suitable salts or saline solutions or buffer solutions for controlling the pH.21. Method according to embodiment 20, wherein said base is triethylamine.22. Method according to embodiments 20 or 21, wherein said buffer solution is PBS. 23. Method according to embodiments 19-22, wherein the aqueous mixture is added with or replaced with a culture medium suitable for keeping the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in viable and growing conditions containing in addition optionally nutrients and / or growth factors according to embodiments 14 or 15. 24. Method according to embodiments 19 or 23, wherein to the aqueous mixture or the culture medium one or more drugs and / or factors and / or active substances are added, capable of modulating progenitor cells, such as non-embryo- derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or ob- tained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving riseto macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchy-mal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, action and / or keeping them viable and / or stimulating their metab- olism and / or stimulating their growth and development and / or stimulating their differentiation and / or stimulating the interaction and mutual interconnectivity.

[0015] P70149PC00 25. Use of a double interpenetrating polymeric matrix hydrogel according to anyone of the embodiments 1-18, or ob- tained with any method according to anyone of the embodiments 19 – 24, for preparing an implant comprising progen- itor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, said implant for, integrally or partially, regener- ating or reconstructing damaged tissues.26. Use according to embodiment 25, where said tissue is selected from the group comprising spinal cord tissue, braintissue, or peripheral nerves. 27. A double interpenetrating polymeric matrix hydrogel for use according to anyone of the embodiments 25 or 26, comprising cells from the stromal vascular fraction wherein said cells are obtained by harvesting and enrichment out of autologous fat-tissue, duly processed enzymatic or purely mechanical means or a mixture thereof. 28. A double interpenetrating polymeric matrix hydrogel according to anyone of the embodiments 1-18, or obtained with any method according to anyone of the embodiments 19 – 24, for use in neuronal tissue repair, in particular acute and subacute spinal cord injury repair, chronic or TBI repair, or peripheral nerve injury repair. 29. A double interpenetrating polymeric matrix hydrogel according to anyone of the embodiments 1-18 or obtained with any method according to anyone of the embodiments 19 – 24, for use in human or veterinary surgical or wound treat- ment, where the intimate mesh structure of the hydrogel allows cells entrapment and long-lasting cells survival. 30. A double interpenetrating polymeric matrix hydrogel for use according to anyone of the embodiments 26-29, con- taining cells from the stromal vascular fraction wherein said cells are obtained by harvesting and enrichment out of autologous fat-tissue, duly processed enzymatic or purely mechanical means or a mixture thereof, in order to obtain a sufficient number of cells that can then spontaneously generate extracellular matrix inside the hydrogel.31. Method according to embodiments 19 – 24 wherein the entrapment of: progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or ob- tained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchy- mal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, is connected to the specific composition and mesh size of the double interpenetrating polymeric matrix hydrogel according to the embodiments 1 or 2. 32. Method according to embodiments 19 – 24 wherein the re-hydration from the frozen form, preferably its double lyophilized for, of the double interpenetrating polymeric matrix hydrogel according to the embodiments 1 or 2 helps the driving force and soaking up of: progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissueor obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment.33. A method for human or veterinary surgical or wound treatment, or for, integrally or partially, regenerating or recon- structing damaged tissues, wherein, preferably said tissue is selected from the group comprising spinal cord tissue, brain tissue, or peripheral nerves, or for neuronal tissue repair, in particular acute and subacute spinal cord injury repair, or for chronic or TBI repair, or for peripheral nerve injury repair, said method comprising administering, preferably in an effective amount thereof, of a double interpenetrating polymeric matrix hydrogel, preferably in its double lyophilized form, comprising:

[0016] P70149PC00 - an acrylic acid polymer or carbomer, - a natural polymer selected from agarose, chitosan, dextran, a polysaccharide or a glycosaminoglycan, - a cross-linking agent selected from glycerol, propylene glycol, ethylene glycol, triethylamine or a mixture thereof, - added chemical moieties by click chemistry, further containing progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autolo- gous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtainedfrom medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, in particular in sucha way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, placed on its external surface or hosted within a tridimensional lattice formed by said polymeric matrix, pref- erably said tridimensional lattice specifically tuned to allow cells entrapment and survival over time, to a subject in need thereof. 34. Method according to embodiment 33, wherein for subacute spinal cord injury repair, in particular in a human surgical application, the interpenetrating polymeric matrix hydrogel in a form of freeze-dried hydrogel (2), swollen (3) and loaded with fat-tissue-derived stem cells (1) on its external surface or hosted within a tridimensional lattice formed by said polymeric matrix (4), is applied (5) nearby (6) the lesion zone (7), where laminectomy has been performed (6), being the hydrogel loading the non-embryo-derived stem cells in epidural position (8) and where the cellular healing factors cross over (9) the lesion zone (7). 35. Method of human or veterinary surgical or wound treatment wherein a double hydrogel according to embodiments 3 - 18 is administered preferably in an affective amount thereof to a subject in need thereof. 36. Double interpenetrating polymeric matrix hydrogel according to embodiments 3 – 18, for use in, integrally or par- tially, regenerating or reconstructing damaged tissues, preferably said tissue is selected from the group comprisingspinal cord tissue, brain tissue, or peripheral nerves.37. Double interpenetrating polymeric matrix hydrogel according to embodiments 3 - 18 for use as a medicament. 38. Double interpenetrating polymeric matrix hydrogel according to embodiments 3- 18, for use in neuronal tissue repair, such as acute and subacute spinal cord injury repair, chronic or TBI repair, or peripheral nerve injury repair. 39. Double interpenetrating polymer matrix hydrogel according to embodiments 3-18, for use in human or veterinary surgical or wound treatment. 40. Double interpenetrating polymer matrix hydrogel of embodiment 37 for use in treating damaged or injured tissue selected from spinal cord tissue, brain tissue and peripheral nerves. 41. The double interpenetrating polymer matrix hydrogel of embodiment 40, wherein the damaged or injured tissue is affected by acute and subacute spinal cord injury, chronic or post-traumatic brain injury (traumatic brain injury) or pe- ripheral nerve injury. 42. Double interpenetrating polymeric matrix hydrogel according to any embodiments 1 - 18, double lyophylised, pref- erably wherein the chemical moieties added by click chemistry are selected from the group comprising RGD or GFOGER sequences or gelatine, more preferably said double interpenetrating polymeric matrix hydrogel according to any embodiments 1 - 18, double lyophylised, wherein the chemical moieties added by click chemistry are selected fromthe group comprising RGD or GFOGER sequences or gelatine, further contains progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or ob- tained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchy- mal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, placed on its external surface or hosted within a tridimensional

[0017] P70149PC00 lattice formed by said polymeric matrix, even more preferably said double interpenetrating polymeric matrix hydrogel according to any embodiments 1 - 18, double lyophylised, wherein the chemical moieties added by click chemistry are selected from the group comprising RGD or GFOGER sequences or gelatine, further contains progenitor cells and further containing one or more drugs and / or factors and / or active substances, capable of modulating the action of the progenitor cells, and / or keeping them viable and / or stimulating their metabolism and / or stimulating their growth and development and / or stimulating their differentiation and / or stimulating the interaction and mutual interconnectivity. 43. Method for preparing a double interpenetrating polymeric matrix hydrogel according to any embodiments 19 – 24 wherein the double interpenetrating polymeric matrix hydrogel according to any embodiments 1 – 18 is lyophilized one, two or more times.The following, not limitative, examples describe embodiments of the invention.EXAMPLES EMBODIMENTS ACCORDING TO THE INVENTION In order to show the feasibility, reproducibility and efficacy of a double interpenetrating polymeric matrix hydrogel ac- cording to the present invention, EXAMPLE 1 – Hydrogel preparation The following procedure describes the formation of an hydrogel with a percentage of carbomer equal to 0.5% and agarose 0.25%. Reactants necessary for the preparation of 100 mL of hydrogel: - Carbomer 974 P (CAS 151687-96-6, Fagron, Bufa, The Netherlands): 0.5 g - Agarose (CAS 9012-36-6, Invitrogen Corp., Carlsbad, USA): (0.25 g) - Glycerol (CAS 56-81-5, Merck Chemicals, Germany): 1 mL - Polyethylene glycol 2000 (CAS 25322-68-3, Merck Chemicals, Germany): 6 g - Sodium hydroxide (CAS 1310-73-2, preparazione ad alta purezza, Sigma-Aldrich, Germany): 0.5 mL - PBS: 100 mLOperative procedure for hydrogel synthesis:1. weight carbomer in a pyrex beaker and add PBS then mix at 1400 rpm for 30 min at room temperature (T=25°C); 2. add glycerol and polyethylene glycol, mix at the same velocity for 20 min at room temperature; 3. leave the suspension to rest for 60 min at room temperature; 4. neutralize the pH using sodium hydroxide and mix at 250 rpm: 5. weight and add agarose. Then mix at 250 rpm for 5 min at room temperature; 6. heat the solution with electromagnetic radiation (500W) for about 1 min until reach 80°C; 7. dilute the gelling solution with PBS 1:1 and cast in the final shape; 8. freeze the gels with dry ice and then lyophilize which can be done one or more times”. EXAMPLE 2 – Preparation of hydrogel functionalized with RGD through click chemistry We proceed with the same procedure described in example 1 where 0.01 g of carbomer are substituted with 0.1g of polyacrylic acid functionalized with RGD using click chemistry. Reactants necessary to produce polyacrylic acid func- tionalized with RGD are: - solution of polyacrylic acid 135,000 Da 35% w / w (CAS 9003-01-4, Merck Chemicals, Germany)- distilled water- 1-hydroxybenzotriazole hydrate (CAS 123333-53-9, Merck Chemicals, Germany) - propargylamine hydrochloride (CAS 15430-52-1, Merck Chemicals, Germany) - acetonitrile (CAS 75-05-8, Merck Chemicals, Germany) - ethyldimethylaminopropyl carbodiimide (CAS 25952-53-8, Merck Chemicals, Germany) - hydrochloric acid (CAS 7647-01-0, Merck Chemicals, Germany) - tetrahydrofuran (CAS 109-99-9, Merck Chemicals, Germany)

[0018] P70149PC00 - RGD-azide (Cymit Quimica, Spin) - copper iodide (CAS 7681-65-4, Merck Chemicals, Germany) - sodium ascorbate (CAS 134-03-2, Merck Chemicals, Germany) Procedure for polymer functionalization: 1. Dissolve the solution of polyacrylic acid (200 mg) at 25°C in distilled water (15 mL) and propargylamine hy- drochloride (15,4 mg) using a 50 mL round bottom flask mixing at 1400 rpm. 2. Dissolve 1-hydroxybenzotriazole hydrate in a solution 1:1 acetonitrile / water (total 14 mL) at 50°C and add to the mix at point 1; 3. Add (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (53.6 mg) in the bottom round flask and reach pH equalto 5.5 using hydrochloric acid 1M missing at 1400 rpm for 24 h. Then dialyze the product for 3 days in water andlyophilize. Functionalization degree will be controlled using NMR and should be in the range between 8% and 12%; 4. Dissolve the polymer obtained in point 3 (78 mg) in tetrahydrofuran (5 mL) and distilled water (10 mL) together with RGD-azide (25 mg) and add copper iodide (2.2 mg) and sodium ascorbate (2.2 mg); 5. Mix at 1400 rpm for 24 h at 60°C. Then dialyze the product for 3 days in water and lyophilize. Polymer func- tionalization will be checked using NMR.

[0019] P70149PC00

Claims

CLAIMS 1. Double interpenetrating polymeric matrix hydrogel comprising: - an acrylic acid polymer or carbomer, - a natural polymer selected from agarose, chitosan, dextran, a polysaccharide or a glycosaminoglycan, - a cross-linking agent selected from glycerol, propylene glycol, ethylene glycol, triethylamine or a mixture thereof, - added chemical moieties by click chemistry.

2. Double interpenetrating polymeric matrix hydrogel according to claim 1, wherein the added chemical moieties are selected from the group comprising RGD or GFOGER sequences or gelatine.

3. Double interpenetrating polymeric matrix hydrogel according to claims 1 - 2 further containing progenitor cells, suchas non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fattissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, placed on its external surface or hosted within a tridimen- sional lattice formed by said polymeric matrix.

4. Double interpenetrating polymeric matrix hydrogel according to claim 3, further comprising one or more drugs and / or factors and / or active substances, capable of modulating the action of the progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macro- phages, antigen presenting cells (APCs), preferably non-embryo-derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, and / or keeping them viable and / or stimulating their metabolism and / or stim-ulating their growth and development and / or stimulating their differentiation and / or stimulating the interaction and mu-tual interconnectivity.

5. A double interpenetrating polymeric matrix hydrogel according to anyone of claims 3-4, wherein the non-embryo- derived stem cells are mesenchymal stem cells.

6. A double interpenetrating polymeric matrix hydrogel according to anyone of claim 3-5, further comprising immune competent cells.

7. A double interpenetrating polymeric matrix hydrogel according to claim 6, wherein the immune competent cells are autologous cells.

8. A double interpenetrating polymeric matrix hydrogel according to claim 7, wherein the autologous cells are autolo- gous cells obtained from fat tissue, or via enrichment.

9. Double interpenetrating polymeric matrix hydrogel according to claim 7, wherein the autologous cells are obtained in such a way to ensure a sufficient number of cells in order to let them spontaneously generate extracellular matrix inside the hydrogel, also in such a number to allow them to spontaneously generate extracellular matrix inside the hydrogel.

10. Double interpenetrating polymeric matrix hydrogel according to claim 7, wherein the autologous cells are autologouscells obtained from medullary tissue.

11. Double interpenetrating polymeric matrix hydrogel according to anyone of claims 1-10, wherein acrylic or polymeric carbomer is functionalized by click chemistry.

12. Use of a double interpenetrating polymeric matrix hydrogel according to claims 1-2, for preparing an implant com- prising progenitor cells, such as non-embryo-derived stem cells, immune competent cells, such as autologous cells, such as those obtained from fat tissue or obtained from medullary tissue or via enrichment, endothelial progenitors, neuroprogenitors, and monocytes giving rise to macrophages, antigen presenting cells (APCs), preferably non-embryo-P70149PC00derived stem cells, in particular mesenchymal stem cells, more preferably non-embryo-derived stem cells and immune competent cells, even more preferably autologous cells, such as those obtained from fat tissue or obtained from me- dullary tissue or via enrichment, most preferably autologous cells obtained via enrichment, said implant for, integrally or partially, regenerating or reconstructing damaged tissues.

13. Double interpenetrating polymeric matrix hydrogel according to claims 3 – 11, for use in, integrally or partially, regenerating or reconstructing damaged tissues, preferably said tissue is selected from the group comprising spinal cord tissue, brain tissue, or peripheral nerves.

14. Double interpenetrating polymeric matrix hydrogel according to claims 3 - 11 for use as a medicament.

15. Double interpenetrating polymeric matrix hydrogel according to claims 3- 11, for use in neuronal tissue repair, suchas acute and subacute spinal cord injury repair, chronic or TBI repair, or peripheral nerve injury repair.

16. Double hydrogel according to claims 3-11, for use in human or veterinary surgical or wound treatment.

17. Double interpenetrating polymer matrix hydrogel of claim 14 for use in treating damaged or injured tissue selected from spinal cord tissue, brain tissue and peripheral nerves.

18. The double interpenetrating polymer matrix hydrogel of claim 13, wherein the damaged or injured tissue is affected by acute and subacute spinal cord injury, chronic or post-traumatic brain injury (traumatic brain injury) or peripheral nerve injury.P70149PC00

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