Methods and compositions for improving wound healing

Abstract

The present disclosure relates to compositions and methods for improving wound healing and tissue regeneration. More specifically, the present disclosure relates to compositions that direct the immune response within a wound towards a pro-regenerative response, and methods of using such compositions. Such compositions and methods are particularly useful in altering the immune response induced by a medical implant in order to avoid scarring and fibrosis at the implantation site.

Description

Technical Field

[0001] Priority Paragraph This application claims the benefit of priority of U.S. Provisional Application No. 63 / 367,994, filed on July 8, 2022, and U.S. Provisional Application No. 63 / 488,122, filed on March 2, 2023, under 35 U.S.C. § 119, the contents of which are hereby incorporated by reference in their entirety.

[0002] Description of Research and Development Sponsored by the Federal Government This research was supported by the Intramural Research Program of the National Institute of Biomedical Imaging and Bioengineering, an agency within the National Institutes of Health. The United States Government has certain rights in this invention.

[0003] The field of the present disclosure generally relates to wound treatment and tissue engineering.

Background Art

[0004] The goal of tissue engineering is to replace the function of missing or damaged tissues and organs. This can be achieved using biomaterial scaffolds that integrate damaged or missing tissues such as skin grafts and assist in their regeneration, and medical device implants made of synthetic materials that replace the function or aesthetics of those tissues such as artificial knee joint replacements (see, e.g., Patent Document 1, which is hereby incorporated by reference in its entirety). However, whenever a biomaterial or medical device is implanted into the human body, it necessarily alters homeostasis and induces a cascade of immune responses, which may result in positive outcomes such as scaffold integration and tissue growth, or may result in immune-mediated pathologies such as implant fibrosis or excessive inflammation and damage to surrounding tissues. Therefore, to create next-generation medical devices that can function within the immune system, it is necessary to understand the mechanisms by which the immune system interacts with artificial materials.

[0005] The field of evaluating the immune response to artificial materials has grown over the past decade, and many discoveries have been made regarding how immune cells are involved in the acceptance or rejection of materials and fibrosis. Merging with the fields of wound healing and developmental biology, several cells are involved in these processes. Specifically, type 2 immunity, including eosinophils, M2 macrophages, and Th2 T cells, as well as the signaling protein interleukin (IL)-4, has been described in the healing of various tissues, including the liver, nerves, skin, and muscle. Regulatory T cells and related cytokines (IL-10) have also been described in the healing process and are utilized in material acceptance strategies to attempt to suppress excessive immune activation. Regarding pathology, Th17 T cells, which are involved in tissue fibrosis and autoimmunity, are also associated with device fibrosis in mice and humans. The crosstalk between these immune cells, such as T cells and macrophages, together with fibroblasts, generates a complex network of cell signaling where multiple factors contribute to regeneration and acceptance, or inflammation and fibrosis.

[0006] To date, research has begun to clarify the cell processes involved in wound healing and fibrosis, but the success of treatments using existing methods has remained inconsistent, and the treatment of wounds and the avoidance of fibrosis remain challenges. Therefore, better and more specific treatments are needed that enable the regeneration of tissue within wounds without accompanying fibrosis. The present disclosure addresses that need.

Prior Art Documents

Patent Documents

[0007]

Patent Document 1

Summary of the Invention

[0008] One aspect is a method comprising administering to a site of a wound a therapeutic composition that induces a regeneration-promoting environment within and / or around the tissue in the wound.

[0009] One aspect is a method of altering the immune response to a wound, the method comprising administering to the site of the wound a therapeutic composition that induces a regeneration-promoting environment within and / or around the tissue of the wound.

[0010] One aspect is a method of treating a wound in an individual, the method comprising administering to the site of the wound a therapeutic composition that induces a regeneration-promoting environment within and / or around the tissue of the wound.

[0011] One aspect is a method of treating an individual having a wound, the method comprising administering to the site of the wound a therapeutic composition that induces a regeneration-promoting environment within and / or around the tissue of the wound.

[0012] One aspect is a method of reducing or preventing fibrosis of a wound, the method comprising administering to the site of the wound a therapeutic composition that induces a regeneration-promoting environment within and / or around the tissue of the wound.

[0013] One aspect is a method of implanting a medical device into an individual, the method comprising introducing the medical device into tissue within the individual and administering to the site of the implanted medical device a therapeutic composition that induces a regeneration-promoting environment within and / or around the tissue of the wound.

[0014] One aspect is a method of implanting a medical device into an individual, the method comprising introducing the medical device into tissue within the individual, wherein a therapeutic composition that induces a regeneration-promoting environment within and / or around the tissue of the wound is applied to the medical device prior to implanting the medical device into the individual.

[0015] In these aspects, the wound can be a wound due to a burn, contusion, plasma tumor, hematoma, laceration, detachment, puncture, surgical wound, incision, ulcer, or pressure injury. In these aspects, administering the therapeutic composition can include introducing the therapeutic composition into the wound cavity and / or the tissue around the wound cavity, which can include applying the therapeutic composition (e.g., topically) to the skin around the wound cavity.

[0016] In these embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste, which can include nanoparticles or microspheres. In these embodiments, the therapeutic composition can be formulated as an immediate-release composition or a sustained-release composition. In these embodiments, the therapeutic composition can release the therapeutic agent over approximately 5 minutes, over approximately 10 minutes, over approximately 30 minutes, over approximately 1 hour, over approximately 2 hours, over approximately 12 hours, over approximately 24 hours, over approximately 48 hours, over approximately 5 days, over approximately 1 week, or over approximately 1 month or more.

[0017] In these embodiments, the therapeutic composition can include a therapeutic agent that induces a regeneration-promoting environment. The therapeutic agent can steer the immune response away from a TH1-type response. The therapeutic agent can induce a TH2-driven immune environment. The therapeutic agent can increase the number of M2 macrophages in and / or around the tissue of the wound. The therapeutic agent can induce the influx of M2 macrophages in and / or around the tissue of the wound and / or the therapeutic agent can induce the local proliferation of M2 macrophages in and / or around the tissue of the wound. The therapeutic agent can induce an increase in the number of conventional dendritic cells (cDC1) in and / or around the tissue of the wound. The therapeutic agent can induce the influx of cDC1 in and / or around the tissue of the wound and / or the therapeutic agent can induce the local proliferation of cDC1 in and / or around the tissue of the wound. cDC1 can be cross-presenting dendritic cells. cDC1 can be XCR1 + CD103 + positive dendritic cells.

[0018] In these embodiments, the therapeutic agent can include peptides, proteins, glycoproteins, lipoproteins, lipids, sugars, polysaccharides, glycosaminoglycans, nucleic acid molecules, organic molecules, and combinations thereof. In these embodiments, the therapeutic agent can include a decellularized extracellular matrix (ECM), or a component derived therefrom, which can include degradation products of the ECM. In these embodiments, the therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and degradation products thereof. The therapeutic agent can include matricains, which can be metastatin, aifestin, canstatin, tetrastatin, pentastatin, lumstatin, hexastatin, endostatin, neostatins, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastocan, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent can include DAMPs.

[0019] In certain embodiments, the medical device can be an implant, which can be selected from the group consisting of breast implants, stents, ports, shunts, hip implants, knee implants, cochlear implants, hernia or other surgical meshes, intraocular lens implants, pacemakers, metal / surgical screws, rods, or pins, artificial intervertebral discs, and spinal fixation devices. In certain embodiments, the implant can include metals and metal alloys, plastic polymers, ceramics, hydrogels, and composite materials, which can include, but are not limited to, silicone, polyethylene, stainless steel, titanium, zirconia, polyurethane foam, polylactic acid, amalgam, gold, alumina, silicate, chromium, cobalt, and molybdenum.

[0020] One aspect is a therapeutic composition for treating a wound, the therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment within the wound and / or within the tissue surrounding the wound. The therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste, which can include nanoparticles or microspheres. The therapeutic composition can be formulated as an immediate-release composition or a sustained-release composition. The therapeutic composition can release the therapeutic agent over approximately 5 minutes, over approximately 10 minutes, over approximately 30 minutes, over approximately 1 hour, over approximately 2 hours, over approximately 12 hours, over approximately 24 hours, over approximately 48 hours, over approximately 5 days, over approximately 1 week, or over approximately 1 month or more. In certain aspects, the therapeutic composition can include a therapeutic agent that induces a regeneration-promoting environment. The therapeutic agent can steer the immune response away from a TH1-type response. The therapeutic agent can induce a TH2-driven immune environment. The therapeutic agent can increase the number of M2 macrophages within the wound and / or within the tissue surrounding the wound. The therapeutic agent can induce the influx of M2 macrophages within the wound and / or within the tissue surrounding the wound, and / or the therapeutic agent can induce the local proliferation of M2 macrophages within the wound and / or within the tissue surrounding the wound. The therapeutic agent can induce an increase in the number of conventional dendritic cells (cDC1) within the wound and / or within the tissue surrounding the wound. The therapeutic agent can induce the influx of cDC1 within the wound and / or within the tissue surrounding the wound, and / or the therapeutic agent can induce the local proliferation of cDC1 within the wound and / or within the tissue surrounding the wound. The cDC1 can be cross-presenting dendritic cells. The cDC1 can be XCR1 + CD103 +It can be a dendritic cell. The therapeutic agent can include peptides, proteins, glycoproteins, lipoproteins, lipids, sugars, polysaccharides, glycosaminoglycans, nucleic acid molecules, organic molecules, and combinations thereof. In certain embodiments, the therapeutic agent can include a decellularized extracellular matrix (ECM), or a component derived therefrom, which can include degradation products of the ECM. The therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and degradation products thereof. The therapeutic agent can include matricines, which can be metastatin, aifestin, canstatin, tetrastatin, pentastatin, ramstatin, hexastatin, endostatin, neostatins, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. The therapeutic agent can include DAMPs.

[0021] One aspect of the present disclosure is a method of mobilizing NK cells to a wound, or tissue proximal to a wound, the method comprising administering to the site of the wound a therapeutic agent or therapeutic composition of the present disclosure.

[0022] One aspect of the present disclosure is a method of activating NK cells, the method comprising contacting the NK cells with a therapeutic agent of the present disclosure.

[0023] One aspect of the present disclosure is a method for inducing an increase in the expression of Xcl1 in NK cells, the method comprising contacting the NK cells with a therapeutic agent of the present disclosure.

[0024] One aspect of the present disclosure is a kit comprising a therapeutic composition of the present disclosure. The kit may also include a needle, syringe, vial, applicator, and instructions for using the therapeutic composition to treat a wound.

[0025] One aspect of the present disclosure is a therapeutic composition of the present disclosure when used for the treatment of a wound, the therapeutic composition inducing a regeneration-promoting environment in the wound and / or in the tissue surrounding the wound.

[0026] One aspect of the present disclosure is a therapeutic composition of the present disclosure when used for changing the immune response to a wound, the therapeutic composition inducing a regeneration-promoting environment in the wound and / or in the tissue surrounding the wound.

[0027] One aspect of the present disclosure is a therapeutic composition of the present disclosure when used for reducing or preventing fibrosis of a wound, the therapeutic composition inducing a regeneration-promoting environment in the wound and / or in the tissue surrounding the wound.

[0028] This patent or application documents include at least one drawing created in color. Copies of this patent or patent application publication that include color drawings are provided by the Patent Office upon request and payment of the required fee.

Brief Description of the Drawings

[0029]

Figure 1

Figures 2A - B

Figure 2C

Figure 3A

Figure 3B

Figure 3C

Figure 3D

Figure 4A

Figure 4B

Figure 4C

Figure 4D

Figure 4E

Figure 5A

Figure 5B

Figure 6A

Figure 6B

Figure 6C

Figure 7A

Figure 7B

Figure 7C

Figure 7D

Figure 7E

Figure 7F

Figure 7G

Figure 7H

Figure 8

Figures 9A - B

Figure 9C

Figure 10

Figure 11

Figure 12

Figure 13

Figure 14

Figure 15

Figure 16 - 1

Figure 16 - 2

Figure 16 - 3

Figure 17

Figure 18 - 1

Figure 18 - 2

Figure 18 - 3

Figure 19

Figure 20

Figure 21A

Figure 21B

Figure 21C

Figure 21D

Figure 21E

Figure 22

Figure 23

Figure 24A

Figure 24B

Figure 24C

Figure 25

Figure 26A

Figure 26B

Figure 26C

Figure 26D

Figure 27

Figure 28

Figure 29

Figure 30

Mode for Carrying Out the Invention

[0030] The present disclosure relates to methods and compositions for improving wound healing. More specifically, the present disclosure relates to compositions that direct the immune response within a wound towards a pro-regenerative response, and methods of using such compositions. The disclosed compositions and methods are particularly useful in altering the immune response elicited in response to an implanted device. Implanted devices often cause scarring and fibrosis at the implantation site. Thus, the methods of the present disclosure can generally be carried out by introducing into the site of the wound a composition of the present disclosure that induces a pro-regenerative environment within the wound. In certain embodiments, the pro-regenerative environment comprises the novel dendritic cells described herein.

[0031] Before further describing the invention, it is to be understood that the invention is not limited to the particular embodiments described, and as such, can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting, since the scope of the present disclosure is defined only by the claims.

[0032] It is noted that, as used in this specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. For example, a compound refers to one or more compound molecules. Thus, the terms “a,” “an,” “one or more,” and “at least one” can be used interchangeably. Similarly, the terms “comprising,” “including,” and “having” can be used interchangeably. It is further noted that the claims can be drafted to exclude any optional element. Thus, this description is intended to serve as antecedent basis for use of exclusive terminology, such as “solely,” “only,” and the like, in connection with the listing of claim elements, or use of “negative” limitations.

[0033] The publications disclosed in this specification are provided only with respect to their disclosure prior to the filing date of this application. Nothing in this specification should be construed as an admission that the present disclosure is not entitled to antedate such publications for the purposes of prior disclosure. Further, the provided publication dates may be different from the actual publication dates and may need to be independently verified. All publications mentioned in this specification are incorporated herein by reference for the purpose of disclosing and describing the methods and / or materials for which those publications are cited.

[0034] Unless otherwise defined, all technical and scientific terms used in this specification have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods and materials are described hereinafter. Terms and expressions common to various aspects disclosed in this specification are defined below.

[0035] Wound healing involves various growth factors and cytokines that control cell growth, cell differentiation, and cell proliferation. Wound healing generally occurs in stages, although these stages may overlap to some extent. Following tissue injury, epithelial cells and / or endothelial cells release inflammatory mediators that initiate the antifibrinolytic coagulation cascade, which induces blood clot formation. This is followed by the inflammatory and proliferative phases, where white blood cells are mobilized and then activated and induced to proliferate by chemokines and growth factors. Activated white blood cells secrete fibrosis-promoting cytokines such as IL-13 and TGF-β. Stimulated epithelial cells, endothelial cells, and myofibroblasts produce matrix metalloproteinases (MMPs) that break down the basement membrane, as well as additional cytokines and chemokines that mobilize and activate neutrophils, macrophages, T cells, B cells, and eosinophils, which are important components of tissue regeneration. Activated macrophages and neutrophils clear tissue debris, dead cells, and invading organisms. Immediately after the initial inflammatory phase, myofibroblasts produce ECM components and endothelial cells form new blood vessels. Myofibroblasts can be derived from locally mobilized mesenchymal cells from the bone marrow (which are known as fibroblasts), or they can be induced by epithelial-mesenchymal transition (EMT). In the subsequent remodeling and maturation phases, activated myofibroblasts stimulate wound contraction. Collagen fibers also become more organized, blood vessels recover normally, scar tissue is removed, and epithelial cells and / or endothelial cells divide and migrate across the basement layer, resulting in the regeneration of epithelium or endothelium, respectively, and the damaged tissue returning to its normal appearance. The aforementioned process generally describes the regeneration-promoting response to a wound. However, in certain cases, such as chronic wounds, the normal healing process is disrupted. Persistent inflammation, tissue necrosis, and infection lead to chronic activation of myofibroblasts and excessive accumulation of ECM components, promoting the formation of permanent fibrous scars. However, the inventors have discovered that by treating a wound with an appropriate composition, the immune response within the wound can be diverted from a fibrotic response towards a regeneration-promoting response.

[0036] One aspect of the present disclosure is a method comprising administering to a site of a wound a therapeutic composition that induces a regeneration-promoting environment within and / or in the tissue surrounding the wound.

[0037] The term "wound" refers to an injury to the integrity of living tissue such as skin (including epidermis, dermis, and subcutaneous), mucosa, and organ tissue (e.g., muscle, lung tissue, etc.). Wounds suitable for use in the methods of the present disclosure can be either closed wounds or open wounds. Wounds of the present disclosure include, but are not limited to, burns, contusions, plasma tumors, hematomas, lacerations, exfoliations, punctures, surgical wounds (e.g., incisions), ulcers, and wounds due to compression injuries. Thus, in one aspect, the wound is selected from the group consisting of burns, contusions, plasma tumors, hematomas, lacerations, exfoliations, punctures, surgical wounds (e.g., incisions), ulcers, and wounds due to compression injuries.

[0038] "Administering to the site of the wound", "applying to the wound", etc. mean introducing the therapeutic composition into the wound cavity (e.g., incision) and / or the tissue surrounding the wound cavity. The most visible sign of a wound such as an incision is the resulting open cavity, but the tissue surrounding the cavity is also affected, and the infiltration and proliferation of immune cells in such tissue contribute to wound healing. Thus, in certain aspects, the introduction of the therapeutic composition may include or be exclusive to the tissue surrounding the wound cavity. In certain aspects, the location of introduction can be determined based on the physical properties of the therapeutic composition. For example, if the composition is a cream or foam, it may be best to apply the composition to the skin surface of the wound cavity. Alternatively, if the composition is a liquid, the composition can be applied to the skin within the wound cavity and / or the composition can be injected into the tissue within or around the wound cavity.

[0039] A "therapeutic composition" is a composition comprising a therapeutic agent that induces a regeneration-promoting environment at and around the location where the therapeutic composition is administered to an individual. The therapeutic compositions of the present disclosure can include any formulation suitable for the delivery of the therapeutic agent such that the therapeutic agent can induce a regeneration-promoting environment. In certain embodiments, the therapeutic composition can be formulated as an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles.

[0040] In certain embodiments, the therapeutic composition can include at least one additional agent (e.g., a pharmaceutically acceptable excipient) such as a buffering agent, a stabilizing agent, a chelating agent, an antioxidant, a preservative, and any mixture thereof.

[0041] In certain embodiments, the therapeutic composition can be formulated as a sustained-release composition. A sustained-release composition is a composition that releases the active ingredient (e.g., the therapeutic agent) slowly over a period of time rather than all at once. Methods for making sustained-release compositions are known to those of skill in the art.

[0042] "Therapeutic agent" refers to a molecule or combination of molecules that, when administered to an individual, induces a regeneration-promoting response at the location where the therapeutic composition is administered to the individual, preferably proximal thereto. The therapeutic agent can be any type of molecule that can induce a regeneration-promoting environment. In certain embodiments, the therapeutic agent can include peptides, proteins, glycoproteins, lipoproteins, lipids, sugars, polysaccharides, glycosaminoglycans, nucleic acid molecules, organic molecules, and any combination thereof. Thus, the therapeutic agent can include more than one type of molecule. The therapeutic agents of the present disclosure can be isolated from biological materials (e.g., cells, organisms, etc.), produced using recombinant DNA technology, chemically synthesized, or produced using a combination of such techniques. The term "isolated" does not indicate a particular degree of isolation.

[0043] In certain embodiments, the therapeutic agent can include a decellularized extracellular matrix (ECM) or one or more components derived therefrom. The ECM is the acellular portion of tissues and organs and includes a network of proteins (e.g., collagen, elastin, laminin) as well as other molecules such as proteoglycans and polysaccharides. After injury, enzymes (especially MMPs) are activated and these activated enzymes degrade ECM proteins such as collagen and elastin. These degradation products act, among other things, to limit the migration of stromal cells and accelerate the proliferation of fibroblasts. Thus, in certain embodiments, the therapeutic agent can include an ECM or one or more components derived therefrom. In certain embodiments, the therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, and agrin.

[0044] The dynamic remodeling of the ECM is necessary for wound healing. During remodeling, the ECM undergoes proteolytic processing to release bioactive matrix fragments called "matricines". Examples of matricines are disclosed in WO2022 / 055974, which is hereby incorporated by reference in its entirety. Matricines have been shown to promote cell infiltration, progressive tissue damage, or wound healing. Thus, these signals represent an important effector mechanism of the ECM for cell signaling and the transport of cells to target organs. In certain embodiments, the therapeutic agent may comprise one or more degradation products of the ECM. In certain embodiments, the therapeutic agent may comprise a matricine. In certain embodiments, the therapeutic agent may comprise one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, and agrin. In certain embodiments, the therapeutic agent comprises a matricine selected from the group consisting of metastatin, aifestin, canstatin, tetrastatin, pentastatin, ramstatin, hexastatin, endostatin, neostatins, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof.

[0045] In certain embodiments, the therapeutic agent may include one or more damage - associated molecular patterns (DAMPs). Damage - associated molecular patterns (DAMPs) are endogenous danger molecules that are released from damaged or dying cells and activate the innate immune system, for example, by interacting with pattern recognition receptors (PRRs). DAMPs can be derived from various sources, such as extracellular proteins, such as biglycan and tenascin C, intracellular proteins, such as high - mobility group box 1 (HMGB1), histones, S100 proteins, heat - shock proteins (HSPs), and plasma proteins such as fibrinogen, Gc - globulin, and serum amyloid A (SAA). Examples of DAMPs include, but are not limited to, biglycan, decorin, versican, LMW hyaluronan, heparan sulfate, fibronectin containing the EDA domain, fibrinogen, tenascin C, uric acid, S100 proteins, heat - shock proteins, adenosine triphosphate (ATP), F - actin, cyclophilin A, amyloid - beta (Aβ), histones, HMGB1, HMGN1, IL - 1α, IL - 33, SAP130, deoxyribonucleic acid (DNA), ribonucleic acid (RNA), mitochondrial DNA (mtDNA), transcription factor A mitochondrial (TFAM), formyl peptides, mitochondrial reactive oxygen species (mROS), calreticulin, defensins, cathelicidin (LL37), eosinophil - derived neurotoxin, granules, syndecan, and glypican, but are not limited thereto.

[0046] In some embodiments, the therapeutic agent may include a compound that induces the production or release of ECM components, or one or more components, matricines, or DAMPs therefrom.

[0047] As used herein, a "regeneration promoting environment" refers to the composition of factors (e.g., immune cells, cytokines, etc.) within and around a wound that steers the healing process away from fibrosis, promotes the regeneration of tissue within the wound, such that the damaged tissue returns to its normal appearance. As described above, injury initiates a cascade of events that cause ECM and tissue remodeling, as well as the mobilization of cells to the wound site for host defense and tissue repair. After injury, some of the most rapidly responsive cells are polymorphonuclear cells, including neutrophils, eosinophils, and basophils. Neutrophils are phagocytic cells that scavenge debris and bring about the mobilization of other cells, such as macrophages. Thus, within the first few days after injury, a set of diverse immune cells are mobilized to the site of injury. In certain embodiments, a therapeutic agent induces a production promoting environment that includes a Th2-driven immune environment. The TH2 environment is characterized by the presence of eosinophils, basophils, mast cell degranulation, and M2 macrophages, among which the M2 macrophages are associated with wound healing and repair. Additionally, the Th2 environment exhibits elevated levels of one or more cytokines important for the induction of a humoral immune response, such as interleukin-4 (IL-4), IL-5, IL-10, and IL-13. In certain embodiments, the Th2-driven environment may include an eosinophil-predominant granulocyte compartment at the site of injury. In certain embodiments, the therapeutic agents of the present disclosure may induce the influx of macrophages having an M2 phenotype. In certain embodiments, the therapeutic agents of the present disclosure may induce the local proliferation of M2 macrophages at the site of injury. In certain embodiments, the therapeutic agents of the present disclosure may induce the differentiation of macrophages into M2 macrophages. In certain embodiments, M2 macrophages within a Th2-driven environment may include high levels of CD206, CD301b, and CD169. Such levels indicate the local proliferation of tissue resident cells. Thus, the therapeutic agents of the present disclosure may induce an increase in the number of M2 macrophages having high levels of CD206, CD301b, and CD169 within the wound.

[0048] The second most common antigen-presenting cell (APC) found within the post-injury wound environment is the dendritic cell, e.g., CD11c + CD11b lo / negIt is a dendritic cell. The inventors have discovered that the therapeutic composition of the present disclosure induces a regeneration-promoting environment including an increase in the number of conventional dendritic cells (cDC1). cDC1 cells can initiate a new T cell response. In addition, they can attract T cells, secrete cytokines, and enhance local cytotoxic T cell function. cDC1 cells can also induce tolerance. In certain embodiments, the therapeutic agent of the present disclosure can induce an increase in the number of conventional dendritic cells (cDC1). Such cells can be cross-presenting dendritic cells, which are XCR1 + CD103 + and can be dendritic cells. In certain embodiments, the therapeutic composition of the present disclosure induces the influx of XCR1+CD103+cDC1. In certain embodiments, the therapeutic composition of the present disclosure induces the local proliferation of XCR1+CD103+cDC1. In certain embodiments, the therapeutic composition of the present disclosure induces the differentiation of dendritic cells into XCR1+CD103+cDC1.

[0049] In certain embodiments, the therapeutic agent can induce a therapeutic environment that may include an increase in NK cells. Such an increase may be due to an increase in the mobilization of NK cells to the site of the therapeutic agent or an increase in the replication of NK cells. In certain embodiments, such NK cells can be CD49b+TCRβ-. In certain embodiments, such NK cells can exhibit upregulation of Xcl1 gene expression that can be induced by the therapeutic agent.

[0050] One aspect of the present disclosure is a method of altering the immune response to a wound in an individual, the method comprising administering to the site of the wound a therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment within and / or around the tissue of the wound, thereby altering the immune response to the wound. In certain embodiments, the wound can include wounds due to burns, contusions, plasma tumors, hematomas, lacerations, avulsions, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce a regeneration-promoting immune response within the wound or in the tissue proximal to the wound. In certain embodiments, the regeneration-promoting immune response can include an eosinophil-predominant granulocyte compartment. In certain embodiments, the therapeutic agent can induce a TH2-driven immune response. In certain embodiments, the therapeutic agent can induce an influx of M2 macrophages to the wound. In certain embodiments, the therapeutic agent can induce local proliferation of M2 macrophages within the wound. In certain embodiments, the therapeutic agent can induce macrophages to differentiate into M2 macrophages. In certain embodiments, the Th2-driven immune response can include macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can induce local proliferation of tissue-resident macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can cause enrichment of cross-presenting dendritic cells in the wound or in the tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce an influx of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce local proliferation of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce dendritic cells to differentiate into cross-presenting dendritic cells. In certain embodiments, the cross-presenting dendritic cells can include cDC1 cells. In certain embodiments, the cross-presenting dendritic cells can be XCR1+CD103+ dendritic cells. In certain embodiments, the cross-presenting dendritic cells can express intermediate levels of CD86. In certain embodiments, the therapeutic agent can, in an individual, CD44 + CD26L- can induce an increase in T cells. In certain embodiments, CD44 + CD26L - T cells can include CD4+ cells. In certain embodiments, CD44 + CD26L - T cells can include CD8+ cells. In certain embodiments, the therapeutic agent can include one or more components of the ECM. In certain embodiments, the therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, and agrin. In certain embodiments, the therapeutic agent can include one or more degradation products of one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent includes matricines selected from the group consisting of metastatin, arresten, canstatin, restin, pentastatin, laminastatin, hexastatin, endostatin, neostatins, angiostatin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent can include one or more DAMPs.

[0051] One aspect of the present disclosure is a method of treating an individual having a wound, the method comprising administering to the site of the wound a therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment within and / or around the tissue of the wound, thereby treating the individual. In certain embodiments, the wound can include wounds due to burns, contusions, plasma tumors, hematomas, lacerations, exfoliations, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce a regeneration-promoting immune response within the wound or in the tissue proximal to the wound. In certain embodiments, the regeneration-promoting immune response can include an eosinophil-predominant granulocyte compartment. In certain embodiments, the therapeutic agent can induce a TH2-driven immune response. In certain embodiments, the therapeutic agent can induce the influx of M2 macrophages into the wound. In certain embodiments, the therapeutic agent can induce the local proliferation of M2 macrophages within the wound. In certain embodiments, the therapeutic agent can induce macrophages to differentiate into M2 macrophages. In certain embodiments, the Th2-driven immune response can include macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can induce the local proliferation of tissue-resident macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can cause the enrichment of NK cells in the wound or in the tissue proximal to the wound. Such enrichment can result from an increase in the mobilization of NK cells or an increase in the local proliferation of NK cells. In certain embodiments, such NK cells can be CD49b+TCRβ-. In certain embodiments, such NK cells can exhibit upregulation of Xcl1 gene expression, which can be induced by the therapeutic agent.

[0052] In certain embodiments, the therapeutic agent can cause enrichment of cross-presenting dendritic cells in the wound or tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce influx of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce local proliferation of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce differentiation of dendritic cells into cross-presenting dendritic cells. In certain embodiments, the cross-presenting dendritic cells can include cDC1 cells. In certain embodiments, the cross-presenting dendritic cells can be XCR1+CD103+ dendritic cells. In certain embodiments, the cross-presenting dendritic cells can express intermediate levels of CD86. In certain embodiments, the therapeutic agent can increase CD44 + CD26L - in an individual. In certain embodiments, CD44 + CD26L - T cells can include CD4+ cells. In certain embodiments, CD44 + CD26L -T cells may include CD8+ cells. In certain embodiments, the therapeutic agent may include one or more components of the ECM. In certain embodiments, the therapeutic agent may include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent may include one or more degradation products of one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent includes matricines selected from the group consisting of metastatin, aifestin, canstatin, tetrastatin, pentastatin, ramstatin, hexastatin, endostatin, neostatins, anasterin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent may include one or more DAMPs.

[0053] One aspect of the present disclosure is a method of treating a wound in an individual, the method comprising administering to the site of the wound a therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment within and / or around the tissue of the wound, thereby treating the wound. In certain embodiments, the wound can include wounds due to burns, contusions, plasma tumors, hematomas, lacerations, exfoliations, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce a regeneration-promoting immune response within the wound or in the tissue proximal to the wound. In certain embodiments, the regeneration-promoting immune response can include an eosinophil-predominant granulocyte compartment. In certain embodiments, the therapeutic agent can induce a TH2-driven immune response. In certain embodiments, the therapeutic agent can induce an influx of M2 macrophages into the wound. In certain embodiments, the therapeutic agent can induce local proliferation of M2 macrophages within the wound. In certain embodiments, the therapeutic agent can induce macrophages to differentiate into M2 macrophages. In certain embodiments, the Th2-driven immune response can include macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can induce local proliferation of tissue-resident macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can cause enrichment of NK cells in the wound or in the tissue proximal to the wound. Such enrichment can be due to an increase in mobilization of NK cells or an increase in local proliferation of NK cells. In certain embodiments, such NK cells can be CD49b+TCRβ-. In certain embodiments, such NK cells can exhibit upregulation of Xcl1 gene expression, which can be induced by the therapeutic agent. In certain embodiments, the therapeutic agent can cause enrichment of cross-presenting dendritic cells in the wound or in the tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce an influx of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce local proliferation of cross-presenting dendritic cells.In certain embodiments, the therapeutic agent can induce dendritic cells to differentiate into cross-presenting dendritic cells. In certain embodiments, the cross-presenting dendritic cells can include cDC1 cells. In certain embodiments, the cross-presenting dendritic cells can be XCR1+CD103+ dendritic cells. In certain embodiments, the cross-presenting dendritic cells can express intermediate levels of CD86. In certain embodiments, the therapeutic agent is CD44 in an individual. + CD26L - can induce an increase in T cells. In certain embodiments, CD44 + CD26L - T cells can include CD4+ cells. In certain embodiments, CD44 + CD26L -T cells can include CD8+ cells. In certain embodiments, the therapeutic agent can include one or more components of the ECM. In certain embodiments, the therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, and agrin. In certain embodiments, the therapeutic agent can include one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, and agrin. In certain embodiments, the therapeutic agent includes matricines selected from the group consisting of metastatin, arresten, canstatin, tetrastatin, pentastatin, laminstatin, hexastatin, endostatin, restin 1, restin 2, restin 3, restin 4, endostatin, neostatins, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, and tumstatin. In certain embodiments, the therapeutic agent can include one or more DAMPs.

[0054] One aspect of the present disclosure is a method of implanting a medical device into an individual, the method comprising introducing the medical device into tissue within the individual and administering a therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment within and / or around the tissue at the site of the implanted medical device. In certain embodiments, the wound can include wounds due to burns, contusions, seromas, hematomas, lacerations, detachments, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce a regeneration-promoting immune response within the wound or in tissue proximal to the wound. In certain embodiments, the regeneration-promoting immune response can include an eosinophil-predominant granulocyte compartment. In certain embodiments, the therapeutic agent can induce a TH2-driven immune response. In certain embodiments, the therapeutic agent can induce an influx of M2 macrophages into the wound. In certain embodiments, the therapeutic agent can induce local proliferation of M2 macrophages within the wound. In certain embodiments, the therapeutic agent can induce macrophages to differentiate into M2 macrophages. In certain embodiments, the Th2-driven immune response can include macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can induce local proliferation of tissue-resident macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can cause enrichment of NK cells in the wound or in tissue proximal to the wound. Such enrichment can be due to an increase in mobilization of NK cells or an increase in local proliferation of NK cells. In certain embodiments, such NK cells can be CD49b+TCRβ-. In certain embodiments, such NK cells can exhibit upregulation of Xcl1 gene expression, which can be induced by the therapeutic agent. In certain embodiments, the therapeutic agent can cause enrichment of cross-presenting dendritic cells in the wound or in tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce an influx of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce local proliferation of cross-presenting dendritic cells.In certain embodiments, the therapeutic agent can induce dendritic cells to differentiate into cross-presenting dendritic cells. In certain embodiments, the cross-presenting dendritic cells can include cDC1 cells. In certain embodiments, the cross-presenting dendritic cells can be XCR1+CD103+ dendritic cells. In certain embodiments, the cross-presenting dendritic cells can express intermediate levels of CD86. In certain embodiments, the therapeutic agent is CD44 in an individual. + CD26L - can induce an increase in T cells. In certain embodiments, CD44 + CD26L - T cells can include CD4+ cells. In certain embodiments, CD44 + CD26L -T cells can include CD8+ cells. In certain embodiments, the therapeutic agent can include one or more components of the ECM. In certain embodiments, the therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent can include one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent can include matricines selected from the group consisting of metastatin, aifestin, canstatin, tetrastatin, pentastatin, ramstatin, hexastatin, endostatin, restin 1, restin 2, restin 3, restin 4, endostatin, neostatine, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent can include one or more DAMPs. "Medical device" refers to any device implanted in an individual for the purpose of improving the health and / or function of the individual. An example of a medical device is an implant. Examples of implants include, but are not limited to, breast implants, stents, ports, shunts, hip implants, knee implants, cochlear implants, hernia surgical mesh implants, intraocular lens implants, pacemakers, metal / surgical screws, metal / surgical rods, metal / surgical pins, artificial intervertebral discs, and spinal fixation devices.Such implants can be made using, for example, metals and metal alloys, plastic polymers, ceramics, hydrogels, and composite materials, including but not limited to silicone, polyethylene, stainless steel, titanium, zirconia, polyurethane foam, polylactic acid, amalgam, gold, alumina, silicate, chromium, cobalt, and molybdenum.

[0055] One aspect of the present disclosure is a method of reducing or preventing fibrosis of a wound, the method comprising administering to the site of the wound a therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment within and / or around the tissue of the wound, thereby reducing or preventing fibrosis of the wound. In certain embodiments, the wound can include wounds due to burns, contusions, seromas, hematomas, lacerations, detachments, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce a regeneration-promoting immune response within the wound or in the tissue proximal to the wound. In certain embodiments, the regeneration-promoting immune response can include an eosinophil-predominant granulocyte compartment. In certain embodiments, the therapeutic agent can induce a TH2-driven immune response. In certain embodiments, the therapeutic agent can induce influx of M2 macrophages into the wound. In certain embodiments, the therapeutic agent can induce local proliferation of M2 macrophages within the wound. In certain embodiments, the therapeutic agent can induce differentiation of macrophages into M2 macrophages. In certain embodiments, the Th2-driven immune response can include macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can induce local proliferation of tissue-resident macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can cause enrichment of NK cells in the wound or in the tissue proximal to the wound. Such enrichment can be due to an increase in mobilization of NK cells or an increase in local proliferation of NK cells. In certain embodiments, such NK cells can be CD49b+TCRβ-. In certain embodiments, such NK cells can exhibit upregulation of Xcl1 gene expression, which can be induced by the therapeutic agent. In certain embodiments, the therapeutic agent can cause enrichment of cross-presenting dendritic cells in the wound or in the tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce influx of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce local proliferation of cross-presenting dendritic cells.In certain embodiments, the therapeutic agent can induce dendritic cells to differentiate into cross-presenting dendritic cells. In certain embodiments, the cross-presenting dendritic cells can include cDC1 cells. In certain embodiments, the cross-presenting dendritic cells can be XCR1+CD103+ dendritic cells. In certain embodiments, the cross-presenting dendritic cells can express intermediate levels of CD86. In certain embodiments, the therapeutic agent is CD44 in an individual. + CD26L - can induce an increase in T cells. In certain embodiments, CD44 + CD26L - T cells can include CD4+ cells. In certain embodiments, CD44 + CD26L -T cells may include CD8+ cells. In certain embodiments, the therapeutic agent may include one or more components of the ECM. In certain embodiments, the therapeutic agent may include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent may include one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent includes matricines selected from the group consisting of metastatin, aifestin, canstatin, tetrastatin, pentastatin, ramstatin, hexastatin, endostatin, restin 1, restin 2, restin 3, restin 4, endostatin, neostatins, anasterin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent may include one or more DAMPs.

[0056] One aspect of the present disclosure is a therapeutic composition for treating a wound, the composition comprising a therapeutic agent that induces a regeneration-promoting environment within the wound and / or within the tissue surrounding the wound. In certain embodiments, the wound can include wounds due to burns, contusions, plasma tumors, hematomas, lacerations, exfoliations, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, a spray, a lotion, a gel, a cream, a foam, a solution, a suspension, an emulsion, a hydrogel, or a paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce a regeneration-promoting immune response within the wound or within the tissue proximal to the wound. In certain embodiments, the regeneration-promoting immune response can include an eosinophil-predominant granulocyte compartment. In certain embodiments, the therapeutic agent can induce a TH2-driven immune response. In certain embodiments, the therapeutic agent can induce the influx of M2 macrophages into the wound. In certain embodiments, the therapeutic agent can induce the local proliferation of M2 macrophages within the wound. In certain embodiments, the therapeutic agent can induce macrophages to differentiate into M2 macrophages. In certain embodiments, the Th2-driven immune response can include macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can induce the local proliferation of tissue-resident macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can cause the enrichment of NK cells in the wound or in the tissue proximal to the wound. Such enrichment can be due to an increase in the mobilization of NK cells or an increase in the local proliferation of NK cells. In certain embodiments, such NK cells can be CD49b+TCRβ-. In certain embodiments, such NK cells can exhibit upregulation of Xcl1 gene expression, which can be induced by the therapeutic agent. In certain embodiments, the therapeutic agent can cause the enrichment of cross-presenting dendritic cells in the wound or in the tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce the influx of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce the local proliferation of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce dendritic cells to differentiate into cross-presenting dendritic cells.In certain embodiments, the cross-presenting dendritic cells can include cDC1 cells. In certain embodiments, the cross-presenting dendritic cells can be XCR1+CD103+ dendritic cells. In certain embodiments, the cross-presenting dendritic cells can express intermediate levels of CD86. In certain embodiments, the therapeutic agent is CD44 in an individual. + CD26L - can induce an increase in T cells. In certain embodiments, CD44 + CD26L - T cells can include CD4+ cells. In certain embodiments, CD44 + CD26L -T cells can include CD8+ cells. In certain embodiments, the therapeutic agent can include one or more components of the ECM. In certain embodiments, the therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent can include one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent includes matricines selected from the group consisting of metastatin, arresten, canstatin, tetrastatin, pentastatin, laminastatin, hexastatin, endostatin, neostatins, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent can include one or more DAMPs.

[0057] One aspect of the present disclosure is a method of mobilizing NK cells to a wound or tissue proximal to a wound, the method comprising administering to the site of the wound a therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment within the wound and / or in the tissue surrounding the wound, thereby mobilizing NK cells to the wound or tissue proximal to the wound. The wound can include wounds due to burns, contusions, plasma tumors, hematomas, lacerations, exfoliations, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce an increase in the mobilization of NK cells to the wound or tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce an increase in the local proliferation of NK cells. In certain embodiments, the NK cells can be CD49b+TCRβ-. In certain embodiments, the NK cells can exhibit upregulation of Xcl1 gene expression, which can be induced by the therapeutic agent. In certain embodiments, the therapeutic agent can include one or more components of the ECM. In certain embodiments, the therapeutic agent can include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent can include one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof.In certain embodiments, the therapeutic agent comprises a matricine selected from the group consisting of metallastatin, aifestin, canstatin, tetrastatin, pentastatin, ramstatin, hexastatin, endostatin, restin 1, restin 2, restin 3, restin 4, endostatin, neostatine, anasterin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent may comprise one or more DAMPs.

[0058] One aspect of the present disclosure is a method of activating NK cells, the method comprising contacting the NK cells with a therapeutic agent of the present disclosure. In certain embodiments, the therapeutic agent causes replication of the NK cells. In certain embodiments, the therapeutic agent causes migration of the NK cells. In certain embodiments, the therapeutic agent increases Xcl1 gene expression in the NK cells. In certain embodiments, the therapeutic agent induces an increase in the secretion of Xcl1 protein. In certain embodiments, contacting comprises introducing the therapeutic agent into a wound or tissue proximal to the wound. In certain embodiments, contacting comprises introducing the therapeutic agent to NK cells in vitro (e.g., tissue culture). In certain embodiments, the therapeutic agent may comprise one or more components of the ECM. In certain embodiments, the therapeutic agent may comprise one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent may comprise one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof.In certain embodiments, the therapeutic agent comprises a matrikine selected from the group consisting of metallastatin, arresten, canstatin, tetrastatin, pentastatin, lumastatin, hexastatin, endostatin, restin 1, restin 2, restin 3, restin 4, endostatin, neostatins, anasterin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent may comprise one or more DAMPs.

[0059] One aspect of the present disclosure is a method of inducing an increase in the expression of Xcl1 in NK cells, the method comprising contacting the NK cells with a therapeutic agent of the present disclosure. In certain embodiments, the therapeutic agent increases Xcl1 gene expression in NK cells. In certain embodiments, the therapeutic agent induces an increase in the secretion of Xcl1 protein. In certain embodiments, contacting comprises introducing the therapeutic agent into a wound or tissue proximal to the wound. In certain embodiments, contacting comprises introducing the therapeutic agent to NK cells in vitro (e.g., tissue culture). In certain embodiments, the therapeutic agent may comprise one or more components of the ECM. In certain embodiments, the therapeutic agent may comprise one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent may comprise one or more degradation products from one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent comprises a matrikine selected from the group consisting of metastatin, arresten, canstatin, tetrastatin, pentastatin, lumican, hexastatin, endostatin, neostatins, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent may comprise one or more DAMPs.

[0060] A kit for treating a wound, the kit comprising a therapeutic composition comprising at least a therapeutic agent that induces a regeneration-promoting environment within and / or in the tissue surrounding the wound. In certain embodiments, the wound can include wounds due to burns, contusions, plasma tumors, hematomas, lacerations, exfoliations, punctures, surgical wounds, incisions, ulcers, and pressure injuries. In certain embodiments, the therapeutic composition can include an ointment, spray, lotion, gel, cream, foam, solution, suspension, emulsion, hydrogel, or paste. In certain embodiments, the therapeutic composition can include liposomes, microspheres, or nanoparticles. In certain embodiments, the therapeutic agent can induce a regeneration-promoting immune response within the wound or in the tissue proximal to the wound. In certain embodiments, the regeneration-promoting immune response can include an eosinophil-predominant granulocyte compartment. In certain embodiments, the therapeutic agent can induce a TH2-driven immune response. In certain embodiments, the therapeutic agent can induce the influx of M2 macrophages into the wound. In certain embodiments, the therapeutic agent can induce the local proliferation of M2 macrophages within the wound. In certain embodiments, the therapeutic agent can induce macrophages to differentiate into M2 macrophages. In certain embodiments, the Th2-driven immune response can include macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can induce the local proliferation of tissue-resident macrophages having high levels of CD206, CD301b, and / or CD169. In certain embodiments, the therapeutic agent can cause the enrichment of NK cells in the wound or in the tissue proximal to the wound. Such enrichment can result from an increase in the mobilization of NK cells or an increase in the local proliferation of NK cells. In certain embodiments, such NK cells can be CD49b+TCRβ-. In certain embodiments, such NK cells can exhibit upregulation of Xcl1 gene expression, which can be induced by the therapeutic agent. In certain embodiments, the therapeutic agent can cause the enrichment of cross-presenting dendritic cells in the wound or in the tissue proximal to the wound. In certain embodiments, the therapeutic agent can induce the influx of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce the local proliferation of cross-presenting dendritic cells. In certain embodiments, the therapeutic agent can induce dendritic cells to differentiate into cross-presenting dendritic cells.In certain embodiments, the cross-presenting dendritic cells can include cDC1 cells. In certain embodiments, the cross-presenting dendritic cells can be XCR1+CD103+ dendritic cells. In certain embodiments, the cross-presenting dendritic cells can express intermediate levels of CD86. In certain embodiments, the therapeutic agent is CD44 in an individual. + CD26L - can induce an increase in T cells. In certain embodiments, CD44 + CD26L - The T cells can include CD4+ cells. In certain embodiments, CD44 + CD26L -T cells may include CD8+ cells. In certain embodiments, the therapeutic agent may include one or more components of the ECM. In certain embodiments, the therapeutic agent may include one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent may include one or more degradation products of one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and any combination thereof. In certain embodiments, the therapeutic agent includes matricines selected from the group consisting of metastatin, aifestin, canstatin, tetrastatin, pentastatin, lambstatin, hexastatin, endostatin, restin 1, restin 2, restin 3, restin 4, endostatin, neostatine, anasterin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan- , elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGRYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, tumstatin, and any combination thereof. In certain embodiments, the therapeutic agent may include one or more DAMPs. The disclosed kit may include additional components, such as needles, syringes, vials, applicators, and instructions for using the composition of the kit to treat a wound.

[0061] This specification uses examples to disclose the present disclosure, including the best mode, and to enable the practice of the present disclosure, including the making and using of any device or system by any person skilled in the art, and the carrying out of any incorporated method. The patentable scope of the present disclosure is defined by the claims, and other examples that occur to those skilled in the art may be included. Such other examples are intended to be within the scope of the claims if they have structural elements that do not differ from the language of the claims, or if they include equivalent structural elements that do not have a substantial difference from the language of the claims.

Examples

[0062] Materials and Methods Preparation of Materials The small intestine was procured from American Yorkshire pigs (Wagner Meats) at 5 - 6 months of age. The submucosa (SIS) was mechanically isolated by removing the muscular layer and then mechanically scraping off the luminal layer. The obtained SIS was rinsed with distilled water and frozen at -80°C until decellularization. After thawing, the SIS was cut into 1-inch segments in a biosafety cabinet and then incubated for 30 minutes with vigorous shaking in 4% ethanol (Fisher Scientific) and 0.1% peracetic acid (Sigma), or on a stirring plate. The obtained decellularized ECM was neutralized by continuous washing with sterile 1×PBS and distilled water. After blotting the liquid with a sterile absorbent pad, the material was transferred to a 50 ml conical tube and frozen at -80°C until freeze-dried for 48 hours. The dried material was filled into a sterile cryomilling container and milled into fine powder. The obtained powder was hydrated with sterile saline to form a thick paste, which was then filled into a 1 ml slip tip syringe and applied to the wound site.

[0063] Polyethylene powder (PE) with a particle size of less than 150 μm was purchased from Goodfellow Cambridge Limited, immersed in distilled water, rinsed with 70% ethanol, and UV-sterilized in ethanol for 30 minutes. It was stored in 70% ethanol until use. The PE particles suspended in ethanol were transferred to Eppendorf tubes and dried overnight in a biosafety cabinet. Since PE is hydrophobic, the sample cannot be filled into a syringe and is applied directly to the wound as a powder.

[0064] Volumetric muscle loss surgery Mice were subjected to bilateral volumetric muscle loss trauma according to the method (1) described above. Briefly, the lower limbs of 6- to 8-week-old female C57BL / 6 WT mice (Jackson laboratory) were shaved with an electric clipper and excess hair was removed with depilatory cream one day before surgery. The next day, the mice were anesthetized in an anesthesia chamber with 4.0% isoflurane in oxygen at a flow rate of 200 cc / min, and subcutaneous injection of buprenorphine was performed for pain management. Subsequently, the mice were maintained with 2.0% isoflurane during this procedure. After the surgical site was sterilized with triple Betadine followed by 70% isopropanol, a 1-cm incision was made in the skin and the fascia over the quadriceps muscle. Using surgical scissors, a 3- to 4-mm defect was created in the mid-belly of the quadriceps muscle by removing one-third of the quadriceps muscle. After removing the muscle, the resulting tissue space was filled with either a uniform volume (50 μl) of polyethylene microparticles or a porcine-derived ECM scaffold. Subsequently, the wound was closed with 3 to 4 wound clips, and this procedure was repeated on the opposite leg. After surgery, the mice were placed under a heat lamp for 2 to 3 minutes to recover from anesthesia. Then, the mice were returned to their cages and placed on a normal diet supplemented with nutrients until the end of the study. The protocol was approved by the NIH Clinical Center Animal Care and Use Committee under animal protocol number NIBIB20-01.

[0065] Flow cytometry At 3, 7, 21, and 42 days post-injury, mice were sacrificed ethically, and then the injured muscle was harvested together with the scaffold. Next, the dissected muscle was minced and digested with a digestion medium (0.5 mg / mL Liberase™ (Sigma) and 0.2 mg / ml DNase I (Roche) in HEPES-supplemented medium) at 37 °C for 45 minutes at 100 rpm on a shaker. Then, the digested suspension was filtered through a 70 μm cell strainer, washed with 1×PBS, and centrifuged at 350 g for 5 minutes at room temperature. Next, the cell pellet was immersed in 5 mM EDTA in 1×PBS solution for 10 minutes to reduce cell aggregation. After a 10-minute incubation, the cells were washed again with 1×PBS and then centrifuged at 350 g for 5 minutes at 4 °C. Next, the pellet was resuspended in 200 μL of a viability dye solution (1:1000 dilution of Live / Dead Blue (Thermo Fisher) in PBS) on ice for 20 minutes and then washed with a wash buffer (1% BSA and 2 mM EDTA in 1×PBS). Next, these cells were stained with either a myeloid panel or a lymphoid panel antibody (Tables 1 and 2) and then incubated at 4 °C for 30 minutes. After incubation, the cells were washed three times with the wash buffer and analyzed on a Cytek Aurora.

[0066] Intracellular staining After surface staining, the cells were fixed, permeabilized using the True-Nuclear™ Transcription Factor Kit (BioLegend), and intracellular staining of FoxP3 and HELIOS antibodies (Table 2) was carried out according to the manufacturer's guidelines. Briefly, after the final wash of surface staining, the cells were resuspended in True-Nuclear™ 1x Fix Concentrate and incubated at 4°C for 45 minutes. After incubation, the cells were centrifuged at 400×g for 10 minutes at 4°C and resuspended in True-Nuclear™ 1x Perm Buffer. The cells were then washed again with 1xPerm buffer and then treated with HELIOS and FoxP3 antibody cocktail (1:100 dilution of antibodies in True-Nuclear™ 1x Perm Buffer) followed by incubation at 4°C for 45 minutes. After incubation, the cells were washed twice with True-Nuclear™ 1x Perm Buffer. After the final wash, the cells were resuspended in wash buffer and analyzed by flow cytometry.

[0067] Histopathology The samples were fixed in 10% neutral buffered formalin for 48 - 72 hours and then transferred to 70% ethanol. Subsequently, the samples were dehydrated through a stepwise ethanol series of 70%, 80%, 95%, and 100% ethanol, cleared in xylene, and embedded in paraffin wax. Next, the quadriceps femoris muscle group was transected transversely to expose the center of the injury, which was then mounted upside down in a paraffin mold. Subsequently, 5 - 7 μm sections were placed on charged slide glasses and baked dry overnight at 56°C. After rehydration, the sections were incubated in Harris hematoxylin for 5 minutes, followed by washing with tap water, decolorized by immersion twice in acidic ethanol, rinsed with ethanol, and stained with hematoxylin and eosin (H&E) by immersion in eosin Y. The samples were then washed with 95% ethanol, dehydrated, covered with a coverslip, and mounted with PerMount. Picrosirius red (PSR) staining was performed according to the manufacturer's instructions. After rehydration, the samples were stained with PSR for 1 hour, washed with water, de-stained by immersion twice in acetic acid, dehydrated, and mounted with PerMount. The slides were imaged with an EVOS microscope (Thermo).

[0068] Immunohistochemistry The samples were rehydrated and then incubated in citrate antigen retrieval buffer for 20 minutes, followed by slow cooling on the bench top for 20 minutes. Endogenous peroxidase was quenched by incubation with 0.3% hydrogen peroxide in 1×PBS for 5 minutes. Samples were stained using the VECTASTAIN® Elite ABC-HRP kit (Rabbit, Vector Laboratories) according to the manufacturer's instructions. Briefly, after washing with 1×PBS, the samples were blocked with 2.5% normal goat serum for 1 hour. The samples were incubated in primary antibody diluted in blocking buffer for 1 hour. Rabbit monoclonal anti-CD103 (AbCam) and anti-E-cadherin (AbCam) were diluted at a 1:100 dilution. After washing the slides three times with 1×PBS, they were incubated with biotinylated secondary antibody for 30 minutes. The slides were washed three times with 1×PBS and then incubated in VECTASTAIN Elite ABC reagent for 30 minutes. The samples were washed three times with 1×PBS and then incubated in ImmPACT® DAB EqV peroxidase (HRP) substrate (Vector Laboratories) for 1 minute 30 seconds (CD103) or 50 seconds (E-cadherin). The slides were washed with tap water and then counterstained with Harris hematoxylin (Sigma) for 5 minutes, rinsed with tap water, destained with acidic ethanol, dehydrated, and mounted with PerMount.

[0069] RNA Isolation and RT-PCR Seven days after injury, the quadriceps muscle group was dissected from the mice and homogenized in 2 ml of 1×PBS for 30 seconds using a mechanical homogenizer at 5000 rpm. The resulting 500 microliters of homogenate was transferred to an Eppendorf tube containing 500 ul of TRI reagent solution (Sigma Aldrich). The sample was vortexed and then stored at -80 °C until RNA isolation. After thawing, 200 ul of chloroform (Sigma Aldrich) was added to each sample, vortexed, and then separated for 5 minutes at room temperature, followed by centrifugation at 8000×g for 15 minutes at 4 °C. The aqueous phase was combined with an equal volume of 70% ethanol and vortexed. The resulting sample was passed through an RNeasy Mini Prep spin column (Qiagen) and then washed according to the manufacturer's instructions by one wash with RW1 Buffer and two washes with RPE Buffer. The membrane was dried and the RNA was eluted in 30 ul of RNAse-free water. The RNA concentration was determined by NanoDrop and quality control was performed, and samples with an A260 / A280 greater than 2 were advanced. The samples were diluted to a concentration of 100 ng / ul and 11 ul was added to the SuperScript reverse transcriptase IV reaction according to the manufacturer's instructions, using random hexamer (ThermoFisher Scientfic) as the primer. The resulting 2 ul of cDNA was added together with 10 ul of TaqMan Fast Advanced Master Mix, 7 ul of nuclease-free water, and 1 ul of FAM-MGB primer / probe: Gusb, Xcl1 (Table 3).

[0070] Protein Isolation and Cytokine Analysis from Mouse Tissues Muscle and lymph node samples were snap-frozen in liquid nitrogen or ethanol-dry ice slurry immediately after dissection and stored until processing. The frozen muscle samples were added to 2 ml of ice-cold 1×PBS containing protease inhibitor (ThermoFisher Scientific) and cut with scissors. The samples were homogenized for 45 seconds using a mechanical homogenizer at 5000 - 6000 rpm on ice. Then, an additional 2.5 ml of ice-cold 1×PBS containing protease inhibitor was added along with 50 μl of 10% Triton-X100, followed by vigorous mixing. After leaving it on ice for 5 minutes, it was aliquoted, snap-frozen in liquid nitrogen, and stored until use. On the day of use, the samples were thawed and centrifuged at 10,000×g for 10 minutes to pellet the debris. Protein concentration was measured by diluting 1:1 with lysis buffer via Pierce BCA Protein Assay Kit (Thermo Scientific). 200 micrograms of protein was loaded onto a Proteome Profiler™ Array, Mouse XL Cytokine Array Kit and assayed according to the manufacturer's instructions. The blot was imaged with a 30-second exposure on a BioRad ChemiDoc.

[0071] Enzyme-linked immunosorbent assay Measurement of XCL-1 in mouse plasma samples was performed using a Mouse XCL-1 SimpleStep ELISA Kit (Abcam). The assay was performed according to the manufacturer's guidelines. Briefly, 50 μL of mouse plasma samples or protein lysates diluted 1:1 with blocking buffer were added to the appropriate wells of a pre-coated 96-well plate. The samples were then treated with 50 μL of antibody cocktail and subsequently incubated for 1 hour at room temperature. After incubation, the mixture in the wells was aspirated and the wells were washed 3 times with wash buffer. After the final wash, 100 μL of TMB chromogenic solution was added to each well and the plate was incubated for 10 minutes. After incubation, 100 μL of stop solution was added to each well, and subsequently, the OD was read at 450 nm.

[0072] Statistics and data analysis Flow cytometry data were deconvolved using the described single spectral controls (Supplementary Tables 1, 2) with SpectroFlo Software (Cytek Biosciences). The resulting unmixed data were exported to.fcs and then analyzed in FlowJo (Supplementary Figures 2, 16). Dimensionality reduction algorithms were run through the FlowJo plugin. t-Distributed Stochastic Neighbor Embedding (t-SNE) was run with the following parameters: learning setup - opt-SNE, number of iterations - 2000, perplexity - 30, KNN algorithm - exact (vantage point tree), gradient algorithm - Barnes-Hut. Uniform Manifold Approximation and Projection (UMAP) was run with the following parameters: Euclidean, nearest neighbors - 15, minimum distance - 0.5, number of components - 2. FlowSOM 3.0.18 was run with the following parameter: number of metaclusters - 30. Clustering was performed on single live immune cells using all parameters except LIVE / DEAD Blue. Data obtained from manual gating were analyzed in GraphPad Prism v9 and R4.1.2. Immunohistochemistry for E-cadherin was quantified by Fiji's autowhite balance (ImageJ) followed by color deconvolution, isolating the DAB channel. Regions of interest were manually outlined and then measured, converting to optical density (OD) readings by taking the logarithm (maximum intensity / average intensity). Each replicate represents quantification of sections from different animals. The resulting OD values were plotted in GraphPad Prism v9 for data display and analysis. Chemiluminescent Proteome Profiler blots were quantified by pixel intensity via MatLab (version R2022a) using Protein Array Toolbox version 2.0.0.1, normalized to background, and then presented in R4.1.2 as fold change relative to uninjured control muscle tissue. After subtracting the housekeeping gene (Gusb), technical replicates of RT-PCR Ct values were averaged, and the mean ΔCt of uninjured controls was subtracted from all ΔCt values, then converted to fold change and presented as 2(- ΔΔ Ct).

[0073] The statistical tests used are described in the figure captions.

[0074] Example 1. Evaluation of the Characteristics of Immune Cell Responses after Injury To evaluate the immune response to artificial materials and tissue regeneration in trauma, bilateral volumetric muscle injury surgery was performed, followed by implantation of materials. After creating a 3-mm defect in the quadriceps muscle group, the resulting void was filled back with either a control (saline), hydrated decellularized extracellular matrix (ECM) powder, or polyethylene powder (PE). ECM provides an example of a regeneration-promoting material, and PE is an example of a fibrosis-promoting material. A 22-color flow cytometry panel was used to examine the bone marrow immune response to material responses and the early stages of wound healing, including markers for granulocytes, macrophages, and dendritic cells, and to phenotypically analyze their activation (Figure 1). Material treatment resulted in an increase in cell infiltration, which peaked 7 days after injury and maintained half of the peak infiltration level until 42 days after injury. In the control treated with saline, cell infiltration peaked initially, by 3 days after injury (Figures 2A - 2C). Structurally, the ECM material degraded over time but was present until 41 days after injury, as observed by picrosirius red staining of collagen. Cells with granular cytoplasm and segmented nuclei associated with neutrophils were seen in the corona around the PE particles, which were subsequently replaced by a fibrous capsule. Cell infiltration into the material area correlated with the total number of cells observed during flow cytometry analysis (Figures 2A - 2C).

[0075] A. The implanted materials mobilize diverse innate immune compartments, with a granulocyte shift from eosinophil-dominant to neutrophil-dominant repertoires by fibrous materials. Flow cytometry showed that various sets of innate immune cells were mobilized to the injured microenvironment by 7 days after injury, which was dependent on material treatment and visualized by dimensionality reduction (Figs. 3A-3D). Granulocytes such as neutrophils, basophils, and eosinophils, as well as mature macrophages and immature monocyte-like bone marrow cells, dendritic cells, and other immune cells (CD45+Lin-) (Fig. 3A) were identified by manual gating. Comparison of responses to different materials revealed differences in the immune repertoire by 7 days after injury (Fig. 3B). Different subpopulations of macrophages and dendritic cells were identified, and the presence of granulocyte and monocyte-like cell populations was confirmed using the FlowSOM algorithm, a self-organizing map (SOM) algorithm for generating clusters based on the expression of markers detected by flow cytometry (Figs. 3C-3D).

[0076] As described above, a high abundance of macrophages (F4 / 80+CD68+) that persisted throughout the injury recovery process was observed (Figs. 4A - 4E). Furthermore, preferential recruitment of neutrophils (Ly6G+) to fibrosis - promoting PE - treated muscle injury was seen. The regeneration - promoting material generated an eosinophil - dominant granulocyte compartment, which had higher autofluorescence than neutrophils and may be due to granulation or phagocytosis of extracellular matrix materials. CD200R3+ basophils were preferentially recruited to untreated control injuries and reached a peak between 7 and 21 days post - injury, while eosinophils in ECM - treated injuries persisted until 7 - 42 days post - injury, and neutrophils in PE - treated injuries reached a peak by 7 days post - injury and slowly declined until 42 days post - injury while maintaining the majority of total immune cells in the microenvironment. Both control and ECM - treated injuries recruited neutrophils initially, but they disappeared by 7 days post - injury. PE - treated injuries recruited a higher level of monocytes compared to other treatments and were preferentially CX3CR1+ cells, which may represent the activation of a pathogenic type 2 immune response that promotes fibrosis as neutrophilic inflammation begins to subside. Dendritic cells were present and persisted at a low proportion (less than 4% of total CD45+ cells) over the time course of the response to injury and material implantation. Macrophages reached a peak initially and began to decrease proportionally with time, shifting mainly from CD11b+F4 / 80+CD68+ cells to CD11b+CD68+F4 / 80 - cells by 42 days post - injury.

[0077] B. CD103+XCR1+ dendritic cells are enriched by the regeneration - promoting scaffold Since prior studies have suggested an adaptive immunity in the integration and regeneration / fibrosis processes of biomaterials, the phenotype of antigen-presenting cells was further investigated. Most of the MHCII+ immune (CD45+) cells in the wound microenvironment were F4 / 80+ macrophages (Figures 5A and 5B). Macrophages in ECM-tx muscle injury had higher levels of CD206, CD301b, and CD169 expression, suggesting local proliferation of tissue-resident cells with type 2 polarization, which is characteristic of Th2-driven inflammation (Figures 6A - 6C). Although representing only 1 - 2% of total CD45+ immune cell infiltration, the second most common APC in the wound space was dendritic cells (CD11c+CD11b lo / neg ). Identification of cross-presenting dendritic cells was determined by the expression of XCR1, a chemokine receptor, and CD103 on dendritic cells (Figures 7A - 7H, Figure 8). CD11b - CD11c + MHCII hi Macrophages also expressed low levels of CD103 and XCR1, but significantly lower than CD11c+CD11b + F4 / 80 + dendritic cells (Figures 8, 9A - 9C). Type 1 conventional dendritic cells (cDC1) were enriched by the regeneration-promoting scaffold, while the fibrosis-promoting scaffold mainly recruited double-negative cells in a pattern that persisted until 42 days after injury (Figures 7A and 7B). cDC1 expressed intermediate levels of the co-stimulatory molecule CD8α, while double-negative cells showed a bimodal distribution with a subset of dendritic cells that may correlate with plasmacytoid DC (pDC) (Figure 10). Neither population expressed CD8α in muscle tissue. Double-negative cells and XCR1 single-positive cells increased proportionally with time in all groups (Figures 7B and 7C), but XCR1 lo CD103 hi dendritic cells (Figure 8, Figures 9A - 9C). Type 1 conventional dendritic cells (cDC1) were enriched by the regeneration-promoting scaffold, while the fibrosis-promoting scaffold mainly recruited double-negative cells in a pattern that persisted until 42 days after injury (Figures 7A and 7B). cDC1 expressed intermediate levels of the co-stimulatory molecule CD86, while double-negative cells showed a bimodal distribution with a subset of dendritic cells that may correlate with plasmacytoid DC (pDC) (Figure 10). Neither population expressed CD8α in muscle tissue. Double-negative cells and XCR1 single-positive cells increased proportionally with time in all groups (Figures 7B and 7C), but XCR1 + CD103 +Conventional dendritic cells reach their peak ratio 3 to 7 days after injury and their count reaches its peak 7 days after injury (Figures 7B and 7C). Throughout the process of wound healing, there were few CD103 single-positive dendritic cells in the wound space (Figures 7B and 7C). When evaluating the cell population by dimensionality reduction and hierarchical clustering algorithms, the FlowSOM algorithm identified this cell type as a distinct cluster, and this cell type was mapped to islands by visualization with both t-SNE and UMAP (Figures 7D and 7E). When applying FlowSOM to the three treatment groups within the same clustering population, more than 70% of the cells within the cDC1 cluster were derived from ECM-treated injuries, while only 15% and 6% were derived from PE-treated and control injuries, respectively (Figures 7F - 7H). The identification of these cells was confirmed in muscle injuries of two C57BL / 6 mouse littermates and Lewis rats, suggesting that this is reproducible and applicable to multiple species of rodents (Figures 11A - 11B).

[0078] Seven days after injury, when CD103+XCR1+ dendritic cells reached their peak in muscle tissue, these dendritic cells were detected at the skin incision site covering the ECM-treated muscle injury, but at a significantly lower rate. This is probably because most of the skin wounds had closed by this time (Figure 12).

[0079] Outside of the local muscle tissue, dendritic cells were detected in the blood and lymph nodes of injured mice. These DCs (CD11b- / lo CD11c + MHCII hi ) had different expressions of markers (XCR1, CD103, and CD8a) related to antigen cross-presentation ability depending on the location (Figures 13A - 13B). As described above, in muscle, the cells expressed both XCR1 and CD103 at high levels and did not express CD8a. In lymph nodes, these cells expressed all three markers of XCR1, CD103, and CD8a, while in blood, most cells were XCR1 - CD103 - CD8a- でThere was. Interestingly, in the blood of PE-treated mice, CD11c + MHCII hi The proportion of DCs was higher, and most of these cells were B220 / CD45R + which suggests a potential preference for the mobilization of circulating pDCs in response to fibrosis-promoting material implants (Figs. 14A - 14C). In the lymph nodes, there was no significant difference in the proportion of dendritic cells, but PE-treated mice had a higher proportion of TCRγδ+ T cells, which is consistent with previous literature on the role of these cells in fibrotic diseases (Fig. 14A). Regarding the chemokines that bind to XCR1, there was no significant difference between treatment groups in the blood XCL-1 levels 7 and 21 days after injury, but in all treatment groups, the amount of blood XCL-1 was significantly higher at the initial stage of the response to the wound (Fig. 15).

[0080] C. The phenotype and mobilization of CD103+XCR1+ dendritic cells depend on adaptive immune activity Dendritic cells can communicate directly with adaptive immune cells such as T cells and B cells through antigen presentation. Therefore, the inverse relationship and role of adaptive immunity in the mobilization and activation of these cells in the scaffold microenvironment were investigated. In RAG-deficient mice lacking T cells and B cells (Rag1 - / - ), a similar proportion of dendritic cells was mobilized into the whole environment, and there was a tendency to be less in Rag1 - / - but XCR1+CD103+cDC1 was significantly less (Figs. 16A - 16F). When evaluating their mobilization based on the expression of XCR1 or CD103, the loss of CD103 expression was the main cause of the loss of these cDC1s.

[0081] Similar to macrophages, in wild-type mice, dendritic cells had high levels of CD301b and CD206 in the presence of the ECM scaffold (Figs. 16G and 16H), and CD103+DCs expressed the highest levels of CD206 compared to other DC subtypes (Figs. 16I&J). ECM scaffold-mediated type 2 upregulation was lost in both CD301b and CD206 in Rag1 - / - mice compared to wild-type mice (Figs. 16G - 16J).

[0082] Example 2. Investigation of the Involvement of T Cells in Material Response A. Initial T cell activation and subsequent induction of regulatory CD8+ T cells are enhanced by regenerative scaffold treatment A 19-color flow cytometry panel was developed to evaluate the behavior of lymphocytes in blood and draining regional lymph nodes (Figure 17). Seven days after injury, the prevalence of CD44+CD62L- CD4 and CD8 T cells in the draining regional lymph nodes was increased in the ECM-treated injury compared to the PE-treated injury. Activated CD4+ T cells in the blood were enriched by ECM treatment 7 days after injury and correlated with prior studies showing a peak in IL-4 expression in the draining regional lymph nodes. This correlated with a low percentage of HELIOS+ regulatory CD4 and CD8 T cells in the draining regional lymph nodes 7 days after injury. In peripheral blood, the prevalence of CD4+ HELIOS+ iTreg was slightly higher compared to control injury, which was higher 21 days after injury. In all treatments, the percentage of HELIOS+ CD8 T cells tended to be higher 21 days after injury compared to 7 days after injury, but was not significant after correction for multiple comparisons (Figures 18A - 18F, Figure 19). In addition to these regulatory cells, the inventors identified a higher percentage of ST2+ regulatory B cells in the ECM-tx injury and a higher percentage of ST2+ pDC in both the ECM-tx and PE-tx injuries when compared to controls. Whether these cells functionally play a regulatory or pathogenic role is unknown. (Figures 20A and 20B).

[0083] B. CD103+XCR1+ adaptive immune cells are induced by trauma and modified by material treatment In addition to these regulatory T cells, a subpopulation of CD103+XCR1+ adaptive immune cells induced by trauma was found (Figure 21A, Figure 22). This population was present in B cells, CD4+ T cells, CD8+ T cells, and γδ T cells and increased transiently over time (Figure 21B). In B cells and CD4+ T cells, most of this population was CD62L- even without injury. For CD8+ and γδ T cells, activation increased over time, with CD8 reaching a peak by 7 days after injury and γδ T cells increasing over 21 days after injury (Figure 21C). In addition to the CD103+XCR1+ population, a CD103 lo XCR1- population, most prominent in CD8+ T cells and γδ T cells, was also present (Figure 21A and 21D). Activation of these cells, determined by loss of CD62L expression, peaked by 7 days after injury (Figure 21E). Interestingly, all of these cells were CD44 lo in both blood and draining regional lymph nodes, suggesting possible antigen-independent activation, although more work is needed to test this hypothesis (Figure 23). Since multiple activated adaptive immune cells were present in sterile injury, these CD103- and XCR-positive adaptive immune cells were evaluated to determine whether they behaved regulatoryly. The proportion of HELIOS+ iTregs that were CD103+XCR1+ was found to increase over time more than that of FoxP3+HELIOS- Tregs (Figures 24A-24C). This pattern holds for both CD4+ iTregs and CD8+ iTregs.

[0084] When comparing the draining regional lymph nodes with peripheral blood, a higher proportion of circulating CD103+XCR1+ adaptive immune cells was observed (Figure 25). In all treatment groups, B cells had the highest proportion of CD103+XCR1+ cells 21 days after injury, followed by γδ T cells, except for ECM-treated mice in which the proportion of these cells in circulation was low. Furthermore, 7 days after injury when the mobilization of CD103+XCR1+ dendritic cells peaked, a strong enrichment of CD103+XCR1+ T cells was observed in the wound space compared to blood and draining regional lymph nodes, suggesting a preferential mobilization of these cells into the wound space.

[0085] C. Upregulation of Xcl1, E-cadherin, and TGFβ-related signaling in the injured microenvironment as potential mediators of CD103+ cell mobilization Seven days after injury, significantly higher Xcl1 mRNA expression was observed in ECM-treated muscle injury compared to uninjured controls (Figure 6A). Within the muscle region, CD103+ cells were readily identified in ECM-treated injuries in various morphologies associated with both myeloid and lymphoid cells. These cells were located both at the injury interface, within the implant, and around the capsule (Figure 26B). In certain cases, these cells were found within clusters of immune infiltration but were also dispersed throughout the injury site. Furthermore, there was upregulation of E-cadherin, a ligand for integrin CD103, at the injury interface compared to more distal uninjured muscle tissue (Figure 26C). This was true for all treatment groups but was most significant with biomaterial implantation. Both ECM and PE induced this and were highly present not only in injured muscle tissue but also in hematoxylin-dense areas of immune infiltration (Figure 26D).

[0086] Assessing the cytokine and chemokine milieu via protein arrays revealed significant differences in immune profiles depending on the type of treatment as well as the presence or absence of injury. Material implantation correlated with increased CCL6 production by neutrophils and macrophages enriched by these materials. Both PE and ECM treatments induced upregulation of myeloperoxidase (MPO), which has been associated with reactive oxygen species (ROS) generation. Interestingly, injury upregulated endoglin, a part of the TGFβ receptor, and TGFβ is a known inducer of CD103 expression, compared with uninjured controls (Figure 27).

[0087] Example 3. Investigation of NK cells involved in material responses To further evaluate the immune response to biomaterials in injury, mice had a 3 mm defect in the quadriceps muscle and three conditions were investigated: untreated (control), polyethylene (PE) was implanted into the injury site to induce a profibrotic response, or extracellular matrix (ECM) was used to promote a pro-regenerative environment. Flow cytometry, ELISA, and RT-PCR were then used to evaluate the immune response at each site of injury. Seven days after injury, NK cells (CD49b + TCRβ - ) was increased in the ECM environment compared to PE and control (2.498% vs. 0.216%, and 0.0734% ± 0.017, P < 0.0001). Gene expression analysis showed that these NK cells expressed high levels of Tgfb1 (Figure 29A). These NK cells expressed XCR1, which is capable of antigen cross-presentation. + CD103 + This was accompanied by an enrichment of conventional dendritic cells (cDC1). Furthermore, ECM treatment induced an increase in XCL-1 levels in the injury microenvironment and peripheral blood (Figure 29B). - / - There was no significant difference in XCL-1 levels between mice and rats, suggesting that XCL-1 secretion is mediated by RAG1-independent cell types such as NK cells. This may be due to the XCR1 expression that promotes regenerative behavior. + CD103 + This correlated with an increase in dendritic cells. - / -In the absence of these cells in the mouse, there were physical symptoms such as necrotic muscle fibers and giant cells more distal from the site of injury, indicating an expansion of trauma beyond the initial injury.

[0088] NK cells were also contacted with fragments of decellularized extracellular matrix (ECM) or low molecular weight hyaluronic acid (LMW-HA) in vitro, and the level of Xcl1 production was measured. After 24 hours of exposure, upregulation of Xcl1 was observed, suggesting that the binding of NK cells to damage-associated molecular patterns mediates Xcl1 secretion.

[0089] When these data were evaluated together, within the first week after injury, in addition to XCL-1, XCR1 + CD103 + dendritic cells and CD49b + NK cells were upregulated. The presence of XCL-1 provides a potential mechanism for + CD103 + cell recruitment. In Rag1 - / - mice, the concentration of XCL-1 was not significantly different compared to wild type, indicating that RAG-independent recruitment is the source of XCL-1. NK cells were found to be present at a higher percentage at the ECM injury site compared to control and PE environments.

[0090] Discussion of Results The results demonstrate the induction of cross-presenting DCs by the regeneration-promoting material in trauma. This is accompanied by MHCII-bearing M2 macrophages, as well as peripheral CD8+ iTregs and ST2+ regulatory B cells, and CD103+XCR1+ adaptive immune cells induced by trauma. The mobilization of cross-presenting dendritic cells and the activation of CD103+XCR1+ CD8 T cells peak early in the response to injury and reach maximum by 7 days after injury. Upregulation of both Xcl1 gene expression in homogenized injured tissue and E-cadherin within the muscle injury site is detected, particularly at the injury-material interface and in the capsule surrounding the material, which may function as a mechanism for the mobilization of these CD103+ cells that may bind to upregulated E-cadherin for migration to the injury site. Furthermore, Rag1 - / - mice have low CD103 expression levels on dendritic cells, and T reg Since TGFβ secreted by T is known to induce CD103 upregulation, this presents a potential mechanism for CD103 upregulation in DCs by injury-related TGFβ secretion by adaptive immune cells.

[0091] These data indicate the balance of antigen cross-presentation during wound recovery, modulated by artificial material implantation. In the context of materials that promote type 2 and regulatory immune responses, such as in the context of decellularized ECM scaffolds, this cross-presentation occurs in an environment more suitable for peripheral tolerance and antigen-specific wound repair. In the context of inflammation-promoting and fibrosis-promoting materials such as PE that favor type 1 and type 17 immune responses, this can lead to the activation of autoimmunity and the formation of autoreactive T cells and antibodies, resulting in distal lesions and the systemic immune dysregulation reported in some patients.

[0092] The presence of CD103+XCR1+ innate immune cells and adaptive immune cells may present homeostatic control of responses to damaged self that expands during trauma after reaction with cross-presenting dendritic cells (Figs. 28 and 30). Prior studies have shown that XCR1+ T cells can be induced via trogocytosis and communication with cross-presenting dendritic cells and are potential targets for cancer immunotherapy. The ECM scaffold introduces a protein source for new exogenous antigens, but both sham injury and PE-treated injury are surgically induced sterile injuries that do not introduce non-self antigens, and thus these cells may be reacting to self-antigens or reacting antigen-independently. Cells and pathways that communicate with cross-presenting dendritic cells have not been previously described in the context of trauma and biomaterial implantation, and explain a novel mechanism of injury and immune responses to damaged self due to trauma.

Claims

**Claim 1** A therapeutic composition for use in treating a wound of an individual, wherein the therapeutic composition comprises a therapeutic agent that induces a regeneration-promoting environment within the wound and / or within the tissue surrounding the wound, and the therapeutic agent comprises a decellularized extracellular matrix (ECM) or a component derived therefrom, said therapeutic composition. **Claim 2** The therapeutic composition according to claim 1, wherein the component derived therefrom comprises a degradation product of the ECM. **Claim 3** The therapeutic composition according to any one of claims 1 or 2, wherein the therapeutic agent comprises one or more components selected from the group consisting of collagen, laminin, fibronectin, elastin, chondroitin sulfate, heparan sulfate, keratan sulfate, hyaluronic acid, perlecan, agrin, and degradation products thereof. **Claim 4** The therapeutic composition according to any one of claims 1 to 3, wherein the therapeutic agent comprises a matricain or a damage-associated molecular pattern (DAMP). **Claim 5** The therapeutic composition according to claim 4, wherein the matricain comprises one selected from the group consisting of metastatin, arresten, canstatin, tetrastatin, pentastatin, lumican, hexastatin, endostatin, neostatine, anastellin, sibstatin, PEX, endorepellin, CUB1CUB2 domain, Ten / 2, Ten11 / 12 / 13, Ten14, kappa elastin, the ectodomain of syndecan-1, the ectodomain of syndecan-2, the ectodomain of syndecan-3, the ectodomain of syndecan-4, elastokine, laminin peptide A13, laminin peptide C16, laminin 332 (laminin 5), DGGYY peptide, GHK tripeptide, VGVAPG peptide, PGP tripeptide, acetylated PGP tripeptide (AcPGP), tenascin-C (TNC), the G3 domain of nidogen-1, and tumstatin. **Claim 6** The therapeutic composition according to any one of claims 1 to 8, wherein the therapeutic agent induces local proliferation of M2 macrophages within the wound and / or within the tissue surrounding the wound. **Claim 7** The therapeutic composition according to any one of claims 1 to 9, wherein the therapeutic agent induces local proliferation and / or mobilization of conventional dendritic cells (cDC1) within the wound and / or within the tissue surrounding the wound. **Claim 8** The therapeutic composition according to claim 10, wherein the cDC1 is a cross-presenting dendritic cell.

9. The cDC1 is XCR1 ＋ CD103 ＋ The therapeutic composition according to any one of claims 9 or 10, which is a dendritic cell.

10. The therapeutic composition according to any one of claims 1 to 12, wherein the therapeutic agent comprises a peptide, a protein, a glycoprotein, a lipoprotein, a lipid, a sugar, a polysaccharide, a glycosaminoglycan, a nucleic acid molecule, an organic molecule, or a combination thereof.

11. A therapeutic composition for mobilizing NK cells to a wound or tissue proximal to the wound, the therapeutic composition comprising a therapeutic agent that induces a regeneration-promoting environment in and / or around the tissue of the wound, the therapeutic agent comprising a decellularized extracellular matrix (ECM), or a component derived therefrom, the therapeutic composition.

Images