Abstract:
The present invention generally relates to compositions and methods for topical or transdermal delivery, for promoting wound healing, reducing scarring, and/or promoting hair growth. Specifically, the present disclosure is directed to a topical arginine silicate formulation that demonstrates surprising improvement in wound healing time leading to more advantageous recovery and reducing hospital costs. Such formulations may also lessen scarring. The formulations may, for example, be applied to skin or to mucous membranes to facilitate wound healing.

Description:
BACKGROUND 
       [0001]    Field 
         [0002]    The present invention generally relates to compositions and methods for topical or transdermal delivery, for promoting wound healing, reducing scarring, and/or promoting hair growth. 
         [0003]    Description of the Related Art 
         [0004]    When barriers are broken, infections occur. Surgery, trauma (for example, scrapes, lacerations, punctures, abrasions, and burns), medical instrumentation (for example, catheterization, ventilation), chronic wounds (for example, diabetic foot ulcers) and a variety of diseases disrupt our natural barriers of defense. Wounds may become contaminated with microbes and may provide an excellent environment for microbial growth. Bleeding and leakage may provide fluids and nutrients that ultimately support microbial growth. Bacterial or fungal colonization, and/or overt infection may occur. Various surgical, trauma and medical settings all involve disruption of our natural barriers of defense and deserve special attention because the outcomes can range from rapid cure, to cure with heavy scarring, to lethal sepsis. 
         [0005]    Antimicrobial treatment of early infections may alter the course of the infection, resulting in more resistant and more dangerous infections. Common antimicrobial strategies focus on the use of selective antibiotics (for example, penicillin for gram-positive organisms) in order to avoid the development of bacteria that are resistant to broad-spectrum antibiotics. Inadvertently, this important strategy can have negative outcomes on an individual patient, where targeted antibiotics result in the emergence of an aggressive, different microorganism (for example,  Pseudomonas ). In this way, treated wounds can become the site for a “parade of pathogens”, where an early, dominant microbial species (for example,  Staph aureus ) is replaced by a second (for example, MRSA, methicillin-resistant  Staph aureus ) and, perhaps, even a third and fourth microbial species (for example, a multi-drug resistant gram negative species). 
         [0006]    Ultimately, the original natural barrier is the ideal defense against such infections. Current methods of wound closure, such as suturing and grafting have limited success; both methods leave the subject vulnerable to infection, and the subject will likely have significant scarring resulting from these procedures. Accordingly, improved methods for promoting wound healing are needed. 
         [0007]    Arginine silicate can be produced by combining arginine, a silicate salt and inositol. Although the products described herein may contain other agents in addition to arginine and silicate, they are referred to throughout the specification as “arginine silicate.” Arginine silicate can be synthesized, for example, by reacting arginine (free base), potassium silicate and (optionally) inositol. Thus, the compositions described herein may contain arginine, silicate, and inositol—which may be referred to throughout the specification as “inositol-stabilized arginine silicate,” “arginine silicate,” “arginine silicate inositol,” “ASI,” “arginine-silicate-inositol complex,” or “complex.” 
       SUMMARY 
       [0008]    The present disclosure is directed to a topical arginine silicate formulation that demonstrates surprising improvement in wound healing time leading to more advantageous recovery and reducing hospital costs. Such formulations may also lessen scarring. The formulations may, for example, be applied to skin or to mucous membranes to facilitate wound healing. 
         [0009]    Embodiments disclosed herein relate to the use of compositions comprising, consisting essentially of, or consisting of arginine, a silicate salt and inositol, arginine-silicate-inositol complexes, or combinations thereof, including pharmaceutically acceptable salts, hydrates, solvates, or mixtures thereof for topical application and for use in, for example, wound healing treatments, treatments to reduce wound scarring, or treatments to reduce inflammation. Embodiments disclosed herein also relate to the use of compositions comprising, consisting essentially of, or consisting of arginine, a silicate salt and inositol, arginine-silicate-inositol complexes, or combinations thereof, including pharmaceutically acceptable salts, hydrates, solvates, or mixtures thereof for topical application and for use in, for example, simulating and/or promoting hair growth. 
         [0010]    In some embodiments, the formulation is a gel. In some embodiments, the formulation is a cream. In some embodiments, the formulation is a lotion. In some embodiments, the formulation is an ointment. In some embodiments, the formulation is a salve. In some embodiments, the formulation is a balm. In some embodiments, the formulation is a suspension. In some embodiments, the formulation is an emulsion. In some embodiments, the formulation is a foam. In some embodiments, the formulation is a solution. In some embodiments, the formulation is embedded, dispersed, coated, or deposited onto or in an adhesive patch. In some embodiments, the formulation is embedded, dispersed, coated, or deposited onto or in a solid “stick,” (that can be rubbed or sprayed onto the skin). 
         [0011]    In some embodiments, the formulation is a hair care product. In some embodiments, the formulation is a shampoo. In some embodiments, the formulation is a conditioner. In some embodiments, the formulation is a leave-in conditioner. In some embodiments, the formulation is a mousse. In some embodiments, the formulation is a pomade. In some embodiments, the formulation is a hair spray. 
         [0012]    In some embodiments, the formulation contains from about 2% to about 8% arginine silicate (w/w), from about 2% to about 7% arginine silicate (w/w), from about 2% to about 6% arginine silicate (w/w), from about 2% to about 5% arginine silicate (w/w), from about 2% to about 4% arginine silicate (w/w), from about 3% to about 4% arginine silicate (w/w), or about 4% arginine silicate(w/w). 
         [0013]    In some embodiments, the formulation contains about 2.1% arginine silicate (w/w). In some embodiments, the formulation contains about 2.2% arginine silicate (w/w). In some embodiments, the formulation contains about 2.3% arginine silicate (w/w). In some embodiments, the formulation contains about 2.4% arginine silicate (w/w). In some embodiments, the formulation contains about 2.5% arginine silicate (w/w). In some embodiments, the formulation contains about 2.6% arginine silicate (w/w). In some embodiments, the formulation contains about 2.7% arginine silicate (w/w). In some embodiments, the formulation contains about 2.8% arginine silicate (w/w). In some embodiments, the formulation contains about 2.9% arginine silicate (w/w). In some embodiments, the formulation contains about 3.0% arginine silicate (w/w). In some embodiments, the formulation contains about 3.1% arginine silicate (w/w). In some embodiments, the formulation contains about 3.2% arginine silicate (w/w). In some embodiments, the formulation contains about 3.3% arginine silicate (w/w). In some embodiments, the formulation contains about 3.4% arginine silicate (w/w). In some embodiments, the formulation contains about 3.5% arginine silicate (w/w). In some embodiments, the formulation contains about 3.6% arginine silicate (w/w). In some embodiments, the formulation contains about 3.7% arginine silicate (w/w). In some embodiments, the formulation contains about 3.8% arginine silicate (w/w). In some embodiments, the formulation contains about 3.9% arginine silicate (w/w). In some embodiments, the formulation contains about 4.0% arginine silicate (w/w). In some embodiments, the formulation contains about 4.1% arginine silicate (w/w). In some embodiments, the formulation contains about 4.2% arginine silicate (w/w). In some embodiments, the formulation contains about 4.3% arginine silicate (w/w). In some embodiments, the formulation contains about 4.4% arginine silicate (w/w). In some embodiments, the formulation contains about 4.5% arginine silicate (w/w). In some embodiments, the formulation contains about 4.6% arginine silicate (w/w). In some embodiments, the formulation contains about 4.7% arginine silicate (w/w). In some embodiments, the formulation contains about 4.8% arginine silicate (w/w). In some embodiments, the formulation contains about 4.9% arginine silicate (w/w). In some embodiments, the formulation contains about 5.0% arginine silicate (w/w). In some embodiments, the formulation contains about 5.1% arginine silicate (w/w). In some embodiments, the formulation contains about 5.2% arginine silicate (w/w). In some embodiments, the formulation contains about 5.3% arginine silicate (w/w). In some embodiments, the formulation contains about 5.4% arginine silicate (w/w). In some embodiments, the formulation contains about 5.5% arginine silicate (w/w). In some embodiments, the formulation contains about 5.6% arginine silicate (w/w). In some embodiments, the formulation contains about 5.7% arginine silicate (w/w). In some embodiments, the formulation contains about 5.8% arginine silicate (w/w). In some embodiments, the formulation contains about 5.9% arginine silicate (w/w). In some embodiments, the formulation contains about 6.0% arginine silicate (w/w). In some embodiments, the formulation contains about 6.1% arginine silicate (w/w). In some embodiments, the formulation contains about 6.2% arginine silicate (w/w). In some embodiments, the formulation contains about 6.3% arginine silicate (w/w). In some embodiments, the formulation contains about 6.4% arginine silicate (w/w). In some embodiments, the formulation contains about 6.5% arginine silicate (w/w). In some embodiments, the formulation contains about 6.6% arginine silicate (w/w). In some embodiments, the formulation contains about 6.7% arginine silicate (w/w). In some embodiments, the formulation contains about 6.8% arginine silicate (w/w). In some embodiments, the formulation contains about 6.9% arginine silicate (w/w). In some embodiments, the formulation contains about 7.0% arginine silicate (w/w). In some embodiments, the formulation contains about 7.1% arginine silicate (w/w). In some embodiments, the formulation contains about 7.2% arginine silicate (w/w). In some embodiments, the formulation contains about 7.3% arginine silicate (w/w). In some embodiments, the formulation contains about 7.4% arginine silicate (w/w). In some embodiments, the formulation contains about 7.5% arginine silicate (w/w). In some embodiments, the formulation contains about 7.6% arginine silicate (w/w). In some embodiments, the formulation contains about 7.7% arginine silicate (w/w). In some embodiments, the formulation contains about 7.8% arginine silicate (w/w). In some embodiments, the formulation contains about 7.9% arginine silicate (w/w). In some embodiments, the formulation contains about 8.0% arginine silicate (w/w). Ranges of concentrations are also contemplated, and may include ranges from or between any two of the foregoing values. 
         [0014]    In some embodiments the arginine silicate is complexed. In some embodiments the arginine silicate is complexed with inositol. In some embodiments, the arginine silicate is not complexed. 
         [0015]    In some embodiments the molar ratio of arginine to silicate is between about 1:4 and 4:1. In some embodiments the molar ratio of arginine to silicate is between about 1:3 and 3:1. In some embodiments the molar ratio of arginine to silicate is between about 1:2 and 2:1. In some embodiments the molar ratio of arginine to silicate is about 1:1. 
         [0016]    In some embodiments the molar ratio of arginine to silicate to inositol is between about 1:1:1 and 3:3:2. In some embodiments the molar ratio of arginine to silicate to inositol is between about 1:1:1 and 2:2:3. In some embodiments the molar ratio of arginine to silicate to inositol is between about 1:1:1 and 2:2:1. In some embodiments the molar ratio of arginine to silicate to inositol is between about 1:1:1 and 2:2:1.5. In some embodiments the molar ratio of arginine to silicate to inositol is about 1:1:1. 
         [0017]    The actual dose of arginine silicate described herein depends on the specific compound, and on the specific composition and method of delivery. In some embodiments, a daily dose may be from about 1.0 mg/kg to about 2,500 mg/kg or more of body weight, from about 2.0 mg/kg or less to about 2,000 mg/kg, from about 3.0 mg/kg to about 1,500 mg/kg of body weight, from about 4.0 mg/kg to about 1,000 mg/kg of body weight, from about 5.0 mg/kg to about 500 mg/kg of body weight, from about 6.0 mg/kg to about 200 mg/kg of body weight, from about 7.0 mg/kg to about 100 mg/kg of body weight, from about 8.0 mg/kg to about 50 mg/kg of body weight, from about 9.0 mg/kg to about 20 mg/kg of body weight, or from about 10 mg/kg to about 15 mg/kg of body weight. 
         [0018]    In some embodiments the composition is applied once per day. In some embodiments the composition is applied at least twice per day. In some embodiments the composition is applied at least three times per day. In some embodiments the composition is applied at least four times per day. 
         [0019]    In some embodiments, the composition is applied directly to the wound. That is to say, in some aspects, the ASI containing composition is placed into direct contact with the wound area with no intervening materials or compositions in between the composition and the wound. In some embodiments, the composition is applied indirectly to the wound, e.g. on a bandage or patch. Thus, in some aspects, a patch or patch may include ASI as an active ingredient. In some embodiments, the composition is applied in the area surrounding the wound. In some embodiments, the composition is applied directly to the wound and to the areas surrounding the wound. In some embodiments, the wound is in skin. In other embodiments, the wound is in a mucous membrane. 
         [0020]    In some embodiments, wound healing time is decreased by about 5%, by about 10%, by about 15%, by about 20%, by about 25%, by about 30%, by about 35%, by about 40%, by about 45%, by about 50%, by about 55%, by about 60%, by about 65%, by about 70%, by about 75%, by about 80%, by about 85%, by about 90%, by about 95%, or by about 99%. 
         [0021]    In some embodiments, scarring is decreased by about 5%, by about 10%, by about 15%, by about 20%, by about 25%, by about 30%, by about 35%, by about 40%, by about 45%, by about 50%, by about 55%, by about 60%, by about 65%, by about 70%, by about 75%, by about 80%, by about 85%, by about 90%, by about 95%, or by about 99%. 
         [0022]    In some embodiments, the composition is applied to non-wounded tissue. In some embodiments, the composition is applied to the scalp. In some embodiments, the composition promotes hair growth. In some embodiments, hair growth is promoted at a shaved area. In some embodiments, hair growth is promoted after treatment with chemotherapy. In some embodiments, hair growth in promoted in balding areas. In some embodiments, the composition increases the rate of hair growth. In some embodiments, the composition increases hair thickness. Use for facilitating or promoting hair growth may be done in conjunction with wound healing or independent of wound healing, and may be applied to broken or unbroken skin to facilitate hair growth. 
         [0023]    The compositions disclosed herein may be used for decreasing wound healing time, decreasing wound scarring, promoting hair growth, and/or reducing inflammation. In some aspects, a method for decreasing wound healing time includes applying a topical composition comprising an effective amount of ASI to a wound, wherein the amount decreases wound healing time in comparison to not applying the composition. In some aspects, a method for decreasing wound scarring includes applying a topical composition comprising an effective amount of ASI to a wound, wherein the amount decreases scarring from the wound in comparison to not applying the composition. In some aspects, a method for promoting hair growth includes applying a topical composition comprising an effective amount of ASI to a treatment area on a mammalian subject, wherein the amount promotes hair growth in the treatment area in comparison to non-treatment areas. In some aspects, a method for reducing inflammation includes applying a topical composition comprising an effective amount of ASI to a treatment area on a mammalian subject, wherein amount reduces inflammation in the treatment area in comparison to non-treatment areas. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0024]      FIG. 1 : Is a set of photographs of the dorsal surface of test rats demonstrating decreased time to wound healing from a laceration upon treatment with a topical arginine-silicate-inositol composition (ASI). Representative rats from the control group (no treatment), ASI 4% group (topical application of 4.4% ASI (w/w)), and ASI %10 group (topical application of 10% ASI (w/w)) one day 0, 5, 10, and 15 are shown. 
           [0025]      FIG. 2 : Is a bar graph showing the effect of topical application of different doses of ASI on hydroxyproline concentration at 5th, 10th and 15th days after wound formation. 
           [0026]      FIG. 3 : Is a set of photographs of gels showing the effect of topical application of different doses of ASI on proteins involved in inflammation and wound healing at 5th, 10th and 15th days after wound formation. Each band is graphically depicted in  FIGS. 4-12  below:  FIG. 4 —inducible nitric oxide synthase (“iNOS”);  FIG. 5 —endothelial nitric oxide synthase (“eNOS”);  FIG. 6 —collagen;  FIG. 7 —matrix metalloproteinase-2 (“MMP-2”);  FIG. 8 —matrix metalloproteinase-9 (“MMP-9”);  FIG. 9 —vascular endothelial growth factor (“VEGF”);  FIG. 10 —endothelial growth factor (“EGF”);  FIG. 11 —and fibroblast growth factor (“FGF”). 
           [0027]      FIG. 4 : Is a bar graph showing the effect of topical application of different doses of ASI on inducible nitric oxide synthase (iNOS) levels at 5th, 10th and 15th days after wound formation. 
           [0028]      FIG. 5 : Is a bar graph showing the effect of topical application of different doses of ASI on endothelial nitric oxide synthase (eNOS) at 5th, 10th and 15th days after wound formation. 
           [0029]      FIG. 6 : Is a bar graph showing the effect of topical application of different doses of ASI on collagen levels at 5th, 10th and 15th days after wound formation. 
           [0030]      FIG. 7 : Is a bar graph showing the effect of topical application of different doses of ASI on matrix metalloproteinase-2 (MMP-2) levels at 5th, 10th and 15th days after wound formation. 
           [0031]      FIG. 8 : Is a bar graph showing the effect of topical application of different doses of ASI on matrix metalloproteinase-9 (MMP-9) levels at 5th, 10th and 15th days after wound formation. 
           [0032]      FIG. 9 : Is a bar graph showing the effect of topical application of different doses of ASI on vascular endothelial growth factor (VEGF) levels at 5th, 10th and 15th days after wound formation. 
           [0033]      FIG. 10 : Is a bar graph showing the effect of topical application of different doses of ASI on endothelial growth factor (EGF) levels at 5th, 10th and 15th days after wound formation. 
           [0034]      FIG. 11 : Is a bar graph showing the effect of topical application of different doses of ASI on fibroblast growth factor (FGF) levels at 5th, 10th and 15th days after wound formation. 
           [0035]      FIG. 12 : Is a set of photographs of gels showing the effect of topical application of different doses of ASI on expression levels of proteins involved in inflammation and wound healing at 5th, 10th and 15th days after wound formation. Each band is graphically depicted in  FIGS. 13-16  below:  FIG. 13 —NFκB;  FIG. 14 —TGF-β;  FIG. 15 —TNF-α;  FIG. 16 —IL-1β. 
           [0036]      FIG. 13 : Is a bar graph showing the effect of topical application of different doses of ASI on NFκB levels at 5th, 10th and 15th days after wound formation. 
           [0037]      FIG. 14 : Is a bar graph showing the effect of topical application of different doses of ASI on TGF-β levels at 5th, 10th and 15th days after wound formation. 
           [0038]      FIG. 15 : Is a bar graph showing the effect of topical application of different doses of ASI on TNF-α levels at 5th, 10th and 15th days after wound formation. 
           [0039]      FIG. 16 : Is a bar graph showing the effect of topical application of different doses of ASI on and IL-β levels at 5th, 10th and 15th days after wound formation. 
           [0040]      FIG. 17 : shows histology sections demonstrating the effect of topical application of ASI on wound healing. Representative sections from the control group (no treatment), ASI 4% group (topical application of 4.4% ASI (w/w)), and ASI %10 group (topical application of 10% ASI (w/w)) one day 0, 5, 10, and 15 are shown. 
       
    
    
     DESCRIPTION 
       [0041]    This disclosure provides compositions and methods of promoting wound healing, including topical and transdermal delivery of arginine silicate. The composition can take the form of a gel, a cream, a lotion, an ointment, a solution, a suspension, a mousse, an emulsion, a solid “stick,” etc., that can be rubbed or sprayed onto the skin, for example, wounded skin. Other aspects of the present invention are generally directed to methods of making or using such compositions, methods of promoting such compositions, and kits including such compositions. 
         [0042]    Arginine silicate inositol complex (“ASP”) is a composition of arginine, silicon and inositol that has been reported for its beneficial effects on vascular health. This disclosure reports the beneficial effect of the composition on wound healing in rats. 
         [0043]    For example, in rats with excision wounds, the granulation tissue was found to form significantly slower in the control group than in the ASI treated rats (4% or 10% ASI ointment (w/w), twice per day). The mean unhealed wound area was significantly smaller and the mean percentage of total wound healing was significantly higher in ASI-treated wounds than in the control wound. Hydroxyproline, collagen and matrix metalloproteinases were measured in the granulated tissue and found to be affected. Inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), collagen, matrix metalloproteinase 2 (MMP-2), MMP-9, vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and various cytokines (TNF-α and IL-1β) measured in this study showed a significant fall in expression level in ASI-treated wounds. The results indicate that a topical application of ASI has beneficial effects on the healing response of wounds. 
         [0044]    The healing of wounds involves a cascade of events characterized by the restructuring of the damaged tissue in an attempt to restore as normal a condition as possible. The process involves considerable complex factors involving the synthesis of the matrix components and cross talks/interactions between several factors such as the cytokines and growth factors. In this study, we studied a number of macroscopic, microscopic changes and the changes at the molecular level in excision wound treated with ASI at two different dose levels in a rather simple and reproducible animal model. 
         [0045]    Synthesis of hydroxyproline and the deposition of collagen observed in the proliferative stage of the healing process is a hallmark of wound healing. The hydroxyproline content, as well as the collagen determined at the site of wound, increased progressively in ASI treated animals on day 5, 10 and 15 after the injury when compared with control group. The results corroborated with the histology of the skin tissue from the treated rats. 
         [0046]    Tissue remodeling is regulated by proteinases and their inhibitors at the levels of expression, deposition, inhibition and activation. In particular, matrix metalloproteinase-9 (MMP-9) is believed to function in the remodeling of the basement membrane zone, because several extracellular matrix proteins in the basement membrane zone, such as type IV collagen, have been identified as substrates of this proteinase. 
         [0047]    MMPs are well known to increase during acute wound healing and it has been shown that a prolonged and excessive production of MMPs (MMP-9 in particular) leads to disordered wound healing. 
         [0048]    In this study, it was observed that MMP-2 and MMP-9 expression in wound sites decreased in ASI treated groups. MMP-2 and MMP-9 expression of ASI treated groups significantly decreased in 5, 10 and 15 days after injury compared with control group. MMP-9 synthesis and secretion are strictly controlled, and can be induced by a variety of physiological stimuli, including cytokines, chemokines and growth factors. Moreover, stress-related chemical and physical factors, such as X-ray radiation, UV light, pH change, and reactive oxygen species, are also known to modulate MMP-9 production in cells. 
         [0049]    Angiogenesis plays a major role in tissue repair and remodeling. Angiogenic factors, such as vascular endothelial cell growth factor (VEGF) or basic fibroblast growth factor (bFGF), enhance the proliferation and/or migration of endothelial cells (ECs) both in vitro and in vivo. These actions of ECs are associated with proteolytic degradation of the extracellular matrix (ECM), which enables ECs to migrate or invade the interstitial space. Plasminogen activators (PAs) convert inactive zymogen plasminogen to the active protease plasmin, which not only degrades fibrin and several ECM proteins, but also activates pro-collagenase to collagenase, a matrix metalloproteinase (MMP). Similarly, the growth factors such as PDGF, TGBβ, and bFGF are pivotal in normal wound repair, driving cell migration, protein synthesis, proliferation, matrix formation, and generally controlling the repair process. The biological activity of many of these growth factors is enhanced by the presence of specific matrix proteins. 
         [0050]    Increased vascular permeability has since been shown to occur during the early phases of wound repair, theoretically allowing deposition of the fibrin-rich matrix necessary for cellular migration. The identification of increased vascular permeability concomitant with increased VEGF production in skin wounds provided evidence for a role for VEGF in wound repair. VEGF production and VEGF-mediated angiogenic activity would rise in the early hypoxic wound and then fall when neovascularization is complete and wound perfusion is restored. 
         [0051]    Previous studies have attempted to correlate the levels of growth factors to the stage of wound healing. The observed levels of individual factors do appear to correspond to the stage of wound healing. It has been reported that the primary sources of surgical wound VEGF are the fibroblast and the macrophage. VEGF has been associated with angiogenesis in numerous pathological situations, including rheumatoid arthritis, tumor growth and, proliferative retinopathy, the angiogenic process being initiated by bFGF and maintained by VEGF. 
         [0052]    The pro-inflammatory cytokines IL-6, TNF-α, and IL-1β all showed high levels on day 1 corresponding to the initial inflammatory response. IL-6 levels then decreased down to day 8, TNF-α levels remained static, and IL-1β levels showed a second peak on postoperative day 6, which may correspond to its other functions in wound healing, for example, matrix synthesis and collagen production. In this study, postoperatively at the 5th, 10th and 15th days, VEGF, EGF and FGF levels were increased in ASI treated groups. These results indicate the role of VEGF in the wound healing. 
         [0053]    Inflammation results in a continuous generation of reactive species, such as the non-radical hydrogen peroxide or the superoxide radical. Nuclear Factor-κB (NF-κB) was determined to play an essential role in inflammation. NF-κB is activated by numerous different stimuli, including protein kinase C activators, cytokines, oxidants, and viruses. Excessive NF-κB activation has been implicated in many disease states associated with chronic inflammation, such as asthma, ulcerative colitis, and joint inflammation. It has been reported that inhibition of NF-κB may reduce inflammation by reducing the local NOx concentrations, and they showed that chronic administration of a recombinant adenovirus expressing an NF-κB super repressor significantly increases the amount of collagen accumulation in subcutaneously implanted polyvinyl alcohol sponges. 
         [0054]    In the current report, levels of NF-κB were significantly decreased and collagen deposition levels were significantly increased in ASI treated groups. The mechanism of this phenomenon may be from decreasing inflammation through the inhibition of NF-κB. NF-κB inhibition effect of ASI may have helped to enhance wound healing. 
         [0055]    In conclusion, the results of the present disclosure suggest that the effects of ASI on wound healing depended on the dose. Beneficial effects were evident after treatment with 4% ASI (w/w), but were lost after 10% ASI (w/w). The results suggest that topical application of 4% ASI (w/w) have beneficial effects on wound healing. 
       Administration and Compositions 
       [0056]    Pharmaceutically-acceptable carriers include, for example, solid or liquid fillers, diluents, hydrotropies, surface-active agents, and encapsulating substances. Optional pharmaceutically-active materials may be included, which do not substantially interfere with the activity of the compound. The amount of carrier employed in conjunction with the compound is sufficient to provide a practical quantity of material for administration per unit dose of the compound. One having ordinary skill in the art would understand additional techniques and compositions for making dosage forms useful in the methods described herein. 
         [0057]    The composition for topical and transdermal delivery can take the form of a gel, a cream, a lotion, an ointment, a solution, a suspension, an emulsion, or embedded, dispersed, coated, or deposited onto an adhesive patch or a solid “stick,” (that can be rubbed or sprayed onto the skin). Topical formulations may generally be comprised of a pharmaceutical carrier, co-solvent, emulsifier, penetration enhancer, preservative system, and emollient. Where the formulation is to be applied to mucous membrane, the formulation may advantageously include a thickener or mucoadhesive. 
         [0058]    The term “pharmaceutically acceptable carrier” or “pharmaceutically acceptable excipient” includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like. The use of such media and agents for pharmaceutically active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active ingredient, its use in the therapeutic compositions is contemplated. In addition, various adjuvants such as are commonly used in the art may be included. One having ordinary skill in the art would understand additional considerations for inclusion of various components in pharmaceutical compositions. 
         [0059]    Some examples of substances, which can serve as pharmaceutically-acceptable carriers or components thereof, are sugars, such as lactose, glucose and sucrose; starches, such as corn starch and potato starch; cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose, and methyl cellulose; powdered tragacanth; malt; gelatin; talc; solid lubricants, such as stearic acid and magnesium stearate; calcium sulfate; vegetable oils, such as peanut oil, cottonseed oil, sesame oil, olive oil, corn oil and oil of the obroma; polyols such as propylene glycol, glycerine, sorbitol, mannitol, and polyethylene glycol; alginic acid; emulsifiers, such as the TWEENS; wetting agents, such sodium lauryl sulfate; coloring agents; flavoring agents; tableting agents, stabilizers; antioxidants; preservatives; pyrogen-free water; isotonic saline; and phosphate buffer solutions. 
         [0060]    Compositions described herein may optionally include other active ingredients. 
         [0061]    Other compositions useful for attaining systemic delivery of the subject compounds include sublingual, buccal and nasal dosage forms. Such compositions typically comprise one or more of soluble filler substances such as sucrose, sorbitol and mannitol; and binders such as acacia, microcrystalline cellulose, carboxymethyl cellulose and hydroxypropyl methyl cellulose. Glidants, lubricants, sweeteners, colorants, antioxidants and flavoring agents disclosed above may also be included. 
         [0062]    A liquid composition, which is formulated for topical ophthalmic use, is formulated such that it can be administered topically to the eye. The comfort may be maximized as much as possible, although sometimes formulation considerations (e.g. drug stability) may necessitate less than optimal comfort. In the case that comfort cannot be maximized, the liquid may be formulated such that the liquid is tolerable to the patient for topical ophthalmic use. Additionally, an ophthalmically acceptable liquid may either be packaged for single use, or contain a preservative to prevent contamination over multiple uses. 
         [0063]    For ophthalmic application, solutions or medicaments are often prepared using a physiological saline solution as a major vehicle. Ophthalmic solutions may preferably be maintained at a comfortable pH with an appropriate buffer system. The formulations may also contain conventional, pharmaceutically acceptable preservatives, stabilizers and surfactants. 
         [0064]    Preservatives that may be used in the pharmaceutical compositions disclosed herein include, but are not limited to, benzalkonium chloride, PHMB, chlorobutanol, thimerosal, phenylmercuric, acetate and phenylmercuric nitrate. A useful surfactant is, for example, Tween 80. Likewise, various useful vehicles may be used in the ophthalmic preparations disclosed herein. These vehicles include, but are not limited to, polyvinyl alcohol, povidone, hydroxypropyl methyl cellulose, poloxamers, carboxymethyl cellulose, hydroxyethyl cellulose and purified water. 
         [0065]    Tonicity adjustors may be added as needed or convenient. They include, but are not limited to, salts, particularly sodium chloride, potassium chloride, mannitol and glycerin, or any other suitable ophthalmically acceptable tonicity adjustor. 
         [0066]    Various buffers and means for adjusting pH may be used so long as the resulting preparation is ophthalmically acceptable. For many compositions, the pH will be between 4 and 9. Accordingly, buffers include acetate buffers, citrate buffers, phosphate buffers and borate buffers. Acids or bases may be used to adjust the pH of these formulations as needed. 
         [0067]    In a similar vein, an ophthalmically acceptable antioxidant includes, but is not limited to, sodium metabisulfite, sodium thiosulfate, acetylcysteine, butylated hydroxyanisole and butylated hydroxytoluene. 
         [0068]    Other excipient components, which may be included in the ophthalmic preparations, are chelating agents. A useful chelating agent is edetate disodium, although other chelating agents may also be used in place or in conjunction with it. 
         [0000]    Methods of Promoting Wound Healing and/or Reducing Scarring 
         [0069]    Some embodiments of the present invention include methods of promoting wound healing with the compositions comprising compounds described herein. Some embodiments of the present invention include methods of reducing scarring with the compounds and compositions comprising compounds described herein. Some methods include administering a compound, composition, pharmaceutical composition described herein to a subject in need thereof. In some embodiments, a subject can be an animal, for example, a mammal, a human. 
         [0070]    “Subject” as used herein, means a human or a non-human mammal, for example, a dog, a cat, a mouse, a rat, a cow, a sheep, a pig, a goat, a non-human primate or a bird, for example, a chicken, as well as any other vertebrate or invertebrate. In some embodiments, the subject is a human. 
         [0071]    In some embodiments a subject in need of the present invention is a subject with an injury or wound to a tissue. Such tissues include, but are not limited to, epithelial tissue, epidermis, dermis, hypodermis, and subcutaneous connective tissue. The types of wounds include, but are not limited to, lesions, burns, lacerations, abrasions, surgical wounds, and punctures. 
         [0072]    Further embodiments include administering a combination of compounds to a subject in need thereof. A combination can include a compound, composition, pharmaceutical composition described herein with an additional medicament. 
         [0073]    Some embodiments include co-administering a compound, composition, and/or pharmaceutical composition described herein, with an additional medicament. By “co-administration,” it is meant that the two or more agents may be found in the patient&#39;s bloodstream at the same time, regardless of when or how they are actually administered. In one embodiment, the agents are administered simultaneously. In some embodiments, administration in combination is accomplished by combining the agents in a single dosage form. In some embodiments, the agents are administered sequentially. In some embodiments, the agents are administered through the same route, such as topically. In some embodiments, the agents are administered through different routes, such as one being administered topically and another being administered orally. 
         [0074]    An “effective amount” or a “therapeutically effective amount” as used herein refers to an amount of a therapeutic agent that is effective to relieve, to some extent, or to reduce the likelihood of onset of, one or more of the symptoms of a disease or condition, and includes curing a disease or condition. “Curing” means that the symptoms of a disease or condition are eliminated; however, certain long-term or permanent effects may exist even after a cure is obtained (such as extensive tissue damage). 
         [0075]    “Treat,” “treatment,” or “treating,” as used herein refers to administering a compound or composition to a subject for prophylactic and/or therapeutic purposes. The term “prophylactic treatment” refers to treating a subject who does not yet exhibit symptoms of a disease or condition, but who is susceptible to, or otherwise at risk of, a particular disease or condition, whereby the treatment reduces the likelihood that the patient will develop the disease or condition. The term “therapeutic treatment” refers to administering treatment to a subject already suffering from a disease or condition. 
         [0076]    To further illustrate this invention, the following examples are included. The examples should not, of course, be construed as specifically limiting the invention. Variations of these examples within the scope of the claims are within the purview of one skilled in the art and are considered to fall within the scope of the invention as described, and claimed herein. The reader will recognize that the skilled artisan, armed with the present disclosure, and skill in the art is able to prepare and use the invention without exhaustive examples. 
       EXAMPLES 
     Example 1 
       [0077]    Animals: In this study, 42 male, 4 months old, Wistar albino rats weighing between 250 and 300 g were used. Animals were housed at 21° C. with a day/night cycle of 12 h. During the study these animals were fed ad libitum standard rodent feed. Guidelines for the care and use of animals approved by the relevant institution were followed and the local ethics committee approved this study. 
         [0078]    Anesthesia: The rats were anaesthetized with single intramuscular injection of 6 mg/kg xylazine hydrochloride (Rompun, Bayer, 23.32 mg/ml) and 85 mg/kg ketamine hydrochlorure (Ketalar, Parke-Davis, 50 mg/ml). 
         [0079]    Test drugs: 4% and 10% arginine-silicate inositol (ASI)(w/w) ointment, and cold cream (Botafarma, 12.5% spermaceti+12% white wax+56% liquid paraffin+0.5% borate of soda+19% distilled water) were used in this study. 
         [0080]    Wound model: The dorsal surfaces of the rats were shaved and prepared with 10% antiseptic povidone-iodine solution (Kim-Pa, Poviiodeks, % 10 povidone-iodine). Then a disposable 1 cm diameter skin punch biopsy tool (Acuderm Inc., Fort Lauderdale, Fla.) was used to create a full-thickness excisional wound. 
         [0081]    The rats were divided into 2 main groups such that each group had 21 randomly selected rats. Each main group was divided within themselves into 3 subgroups of 7 rats each to perform postoperative 5, 10 and 14 days of examination. Immediately after the wounds were made, the wounded areas in the first (n=21) and second (n=21) was covered with cold cream (Botafarma, 12.5% spermaceti+12% white wax+56% liquid paraffin+0.5% borate of soda+19% distilled water) (Control) and 4% or 10% ASI ointment twice a day, respectively. These applications were repeated every day. The wounds were clinically observed in all groups every day. The rats that completed postoperative follow-up periods of 5 (7 rats from each group), 10 (7 rats from each group) and 14 days (7 rats from each group) will be sacrificed. 
         [0082]    Measurements: On every bandage change, the wound boundaries were traced on a transparent sheet with a fine-tipped permanent marker. These transparent sheets were scanned and the areas of wound sites and epithelization fronts were measured with the help of the paint-brush (PB) computer program. The data obtained was calculated with the following formula: Area (cm 2 )=P/K.M, where P is the value of a particular wound site obtained on the PB, K is a rate constant for expressing the values of the PB as cm 2 , and M is the magnitude of tracings after scanning. 
         [0083]    During the qualitative examination, the expansion rate, the day wound contraction started, the fraction of the wound healed with contraction, the rate of wound contraction, the day epithelization was first noticed, the fraction of wound healed with epithelization (final wound size) and the number of days in which wound healing was fully completed was evaluated. The statistical comparison was made with ANOVA. The results were considered significant at P&lt;0.05 and a 95% confidence interval (Kilic et al., 2002). These data are shown in Tables 1-5 and  FIGS. 1-16 . 
         [0084]    Histopathological examination: Burned skin tissue samples were collected after sacrificing the rats for histopathological examination purposes. These tissue samples were fixed in 10% neutral buffered formalin solution, embedded in paraffin wax, cut into 5 μm-thick sections and stained with haematoxyline-eosin and Masson&#39;s trichrome stain for examination by light microscopy. 
         [0085]    Histological scoring was assigned in a blinded manner as described previously (Greenhalgh et al., 1990; Asai et al., 2013). Briefly, each specimen was given a score of 1-12: 1-3, none to minimal cell accumulation and granulation tissue or epithelial migration; 4 to 6, thin, immature granulation dominated by inflammatory cells, but with few fibroblasts, capillaries or collagen deposition and minimal epithelial migration; 7-9, moderately thick granulation tissue, ranging from domination by inflammatory cells to more fibroblasts and collagen deposition; and 10-12, thick vascular granulation tissue dominated by fibroblasts and extensive collagen deposition. 
         [0086]    Statistical analyses: The thickness of granulation tissue was examined and recorded at the center of each wound. Statistically, all data are expressed in millimeters as mean±standard error. The differences between days 5, 10 and 14 was compared using the Mann Whitney U test. The differences between groups was compared using the Kruskal-Wallis test. Statements of statistical significance are based on P&lt;0.05. These analyses were carried out using SPSS statistical analysis system (Relase 10.0, SPSS. Inc.). 
         [0000]    
       
         
               
             
               
               
             
               
               
               
               
               
             
               
               
               
               
               
             
           
               
                 TABLE 1 
               
             
             
               
                   
               
               
                 Changes in wound sizes (mm 2 ) during the healing periods 
               
             
          
           
               
                   
                 Days of Measurements 
               
             
          
           
               
                 Groups 
                 0 (n = 21) 
                 5 (n = 21) 
                 10 (n = 14) 
                 15 (n = 7) 
               
               
                   
               
             
          
           
               
                 Control 
                 19,625 
                 15.33 
                 3.21 
                 1.86 
               
               
                 AS I - I 4% 
                 19,625 
                 8.90 
                 0.31 
                 0.17 
               
               
                 ASI - II 10% 
                 19,625 
                 9.52 
                 0.50 
                 0.29 
               
               
                   
               
               
                 Control: Control group no treatment. 
               
               
                 ASI 4%: Topical application of 4.4% ASI (w/w) 
               
               
                 ASI %10: Topical application of 10% ASI (w/w) 
               
             
          
         
       
     
         [0000]    
       
         
               
             
               
               
             
               
               
               
               
               
             
               
               
               
               
               
             
           
               
                 TABLE 2 
               
             
             
               
                   
               
               
                 Effect of topical application of ASI on wound area (%) 
               
             
          
           
               
                   
                 Days of Measurements 
               
             
          
           
               
                   
                 Groups 
                 5 (n = 21) 
                 10 (n = 14) 
                 15 (n = 7) 
               
               
                   
                   
               
             
          
           
               
                   
                 Control 
                 78.13 
                 16.38 
                 9.46 
               
               
                   
                 ASI - I 4% 
                 45.37 
                 1.57 
                 0.85 
               
               
                   
                 ASI - II 10% 
                 48.53 
                 2.55 
                 1.46 
               
               
                   
                   
               
               
                   
                 Wound area (%) = (Wound area at day X/Wound area at day 0) × 100. 
               
               
                   
                 Control: Control group no treatment. 
               
               
                   
                 ASI 4%: Topical application of 4.4% ASI (w/w) 
               
               
                   
                 ASI %10: Topical application of 10% ASI (w/w) 
               
             
          
         
       
     
         [0000]    
       
         
               
             
               
               
             
               
               
               
               
               
             
               
               
               
               
               
             
           
               
                 TABLE 3 
               
             
             
               
                   
               
               
                 Effect of topical application of ASI on percent of wound contraction 
               
             
          
           
               
                   
                 Days of Measurements 
               
             
          
           
               
                   
                 Groups 
                 5 (n = 21) 
                 10 (n = 14) 
                 15 (n = 7) 
               
               
                   
                   
               
             
          
           
               
                   
                 Control 
                 21.87 
                 83.62 
                 90.54 
               
               
                   
                 ASI - I 4% 
                 54.63 
                 98.43 
                 99.15 
               
               
                   
                 ASI - II 10% 
                 51.47 
                 97.45 
                 98.54 
               
               
                   
                   
               
               
                   
                 Percent of wound contraction on day X = 100 − percentage of wound area on day X 
               
               
                   
                 Control: Control group no treatment. 
               
               
                   
                 ASI 4%: Topical application of 4.4% ASI (w/w) 
               
               
                   
                 ASI %10: Topical application of 10% ASI. =(w/w) 
               
             
          
         
       
     
         [0000]    
       
         
               
             
               
               
             
               
               
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                 TABLE 4 
               
             
             
               
                   
               
               
                 Changes in wound sizes of three groups during the healing periods (mm 2 ). 
               
             
          
           
               
                   
                 Days of Measurements 
               
             
          
           
               
                 Groups 
                 Rats 
                 0 
                 5 
                 10 
                 15 
               
               
                   
               
             
          
           
               
                 Control 
                 1-A 
                 19,625 
                 15 
                 — 
                 — 
               
               
                   
                 1-A 
                 19,625 
                 7 
                 — 
                 — 
               
               
                   
                 1-A 
                 19,625 
                 11 
                 — 
                 — 
               
               
                   
                 1-A 
                 19,625 
                 9 
                 — 
                 — 
               
               
                   
                 1-B 
                 19,625 
                 42 
                 — 
                 — 
               
               
                   
                 1-B 
                 19,625 
                 12 
                 — 
                 — 
               
               
                   
                 1-B 
                 19,625 
                 10 
                 — 
                 — 
               
               
                   
                 2-A 
                 19,625 
                 19 
                 4 
                 — 
               
               
                   
                 2-A 
                 19,625 
                 18 
                 4 
                 — 
               
               
                   
                 2-A 
                 19,625 
                 17 
                 19  
                 — 
               
               
                   
                 2-A 
                 19,625 
                 7 
                 3 
                 — 
               
               
                   
                 2-B 
                 19,625 
                 16 
                 2 
                 — 
               
               
                   
                 2-B 
                 19,625 
                 21 
                 1 
                 — 
               
               
                   
                 2-B 
                 19,625 
                 25 
                 1 
                 — 
               
               
                   
                 3-A 
                 19,625 
                 9 
                 2 
                 1 
               
               
                   
                 3-A 
                 19,625 
                 13 
                 2 
                 1 
               
               
                   
                 3-A 
                 19,625 
                 10 
                 2 
                 1 
               
               
                   
                 3-A 
                 19,625 
                 17 
                 1 
                 3 
               
               
                   
                 3-B 
                 19,625 
                 20 
                 2 
                 3 
               
               
                   
                 3-B 
                 19,625 
                 12 
                 1 
                 1 
               
               
                   
                 3-B 
                 19,625 
                 12 
                 1 
                 3 
               
               
                 ASI - I 4% (w/w) 
                 4-A 
                 19,625 
                 16 
                 — 
                 — 
               
               
                   
                 4-A 
                 19,625 
                 5 
                 — 
                 — 
               
               
                   
                 4-A 
                 19,625 
                 10 
                 — 
                 — 
               
               
                   
                 4-A 
                 19,625 
                 7 
                 — 
                 — 
               
               
                   
                 4-B 
                 19,625 
                 13 
                 — 
                 — 
               
               
                   
                 4-B 
                 19,625 
                 9 
                 — 
                 — 
               
               
                   
                 4-B 
                 19,625 
                 12 
                 — 
                 — 
               
               
                   
                 5-A 
                 19,625 
                 10 
                 1 
                 — 
               
               
                   
                 5-A 
                 19,625 
                 8 
                 0 (Healed) 
                 — 
               
               
                   
                 5-A 
                 19,625 
                 4 
                 0 (Healed) 
                 — 
               
               
                   
                 5-A 
                 19,625 
                 6 
                 1 
                 — 
               
               
                   
                 5-B 
                 19,625 
                 5 
                 1 
                 — 
               
               
                   
                 5-B 
                 19,625 
                 9 
                 0 (Healed) 
                 — 
               
               
                   
                 5-B 
                 19,625 
                 10 
                 0 (Healed) 
                 — 
               
               
                   
                 6-A 
                 19,625 
                 14 
                 1 
                 1 
               
               
                   
                 6-A 
                 19,625 
                 14 
                 0 (Healed) 
                 0 (Healed) 
               
               
                   
                 6-A 
                 19,625 
                 15 
                 0 (Healed) 
                 0 (Healed) 
               
               
                   
                 6-A 
                 19,625 
                 7 
                 4 (apse)  
                 2 (apse)  
               
               
                   
                 6-B 
                 19,625 
                 4 
                 0 (Healed) 
                 0 (Healed) 
               
               
                   
                 6-B 
                 19,625 
                 4 
                 0 (Healed) 
                 0 (Healed) 
               
               
                   
                 6-B 
                 19,625 
                 5 
                 0 (Healed) 
                 0 (Healed) 
               
               
                 ASI - II 10% 
                 7-A 
                 19,625 
                 6 
                 — 
                 — 
               
               
                 (w/w) 
                 7-A 
                 19,625 
                 7 
                 — 
                 — 
               
               
                   
                 7-A 
                 19,625 
                 8 
                 — 
                 — 
               
               
                   
                 7-A 
                 19,625 
                 10 
                 — 
                 — 
               
               
                   
                 7-B 
                 19,625 
                 8 
                 — 
                 — 
               
               
                   
                 7-B 
                 19,625 
                 7 
                 — 
                 — 
               
               
                   
                 7-B 
                 19,625 
                 8 
                 — 
                 — 
               
               
                   
                 8-A 
                 19,625 
                 16 
                 0 (Healed) 
                 — 
               
               
                   
                 8-A 
                 19,625 
                 6 
                 1 
                 — 
               
               
                   
                 8-A 
                 19,625 
                 3 
                 0 (Healed) 
                 — 
               
               
                   
                 8-A 
                 19,625 
                 10 
                 0 (Healed) 
                 — 
               
               
                   
                 8-B 
                 19,625 
                 21 
                 0 (Healed) 
                 — 
               
               
                   
                 8-B 
                 19,625 
                 14 
                 1 
                 — 
               
               
                   
                 8-B 
                 19,625 
                 6 
                 0 (Healed) 
                 — 
               
               
                   
                 9-A 
                 19,625 
                 15 
                 0 (Healed) 
                 0 (Healed) 
               
               
                   
                 9-A 
                 19,625 
                 18 
                 3 
                 0 (Healed) 
               
               
                   
                 9-A 
                 19,625 
                 5 
                 0 (Healed) 
                 0 (Healed) 
               
               
                   
                 9-A 
                 19,625 
                 7 
                 0 (Healed) 
                 1 
               
               
                   
                 9-B 
                 19,625 
                 9 
                 0 (Healed) 
                 0 (Healed) 
               
               
                   
                 9-B 
                 19,625 
                 8 
                 1 
                 0 (Healed) 
               
               
                   
                 9-B 
                 19,625 
                 8 
                 1 
                 1 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
             
               
               
             
               
               
               
               
             
           
               
                 TABLE 5 
               
             
             
               
                   
               
               
                 Effect of 5-day topical application of ASI on the biochemical parameters 
               
             
          
           
               
                   
                 5th day 
               
             
          
           
               
                 ASI 
                 0 
                 4% (w/w) 
                 10% (w/w) 
               
               
                   
               
               
                 BUN, 
                 30.87 ± 1.24  
                 32.20 ± 1.11  
                 30.07 ± 2.69  
               
               
                 mg/dL 
               
               
                 CHOL, 
                 78.86 ± 5.11  
                 92.71 ± 7.75  
                 81.86 ± 7.66  
               
               
                 mg/dL 
               
               
                 HDL, 
                 11.29 ± 0.64  
                 9.86 ± 0.59 
                 14.57 ± 2.86  
               
               
                 mg/dL 
               
               
                 TRIG, 
                 115.71 ± 7.73  
                 152.71 ± 18.88  
                 112.00 ± 7.80  
               
               
                 mg/dL 
               
               
                 AST, 
                 288.14 ± 30.74  
                 319.00 ± 25.02  
                 309.86 ± 32.69  
               
               
                 μ/L 
               
               
                 GLU, 
                 110.14 ± 3.48  
                 98.29 ± 4.31  
                 105.57 ± 6.59  
               
               
                 mg/dL 
               
               
                 ALT, 
                 62.00 ± 4.73  
                 66.86 ± 5.86  
                 64.86 ± 7.85  
               
               
                 μ/L 
               
               
                 ALP, 
                 872.86 ± 141.13 
                 993.86 ± 264.85 
                 706.43 ± 113.24 
               
               
                 μ/L 
               
               
                 ALB, 
                 3.11 ± 0.12 
                 3.36 ± 0.19 
                 3.59 ± 0.20 
               
               
                 g/dL 
               
               
                 TBIL, 
                 0.31 ± 0.02 
                 0.47 ± 0.11 
                 0.35 ± 0.03 
               
               
                 mg/dL 
               
               
                 TP, 
                 6.10 ± 0.22 
                 6.20 ± 0.21 
                 6.49 ± 0.36 
               
               
                 g/dL 
               
               
                 AMY, 
                 725.57 ± 40.08  
                 780.86 ± 36.11  
                 831.43 ± 63.43  
               
               
                 μ/L 
               
               
                 LDL, 
                 44.43 ± 3.85  
                 52.14 ± 5.04  
                 45.14 ± 5.54  
               
               
                 mg/dl 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
             
               
               
             
               
               
               
               
             
           
               
                 TABLE 6 
               
             
             
               
                   
               
               
                 Effect of 10-day topical application of ASI on the biochemical parameters 
               
             
          
           
               
                   
                 10th day 
               
             
          
           
               
                 ASI 
                 0 
                 4% (w/w) 
                 10% (w/w) 
               
               
                   
               
               
                 BUN, 
                 28.57 ± 1.46  
                 30.36 ± 1.47  
                 29.83 ± 1.07  
               
               
                 mg/dL 
               
               
                 CHOL, 
                 93.86 ± 5.56  
                 93.57 ± 10.61 
                 94.29 ± 7.44  
               
               
                 mg/dL 
               
               
                 HDL, 
                 11.43 ± 0.78  
                 14.71 ± 1.39  
                 13.00 ± 1.29  
               
               
                 mg/dL 
               
               
                 TRIG, 
                 123.14 ± 7.90  
                 117.43 ± 15.86  
                 128.14 ± 7.40  
               
               
                 mg/dL 
               
               
                 AST, 
                 252.00 ± 37.38  
                 322.43 ± 33.44  
                 296.71 ± 52.43  
               
               
                 μ/L 
               
               
                 GLU, 
                 94.00 ± 2.07  
                 93.43 ± 3.37  
                 97.14 ± 5.19  
               
               
                 mg/dL 
               
               
                 ALT, 
                 72.14 ± 1.97  
                 77.86 ± 5.81  
                 78.29 ± 4.12  
               
               
                 μ/L 
               
               
                 ALP, 
                 626.57 ± 150.81 
                 801.71 ± 94.77  
                 772.57 ± 90.29  
               
               
                 μ/L 
               
               
                 ALB, 
                 3.54 ± 0.11 
                 3.53 ± 0.09 
                 3.59 ± 0.18 
               
               
                 g/dL 
               
               
                 TBIL, 
                 0.24 ± 0.02 
                 0.24 ± 0.02 
                 0.24 ± 0.02 
               
               
                 mg/dL 
               
               
                 TP, 
                 6.44 ± 0.13 
                 6.19 ± 0.14 
                 6.97 ± 0.41 
               
               
                 g/dL 
               
               
                 AMY, 
                 616.43 ± 83.32  
                 637.89 ± 113.25 
                 821.00 ± 48.22  
               
               
                 μ/L 
               
               
                 LDL, 
                 58.00 ± 4.49  
                 55.57 ± 8.12  
                 49.71 ± 6.22  
               
               
                 mg/dl 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
             
               
               
             
               
               
               
               
             
           
               
                 TABLE 7 
               
             
             
               
                   
               
               
                 Effect of 15-day topical application of ASI on the biochemical parameters 
               
             
          
           
               
                   
                 15th day 
               
             
          
           
               
                 ASI 
                 0 
                 4% (w/w) 
                 10% (w/w) 
               
               
                   
               
               
                 BUN, 
                 28.90 ± 1.81  
                 28.49 ± 3.65  
                 28.59 ± 2.58  
               
               
                 mg/dL 
               
               
                 CHOL, 
                 94.00 ± 6.39  
                 88.43 ± 11.84 
                 72.00 ± 5.16  
               
               
                 mg/dL 
               
               
                 HDL, 
                 11.14 ± 0.51  
                 16.14 ± 3.62  
                 10.29 ± 0.57  
               
               
                 mg/dL 
               
               
                 TRIG, 
                 175.14 ± 10.12  
                 159.29 ± 36.37  
                 135.71 ± 8.76  
               
               
                 mg/dL 
               
               
                 AST, 
                 191.00 ± 24.26  
                 177.86 ± 38.01  
                 163.57 ± 41.71  
               
               
                 μ/L 
               
               
                 GLU, 
                 119.57 ± 3.58  
                 125.14 ± 19.97  
                 110.14 ± 4.34  
               
               
                 mg/dL 
               
               
                 ALT, 
                 89.00 ± 4.49  
                 92.29 ± 15.12 
                 76.86 ± 5.76  
               
               
                 μ/L 
               
               
                 ALP, 
                 956.57 ± 126.03 
                 985.71 ± 184.98 
                 663.86 ± 89.42  
               
               
                 μ/L 
               
               
                 ALB, 
                 3.91 ± 0.08 
                 3.73 ± 0.41 
                 3.53 ± 0.15 
               
               
                 g/dL 
               
               
                 TBIL, 
                  0.2 ± 0.01 
                 0.19 ± 0.02 
                 0.17 ± 0.01 
               
               
                 mg/dL 
               
               
                 TP, 
                 7.13 ± 0.19 
                 11.96 ± 5.60  
                 6.34 ± 0.12 
               
               
                 g/dL 
               
               
                 AMY, 
                 1120.14 ± 37.40  
                 1038.71 ± 115.48  
                 1028.29 ± 70.23  
               
               
                 μ/L 
               
               
                 LDL, 
                 47.86 ± 5.03  
                 39.71 ± 4.97  
                 34.71 ± 4.98  
               
               
                 mg/dl 
               
               
                   
               
             
          
         
       
     
         [0087]    Results: As can be seen in  FIG. 1 , the wounds in the ASI treated animals healed quicker than the control. Surprisingly, the 4% (w/w) ASI cream worked better than the 10% (w/w) ASI cream. Hydroxyproline is a major component of protein collagen.  FIG. 2  shows that hydroxproline concentration in the treatment area increased in the ASI treated animals compared to control. Surprisingly, the 4% (w/w) ASI cream increased hydroxproline concentration to a greater extent that the 10% (w/w) ASI cream. Accordingly, topical application of ASI may increase hydroxproline levels in the treatment area. 
         [0088]    No mortality was observed during this study and all rats survived until the end of the study. The wounds in the control group displayed a greater degree of inflammation compared with ASI groups. The wound sites in ASI treated rats produced less exudate than the control wounds. 
         [0089]    Filling of the wound with granulation tissue to skin level was significantly slower in the control group than in the ASI groups ( FIG. 1  and Tables 1-3). The mean unhealed wound area was significantly smaller and the mean percentage of total wound healing was significantly higher in the ASI-treated wounds than in the control wound (P&lt;0.05). The wound area on completion of 15 days of treatment was 0.85 in the ASI 4% (w/w) treated group, when compared with 9.46, as observed in untreated rats (Table 2). The percent of wound contraction was also maximum in ASI 4% (w/w) treated rats (Table 3). The histopathology of the skin tissue corroborated with the macroscopic findings. In all rats, the healed wound surfaces were covered with a thick epithelial layer. Commonly mononuclear leukocyte type cellular infiltration was seen ( FIG. 17 ). 
         [0090]      FIG. 4  shows that inducible nitric oxide synthase levels in the treatment area decreased in the ASI treated animals compared to control. Accordingly, topical application of ASI may decrease inducible nitric oxide synthase levels.  FIG. 5  shows that endothelial nitric oxide synthase levels in the treatment area decreased in the ASI treated animals compared to control. Accordingly, topical application of ASI may decrease endothelial nitric oxide synthase levels. 
         [0091]      FIG. 6  shows that collagen levels in the treatment area increased in the ASI treated animals compared to control. Surprisingly, the 4% ASI (w/w) cream increased collagen levels to a greater extent that the 10% ASI (w/w) cream. Accordingly, topical application of ASI may hydroxproline levels. Accordingly, topical application of ASI may increase collagen levels in the treatment area. 
         [0092]      FIG. 7  shows that MMP-2 levels in the treatment area decreased in the ASI treated animals compared to control. Accordingly, topical application of ASI may decrease MMP-2 levels in the treatment area.  FIG. 8  shows that MMP-9 levels in the treatment area decreased in the ASI treated animals compared to control. Accordingly, topical application of ASI may decrease MMP-9 levels in the treatment area. 
         [0093]      FIG. 9  shows that VEGF levels in the treatment area increased in the ASI treated animals compared to control. Surprisingly, the 4% ASI (w/w) cream increased VEGF levels to a greater extent that the 10% ASI (w/w) cream. Accordingly, topical application of ASI may increase VEGF levels in the treatment area.  FIG. 10  shows that EGF levels in the treatment area increased in the ASI treated animals compared to control.  FIG. 13  shows that NFκB levels dcreased in the tratement area when compared to control. Accordingly, topical application of ASI may decrease NFκB levels in the treatment area.  FIGS. 14-15  shows that TGF-β and TNF-α levels dcreased in the tratement area when compared to control. Accordingly, topical application of ASI may decrease TGF-β and TNF-α levels in the treatment area.  FIG. 16  shows that IL-1β levels dcreased in the tratement area when compared to control. Accordingly, topical application of ASI may decrease IL-10β levels in the treatment area. 
         [0094]    It was also noted during qualitative examination that the hair on the shaved portion of the rat grew back at a faster rate in the ASI treatment groups than in the control group. 
         [0095]    As can be seen in  FIG. 2 , the topical application of ASI caused a significant increase in hydroxyproline content, with the maximum increase reported with 4% ASI (w/w) on day 10, which remained at the increased level on day 15; although in 10% ASI (w/w) group, there was a small drop in hydroxyproline content (P&lt;0.01). The results of hydroxyproline content corroborated with collagen measured in the wound area ( FIG. 6 ). 
         [0096]    Matrix metalloproteinases (MMP), both MMP-2 and MMP-9, registered a drop in ASI treated animals. MMP-2 and MMP -9 decreased most significantly on day 10 in rats treated with 10% ASI (w/w). Although on day 15, the level of MMP-2 was almost same at both (4% or 10%, (w/w)) dose levels of ASI ( FIGS. 7-8 ; P&lt;0.05). The results, which indicated tissue remodelling, were consistent with the histological findings ( FIG. 17 ). 
         [0097]    There was a significant decrease in iNOS at 10% ASI (w/w) on day 10. ASI 4% (w/w) also caused a reduction in iNOS expression, but to a less extent than ASI 10% (w/w) ( FIG. 4 ; P&lt; 0 . 001 ). On day 15, however, iNOS increased to some extent, but was still significantly lower han the iNOS level in ASI 4% (w/w) treated animals (P&lt;0.05). Overall, ASI caused a decrease in iNOS. The eNOS also showed a similar activity pattern ( FIG. 5 ). The expression of VEGF increased in all groups when compared with the control ( FIG. 9 ; P&lt; 0 . 05 ). The increase was maximum on day 5 in 4% ASI (w/w) treated rats. With time, the study reported a decrease in ASI on day 10 and 15. 
         [0098]    EGF and FGF were also measured in this study. There was an increase in EGF in all groups, with maximum increase being achieved in ASI treated animals on day 5. Subsequently, the level decreased on day 10 ( FIG. 10 ). FGF also showed a similar pattern of activity. It increased on day 5 maximally in ASI 4% (w/w) group, but decreased with time, that is, on day 10 and 15 ( FIG. 11 ; P&lt;0.05). The Western blot analysis of all the above mentioned proteins is shown in  FIG. 3 . 
         [0099]    As shown in  FIG. 13 , there was an increase in NF-κB expression in control group on day 5, which reduced at its own on day 10 and 15 (P&lt;0.001). ASI treated animals, on the other hand, registered a significant decrease in NF-κB on day 5, which further decreased on day 10 and 15, with maximum decrease observed on day 15 at 10% ASI (w/w) (P&lt;0.05). The TGF-β also showed a similar pattern of activity, with ASI causing a decrease in activity when compared with the control on respective days ( FIG. 13 , P&lt;0.05). The TNF-α was at its lowest in ASI 10% (w/w) treated animals on day 10 ( FIG. 14 , P&lt;0.05). The activity of TNF-α in ASI 10% (w/w) group, however, increased on day 15, but was significantly lower when compared with the control value on day 15. ASI treatment also caused a decrease in the level of IL-1β ( FIG. 16 , P&lt;0.05). The Western blot panel showing the expression level of various proteins is shown in  FIG. 12 . 
       Generic Procedure for Examples 2-7 
       [0100]    Animals: In this study, 42 male, 4 months old, Wistar albino rats weighing between 250 and 300 g will be used. Animals will be housed at 21° C. with a day/night cycle of 12 h. During the study these animals will be fed ad libitum standard rodent feed. Guidelines for the care and use of animals approved by the relevant institution were followed and the local ethics committee approved this study. 
         [0101]    Anesthesia: The rats will be anaesthetized with single intramuscular injection of 6 mg/kg xylazine hydrochloride (Rompun, Bayer, 23.32 mg/ml) and 85 mg/kg ketamine hydrochlorure (Ketalar, Parke-Davis, 50 mg/ml). 
         [0102]    Test drugs: 4% and 10% arginine-silicate inositol (ASI) (w/w) ointment and cold cream (Botafarma, 12.5% spermaceti+12% white wax+56% liquid paraffin+0.5% borate of soda+19% distilled water) will be used in this study. 
         [0103]    The rats will be divided into 2 main groups that each group will have randomly selected 21 rats. Each main group will be divided within themselves into 3 subgroups of 7 rats each to perform postoperative 5, 10 and 14 days of examination. Immediately after the wounds will be made (as described in Examples 2-7), the wounded areas in the first (n=21) and second (n=21) will be covered with cold cream (Botafarma, 12.5% spermaceti+12% white wax+56% liquid paraffin+0.5% borate of soda+19% distilled water) (Control) and 4% or 10% arginine-silicate inositol (ASI) (w/w) ointment twice a day, respectively. These applications will be repeated every day. The wounds will be clinically observed in all groups every day. The rats that completes postoperative follow-up periods of 5 (7 rats from each group), 10 (7 rats from each group) and 14 days (7 rats from each group) will be sacrificed. 
         [0104]    For Examples 2-6, injured skin tissue samples undergo histopathological examination. The samples will be collected after sacrificing the rats for histopathological examination purposes. These tissue samples will be fixed in 10% neutral buffered formalin solution, embedded in parafin wax, cut into 5 μm-thick sections and stained with haematoxyline-eosin and Masson&#39;s trichrome stain for examination by light microscopy. 
         [0105]    Histological scoring will be assigned in a blinded manner as described previously (Greenhalgh et al., 1990; Asai et al., 2013). Briefly, each specimen will be given a score of 1-12: 1-3, none to minimal cell accumulation and granulation tissue or epithelial migration; 4 to 6, thin, immature granulation dominated by inflammatory cells, but with few fibroblasts, capillaries or collagen deposition and minimal epithelial migration; 7-9, moderately thick granulation tissue, ranging from domination by inflammatory cells to more fibroblasts and collagen deposition; and 10-12, thick vascular granulation tissue dominated by fibroblasts and extensive collagen deposition. 
         [0106]    Statistical analyses: The thickness of granulation tissue will be examined and recorded at the center of each wound. Statistically, all data are expressed in millimeters as mean±standard error. The differences between days 5, 10 and 14 will be compared using the Mann Whitney U test. The differences between groups will be compared using the Kruskal-Wallis test. Statements of statistical significance are based on P&lt;0.05. These analyses will be carried out using SPSS statistical analysis system (Relase 10.0, SPSS. Inc). 
       Example 2 
       [0107]    Wound model: The dorsal surfaces of the rats will be shaved and prepared with 10% antiseptic povidone-iodine solution (Kim-Pa, Poviiodeks, % 10 povidone-iodine). Then a surgical cautery will be used to create a full-thickness burn wound. 
         [0108]    Measurements: On every bandage change, the wound boundaries will be traced on a transparent sheet with a fine-tipped permanent marker. These transparent sheets will be scanned and the areas of wound sites and epithelization fronts will be measured with the help of the paint-brush (PB) computer program. The data obtained will be calculated with the following formula: Area (cm 2 )=P/K.M, where P is the value of a particular wound site obtained on the PB, K is a rate constant for expressing the values of the PB as cm 2 , and M is the magnitude of tracings after scanning. During the qualitative examination, the expansion rate, the day wound contraction started, the fraction of the wound healed with contraction, the rate of wound contraction, the day epithelization will be first noticed, the fraction of wound healed with epithelization (final wound size) and the number of days in which wound healing will be fully completed will be evaluated. The statistical comparison will be made with ANOVA. The results will be considered significant at P&lt;0.05 and a 95% confidence interval (Kiliç et al., 2002). 
       Example 3 
       [0109]    Wound model: The dorsal surfaces of the rats will be shaved and prepared with 10% antiseptic povidone-iodine solution (Kim-Pa, Poviiodeks, % 10 povidone-iodine). Then the rats will be given a 5 cm by 5 cm abrasion wound. 
         [0110]    Measurements: On every bandage change, the wound boundaries will be traced on a transparent sheet with a fine-tipped permanent marker. These transparent sheets will be scanned and the areas of wound sites and epithelization fronts will be measured with the help of the paint-brush (PB) computer program. The data obtained will be calculated with the following formula: Area (cm 2 )=P/K.M, where P is the value of a particular wound site obtained on the PB, K is a rate constant for expressing the values of the PB as cm 2 , and M is the magnitude of tracings after scanning. During the qualitative examination, the expansion rate, the day wound contraction started, the fraction of the wound healed with contraction, the rate of wound contraction, the day epithelization will be first noticed, the fraction of wound healed with epithelization (final wound size) and the number of days in which wound healing will be fully completed will be evaluated. The statistical comparison will be made with ANOVA. The results will be considered significant at P&lt;0.05 and a 95% confidence interval (Kiliç et al., 2002). 
       Example 4 
       [0111]    Wound model: The dorsal surfaces of the rats will be shaved and prepared with 10% antiseptic povidone-iodine solution (Kim-Pa, Poviiodeks, % 10 povidone-iodine). Then a surgical trochar will be used to create three distinct 2 cm deep puncture wounds. 
         [0112]    Measurements: On every bandage change, the wound boundaries will be traced on a transparent sheet with a fine-tipped permanent marker. These transparent sheets will be scanned and the areas of wound sites and epithelization fronts will be measured with the help of the paint-brush (PB) computer program. The data obtained will be calculated with the following formula: Area (cm 2 )=P/K.M, where P is the value of a particular wound site obtained on the PB, K is a rate constant for expressing the values of the PB as cm 2 , and M is the magnitude of tracings after scanning. During the qualitative examination, the expansion rate, the day wound contraction started, the fraction of the wound healed with contraction, the rate of wound contraction, the day epithelization will be first noticed, the fraction of wound healed with epithelization (final wound size) and the number of days in which wound healing will be fully completed will be evaluated. The statistical comparison will be made with ANOVA. The results will be considered significant at P&lt;0.05 and a 95% confidence interval (Kiliç et al., 2002). 
       Example 5 
       [0113]    Wound model: The dorsal surfaces of the rats will be shaved and prepared with 10% antiseptic povidone-iodine solution (Kim-Pa, Poviiodeks, % 10 povidone-iodine). Then the rats will be given a 5 cm by 5 cm abrasion wound. The ASI treatment in this example is provided as the 4% or 10% ASI (w/w) formulations described above, but embedded into the bandage itself 
         [0114]    Measurements: On every bandage change, the wound boundaries will be traced on a transparent sheet with a fine-tipped permanent marker. These transparent sheets will be scanned and the areas of wound sites and epithelization fronts will be measured with the help of the paint-brush (PB) computer program. The data obtained will be calculated with the following formula: Area (cm 2 )=P/K.M, where P is the value of a particular wound site obtained on the PB, K is a rate constant for expressing the values of the PB as cm 2 , and M is the magnitude of tracings after scanning. During the qualitative examination, the expansion rate, the day wound contraction started, the fraction of the wound healed with contraction, the rate of wound contraction, the day epithelization will be first noticed, the fraction of wound healed with epithelization (final wound size) and the number of days in which wound healing will be fully completed will be evaluated. The statistical comparison will be made with ANOVA. The results will be considered significant at P&lt;0.05 and a 95% confidence interval (Kiliç et al., 2002). 
       Example 6 
       [0115]    Wound model: The dorsal surfaces of the rats will be shaved and prepared with 10% antiseptic povidone-iodine solution (Kim-Pa, Poviiodeks, % 10 povidone-iodine). Then a portacath (infusing normal saline) will be installed. 
         [0116]    Measurements: On every bandage change, the wound boundaries will be traced on a transparent sheet with a fine-tipped permanent marker. These transparent sheets will be scanned and the areas of wound sites and epithelization fronts will be measured with the help of the paint-brush (PB) computer program. The data obtained will be calculated with the following formula: Area (cm 2 )=P/K.M, where P is the value of a particular wound site obtained on the PB, K is a rate constant for expressing the values of the PB as cm 2 , and M is the magnitude of tracings after scanning. During the qualitative examination, the expansion rate, the day wound contraction started, the fraction of the wound healed with contraction, the rate of wound contraction, the day epithelization will be first noticed, the fraction of wound healed with epithelization (final wound size) and the number of days in which wound healing will be fully completed will be evaluated. The statistical comparison will be made with ANOVA. The results will be considered significant at P&lt;0.05 and a 95% confidence interval (Kiliç et al., 2002). 
       Example 7 
       [0117]    Wound model: The dorsal surfaces of the rats will be shaved. 
         [0118]    Measurements: Twice each day, the shaved boundaries will be traced on a transparent sheet with a fine-tipped permanent marker. These transparent sheets will be scanned and the areas of hair growth fronts will be measured with the help of the paint-brush (PB) computer program. The data obtained will be calculated with the following formula: Area (cm 2 )=P/K.M, where P is the value of a particular site obtained on the PB, K is a rate constant for expressing the values of the PB as cm 2 , and M is the magnitude of tracings after scanning. During the qualitative examination, the expansion rate of hair and general appearance of the regrown hair will be noted. Also, twice each day, the length of the regrown hair will be measured. The statistical comparison will be made with ANOVA. The results will be considered significant at P&lt;0.05 and a 95% confidence interval (Kilic et al., 2002). 
         [0119]    Statistical analyses: The length of regrown hair will be examined and recorded at the center of each shaved area. Statistically, all data are expressed in milimeters as mean±standard error. The differences between days 2, 7 and 14 will be compared using the Mann Whitney U test. The differences between groups will be compared using the Kruskal-Wallis test. Statements of statistical significance are based on P&lt;0.05. These analyses will be carried out using SPSS statistical analysis system (Relase 10.0, SPSS. Inc). 
         [0120]    As used in the claims below and throughout this disclosure, by the phrase “consisting essentially of” is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase “consisting essentially of” indicates that the listed elements are required or mandatory, but that other elements are optional and can or cannot be present depending upon whether or not they affect the activity or action of the listed elements. For example, the use of a composition “consisting essentially of ASI” for the treatment of wounds or for promoting hair growth would not include other active ingredients known to promote wound healing or known to improve hair growth, respectively. 
         [0121]    Although the invention has been described with reference to embodiments and examples, it should be understood that numerous and various modifications can be made without departing from the spirit of the invention. Accordingly, the invention is limited only by the following claims.