Curcumin differentially regulates TGF-beta1, its receptors and nitric oxide synthase during impaired wound healing

Wound healing is a highly ordered process, requiring complex and coordinated interactions involving peptide growth factors of which transforming growth factor-beta (TGF-beta) is one of the most important. Nitric oxide is also an important factor in healing and its production is regulated by inducible nitric oxide synthase (iNOS). We have earlier shown that curcumin (diferuloylmethane), a natural product obtained from the plant Curcuma longa, enhances cutaneous wound healing in normal and diabetic rats. In this study, we have investigated the effect of curcumin treatment by topical application in dexamethasone-impaired cutaneous healing in a full thickness punch wound model in rats. We assessed healing in terms of histology, morphometry, and collagenization on the fourth and seventh days post-wounding and analyzed the regulation of TGF-beta1, its receptors type I (tIrc) and type II (tIIrc) and iNOS. Curcumin significantly accelerated healing of wounds with or without dexamethasone treatment as revealed by a reduction in the wound width and gap length compared to controls. Curcumin treatment resulted in the enhanced expression of TGF-beta1 and TGF-beta tIIrc in both normal and impaired healing wounds as revealed by immunohistochemistry. Macrophages in the wound bed showed an enhanced expression of TGF-beta1 mRNA in curcumin treated wounds as evidenced by in situ hybridization. However, enhanced expression of TGF-beta tIrc by curcumin treatment observed only in dexamethasone-impaired wounds at the 7th day post-wounding. iNOS levels were increased following curcumin treatment in unimpaired wounds, but not so in the dexamethasone-impaired wounds. The study indicates an enhancement in dexamethasone impaired wound repair by topical curcumin and its differential regulatory effect on TGF-beta1, it's receptors and iNOS in this cutaneous wound-healing model.     

Efficacy of Butea monosperma on dermal wound healing in rats

Wound healing occurs as a fundamental response to tissue injury. Several natural products have been shown to accelerate the healing process. The present investigation was undertaken to determine the efficacy of topical administration of an alcoholic bark extract of Butea monosperma (B. monosperma) on cutaneous wound healing in rats. Full-thickness excision wounds were made on the back of rat and B. monosperma extract was administered topically. The granulation tissue formed on days 4, 8, 12 and 16 (post-wound) was used to estimate total collagen, hexosamine, protein, DNA and uronic acid. The extract increased cellular proliferation and collagen synthesis at the wound site, as evidenced by increase in DNA, total protein and total collagen content of granulation tissues. The extract treated wounds were found to heal much faster as indicated by improved rates of epithelialization and wound contraction, also confirmed by histopathological examinations. Also, the tensile strength of drug-treated wounds was increased significantly. In addition, we show that B. monosperma possesses antioxidant properties, by its ability to reduce lipid peroxidation. The results clearly substantiate the beneficial effects of the topical application of B. monosperma in the acceleration of wound healing.     

The effects of topical treatment with curcumin on burn wound healing in rats

The present study was designed to determine the role of topical treatment with curcumin (Cur) on burn wound healing in rats. The Wistar-albino rats were randomly allotted into one of three experimental groups: 4th, 8th and 12th day (post burn) and all groups include subgroups which Burn and Burn + Cur. Each group contains 12 animals. Burn wounds were made on the back of rat and Cur was administered topically. At the end of the study, all animals were sacrificed and the wound tissues removed for analyse to biochemical and histopathological changes. There was a significant increase in the hydroxyproline levels in the skin of the Cur groups. Cur treated wounds were found to heal much faster as indicated by improved rates of inflammatory cells, collagen deposition, angiogenesis, granulation tissue formation and epithelialization which were also confirmed by histopathological and biochemical examinations. Our data also indicate that there is a rise in the expression of proliferating cell nuclear antigen in skin tissues of Cur-treated rats in the Burn group. The results clearly substantiate the beneficial effects of the topical application of Cur in the acceleration of wound healing.     

Curcumin as a wound healing agent

Turmeric (Curcuma longa) is a popular Indian spice that has been used for centuries in herbal medicines for the treatment of a variety of ailments such as rheumatism, diabetic ulcers, anorexia, cough and sinusitis. Curcumin (diferuloylmethane) is the main curcuminoid present in turmeric and responsible for its yellow color. Curcumin has been shown to possess significant anti-inflammatory, anti-oxidant, anti-carcinogenic, anti-mutagenic, anti-coagulant and anti-infective effects. Curcumin has also been shown to have significant wound healing properties. It acts on various stages of the natural wound healing process to hasten healing. This review summarizes and discusses recently published papers on the effects of curcumin on skin wound healing. The highlighted studies in the review provide evidence of the ability of curcumin to reduce the body's natural response to cutaneous wounds such as inflammation and oxidation. The recent literature on the wound healing properties of curcumin also provides evidence for its ability to enhance granulation tissue formation, collagen deposition, tissue remodeling and wound contraction. It has become evident that optimizing the topical application of curcumin through altering its formulation is essential to ensure the maximum therapeutical effects of curcumin on skin wounds.     

Effect of nifedipine and amlodipine on dead space wound healing in rats

Effect of two calcium channel blockers (CCBs) nifedipine and amlodipine, was studied on normal and steroid depressed wound healing in albino rats, using the dead space wound model. The drugs enhanced normal healing as evidenced by increase in tensile strength of 10 days old granulation tissue. There was neither a significant change in the hydroxyproline level (or collagen) nor a change in the glycosaminoglycan content in granulation tissue. However, lysyloxidase level was increased significantly. The increase in tensile strength could thus be attributed to better cross-linking and maturation of collagen rather than collagen synthesis per se. The drugs were also able to overcome steroid depressed wound healing. It is likely that the prohealing effects may be related to the improved antioxidant status too, since superoxide dismutase levels were observed to be higher in the CCB- treated animals.     

Effect of sodium diphenylhydantoin on skin wound healing in rats

This study evaluated the effect of phenytoin (sodium diphenylhydantoin) on skin wound healing in a rat model. The study was divided into two parts. In part I, 20 mul of phenytoin (10 mg/ml) was subcutaneously injected into the 3-cm dorsal full-thickness incisional wounds of 14 rats on postoperative days 0, 3, and 6. Twelve rats that received saline injections were used as the controls. The skin samples were harvested and tested for tensile strength and histology. An additional 12 rats with the same incisional wounds were tested for chemokine gene expressions. In part II, 20 mul of phenytoin (10 mg/ml) was applied topically once a day on a 4 x 4 cm area of the open dorsal wounds of 10 rats. Saline was applied to the wounds of the 10 control group rats. The wounds were measured weekly. The results showed that the average tensile strength of the phenytoin-treated wound was 0.49 +/- 0.08 MPa compared with the control group at 0.02 +/- 0.01 MPa (p < 0.05). The density ratio of chemokine monocyte chemotactic protein (MCP-1) to beta-actin in the phenytoin-treated group was also significantly higher than in the control group (p < 0.05). Histologic analysis of the phenytoin group showed a large amount of fibroblast proliferation, collagen synthesis, and neovascularization. Phenytoin-treated wounds were also smaller at 1 to 6 weeks postoperatively than the control group wounds. The authors conclude that the administration of phenytoin can promote wound healing and significantly increase MCP-1 expression. Phenytoin-treated wounds showed significant increase in collagen deposition and neovascularization, which resulted in an increased wound tensile strength and accelerated healing of both open and closed wounds.     

The diabetic rat as an impaired wound healing model: stimulatory effects of transforming growth factor-beta and basic fibroblast growth factor

Two models of wound repair compared the effect of defined, recombinant growth factors on the rate of wound repair in both normal and streptozotocin-induced diabetic rats: subcutaneous implantation of polyvinyl alcohol sponges and incisional wounding. Transverse incisional wounds were made on the dorsal surface of rats and closed with steel sutures. Three days postwounding the rats received a single injection of either transforming growth factor-beta or vehicle alone directly into the wound site. Animals were sacrificed 7, 14, and 21 days postwounding, and fresh and formalin-fixed wound tensile strength were measured. Diabetic rats had expected defects in wound repair, including decreased granulation tissue and reduced amounts of collagen, protein, and DNA. Fresh tensile strength of the diabetic incisions was 53% of normal on Day 7 (p < or = .01) and 29% of normal on Day 21. Fixed tensile strength was 41% of normal on Day 7 (p < or = .01) and fell to 78% of normal by Day 21 (p < or = .01), suggesting that collagen concentrations of diabetic wounds increased towards normal but did not undergo maturation. TGF beta produced a moderate increase in tensile strength of fresh and fixed wounds of diabetic rats, but not to the levels of wounds in untreated normal rats. Sponges fill with granulation tissue, their reproducible rate of organization being measured by histological and biochemical methods. A single injection into sponges 3 days postimplantation of basic fibroblast growth factor, transforming growth factor-beta, or vehicle only, was evaluated at 7 and 9 days postimplantation. In the sponge model, bFGF and TGF beta were each able to induce significant increases in the accumulation of granulation tissue in both diabetic and normal rats. TGF beta increased the collagen content of sponges by 136% in sponges from diabetic animals (p < or = .001), thereby raising the collagen content to that of normal control wounds, while stimulating a 49% (p < or = .02) increase in sponges from normal animals on Day 9. By contrast, the response to bFGF was predominantly an increase in the protein and DNA content of the sponges. These results emphasize the differential effects of the two cytokines in accelerating healing under conditions of defective wound repair.     

The effect of nerve growth factor on the early responses during the process of wound healing

In this study, we investigated the role of nerve growth factor (NGF)-incorporated collagen on wound healing in rats. Full-thickness excision wounds were made on the back of female rats weighing about 150-160 g. Topical application of NGF-incorporated collagen, at a concentration of 1 microg/1.2 mg collagen/cm(2), once a day, for 10 days resulted in complete healing of wounds on the 15th day. The concentrations of collagen, hexosamine and uronic acid in the granulation tissue were determined. The NGF-incorporated collagen-treated rats required shorter duration for the healing with an increased rate of wound contraction. Histological and electron microscopical evaluations were also performed, which reveal the activation of fibroblasts and endoplasmic reticulum and therefore increased level of collagen synthesis due to NGF application. These results clearly indicate that the topical application of NGF-incorporated collagen enhanced the rate of healing of excision wounds.     

The effect of finger millet feeding on the early responses during the process of wound healing in diabetic rats

In the present study, the role of finger millet feeding on skin antioxidant status, nerve growth factor (NGF) production and wound healing parameters in healing impaired early diabetic rats is reported. Hyperglycemic rats received food containing 50 g/100 g finger millet (FM). Non-diabetic controls and diabetic controls received balanced nutritive diet. Full-thickness excision skin wounds were made after 2 weeks prior feeding of finger millet diet. The rate of wound contraction, and the levels of collagen, hexosamine and uronic acid in the granulation tissue were determined. The skin antioxidant status and lipid peroxide concentration were also monitored during the study. In hyperglycemic rats fed with finger millet diet, the healing process was hastened with an increased rate of wound contraction. Skin levels of glutathione (GSH), ascorbic acid and alpha-tocopherol in alloxan-induced diabetic rat were lower as compared to non-diabetics. Altered activities of superoxide dismutase (SOD) and catalase (CAT) were also recorded in diabetics. Interestingly, thiobarbituric acid reactive substances (TBARS) were elevated in the wound tissues of all the groups, when compared to normal (unwounded) skin tissues. However, in diabetic rats the TBARS levels of both normal and wounded skin tissues were significantly elevated (P < 0.001) when compared with control (non-diabetic) and diabetics fed with FM. Impaired production of NGF, determined by ELISA, in diabetic rats was improved upon FM feeding and further confirmed by immunocytochemical observations reflects the increased expression of NGF in hyperglycemic rats supplemented with FM-enriched diet. Histological and electron microscopical evaluations revealed the epithelialization, increased synthesis of collagen, activation of fibroblasts and mast cells in FM-fed animals. Thus, increased levels of oxidative stress markers accompanied by decreased levels of antioxidants play a vital role in delaying wound healing in diabetic rats. However, FM feeding to the diabetic animals, for 4 weeks, controlled the glucose levels and improved the antioxidant status, which hastened the dermal wound healing process.     

The tetrachlorodecaoxygen complex reverses the effect of cortisone on wound healing

Immunosuppression induced by the administration of glucocorticoids will prevent normal wound contraction and normal increases in tensile strength. Vitamin A, anabolic steroids, and growth hormone will, in the presence of glucocorticoids, restore mesenchymal cell proliferation, the accumulation of collagen, and the rate of increase of wound tensile strength. They will not, however, antagonize the inhibition of wound contraction. A novel inorganic agent, the tetrachlorodecaoxygen anion complex (TCDO), known to enhance the migration and activation of macrophages, was tested in a rat model of impaired wound healing using high doses of glucocorticoids. Histology, changes in wound contraction, collagen synthesis, and tensile strength were evaluated. Animals receiving cortisone in combination with TCDO displayed markedly enhanced wound healing, including restoration of tensile strength, collagen synthesis, and wound contraction. The results indicate that TCDO could be a potential agent of wound healing in immunosuppressed patients and anergic wounds.     

Nitric oxide inhibits wound collagen synthesis

Nitric oxide (NO) is a messenger molecule which regulates many physiological functions like immunity, vascular tone and serves as a neurotransmitter. Although it is known to participate in healing process, its role in collagen synthesis is not clear. Therefore, the present investigation was done to study the role of NO in wound collagen synthesis. Rats received full thickness, circular (8 mm), transdermal wounds which were treated with NO releaser, sodium nitroprusside (SNP, 0.001 100 M) topically for 5 days. Wound collagen content estimated in terms of hydroxyproline (HP) and confirmed histochemically was decreased significantly by all SNP doses. L-Arginine, a substrate for nitric oxide synthase (NOS) when applied topically decreased collagen content of the wounded tissues. N-Nitro-L-arginine methyl ester (L-NAME), a competitive inhibitor of NOS, increased wound collagen content significantly as compared to untreated and SNP treated animal wounds when administered intraperitoneally at the doses 3, 10 and 30 mg/kg. Furthermore, histological findings also demonstrated laying down of thick collagen bundles and proliferation of fibroblasts together with prominent angiogenesis in L-NAME treated wound tissues as compared to untreated and SNP treated tissues. N-nitro-D-arginine methyl ester, an inactive isomer, was found to have no effect on wound collagen levels. When L-arginine was administered in L-NAME pretreated rats, it significantly elevated wound HP content. The results indicate that NO plays an important role in regulating the collagen biosynthesis in skin model of a healing wound.     

The effect of disodium cromoglycate on the skin wound healing and collagen content in the wounds of rats

The effect of disodium cromoglycate on skin wound healing and collagen formation in the wounds was studied. Disodium cromoglycate (a mast cell stabilizer) administered to the rats in a dose of 2 mg/animal was found to retard wound healing and markedly increased wound surface in all examined days (3rd, 5th, 7th, 10th, 14th day of healing). The mast cell stabilizer injected directly into wounds decreased collagen content, especially on 10th and 14th day of the healing process.     

Efficacy of l-proline administration on the early responses during cutaneous wound healing in rats

Proline (Pro) plays a versatile role in cell metabolism and physiology. Pro and hydroxypro are major imino acids present in collagen, an important connective tissue protein, essential for wound healing, which is a primary response to tissue injury. This study explains the role of l-pro on cutaneous wound healing in rats when administered both topically and orally. Open excision wounds were made on the back of rats, and 200 μl (200 mg) of pro was administered topically and orally once daily to the experimental rats until the wounds healed completely. The control wounds were left untreated. Granulation tissues formed were removed after day 4 and 8 of post excision wounding, and biochemical parameters such as total protein, collagen, hexosamine, and uronic acid were estimated. Levels of enzymatic and non-enzymatic antioxidants such as catalase, superoxide dismutase, glutathione peroxidase, ascorbic acid, and reduced glutathione were evaluated along with lipid peroxides in the granulation tissues. Tensile strength and period of epithelialization were also measured. It was observed that the treated wounds healed very fast as evidenced by augmented rates of epithelialization and wound contraction, which was also confirmed by histological examinations. The results strappingly authenticate the beneficial effects of the topical administration of l-proline in the acceleration of wound healing than the oral administration and control.     

Effect of Ocimum sanctum Linn. on normal and dexamethasone suppressed wound healing

Ethanolic extract of leaves of O. sanctum was investigated for normal wound healing and dexamethasone depressed healing using incision, excision and dead space wound models in albino rats. The extract of O. sanctum significantly increased the wound breaking strength in incision wound model. The extract treated wounds were found to epithelialize faster and the rate of wound contraction was significantly increased as compared to control wounds. Significant increase in wet and dry granulation tissue weight, granulation tissue breaking strength and hydroxyproline content in dead space wound model was observed. The extract significantly decreased the antihealing activities of dexamethasone in all the wound models. The results indicated that the leaf extract promotes wound healing significantly and able to overcome the wound healing suppressing action of dexamethasone. Histological examination of granulation tissue to determine the pattern of lay-down for collagen confirmed the results.     

Effect of curcumin on protein glycosylation, lipid peroxidation, and oxygen radical generation in human red blood cells exposed to high glucose levels

Curcumin (diferuloylmethane) is the most active component of turmeric. It is believed that curcumin is a potent antioxidant and anti-inflammatory agent. Experimental studies with diabetic animals demonstrate that curcumin supplementation can suppress cataract development and collagen cross-linking, promote wound healing, and lower blood lipids and glucose levels. The mechanism by which curcumin may cause diabetes-associated vascular damage to regress is not known. Erythrocytes were treated with high levels of glucose (mimicking diabetes) in the presence or absence of curcumin (0-10 muM) in the medium at 37 degrees C for 24 h. This study demonstrates that curcumin prevents protein glycosylation and lipid peroxidation caused by high glucose levels using an erythrocyte cell model. This study also suggests that curcumin may inhibit oxygen radical production caused by high glucose concentrations in a cell-free system, and increase glucose utilization in erythrocytes. This provides evidence for a novel mechanism by which curcumin supplementation may prevent the cellular dysfunction associated with diabetes.     

An investigation on skin wound healing in mice with a taurine-chitosan gel formulation

Summary. The process of wound healing begins immediately following surface lesions or just after exposure to radiation, chemical agents or extreme temperatures. Taurine (2-aminoethane sulfonic acid), an amino acid containing sulfur, is found in almost all tissues in mammals, playing various important physio-logical roles in each organ. Taurine exhibits an antioxidant effect and is also known to have effects on cell proliferation, inflammation and collagenogenesis. Many antioxidants have been used to eliminate the negative effects of oxygen free radicals on wound healing. The objective of the present study was to examine the wound healing effect in mice of taurine-chitosan gel, which releases taurine slowly over a long time period. Fifty mM of taurine in 1.5% chitosan polymer (TAU-GEL) and 1.5% chitosan polymer (CHI-GEL) were applied to full thickness skin wounds of mice once a day for seven days. After seven days of treatment, lipid peroxide formation-malondialdehyde (MDA) and hydroxyproline (HPX) levels and the tensile strength of wound tissues were measured. All results were compared with those of the untreated control group (CONT). The structural alterations in the skin layers were also histologically investigated. It was found that locally administered TAU-GEL form significantly increased wound tensile strength by decreasing the MDA and increasing HPX levels. These results were supported by histological findings. All observations suggest that taurine gel may be effective in wound healing. Received January 15, 2001 Accepted June 4, 2001     

pGlcNAc Nanofiber Treatment of Cutaneous Wounds Stimulate Increased Tensile Strength and Reduced Scarring via Activation of Akt1

Treatment of cutaneous wounds with poly-N-acetyl-glucosamine containing nanofibers (pGlcNAc), a novel polysaccharide material derived from a marine diatom, results in increased wound closure, antibacterial activities and innate immune responses. We have shown that Akt1 plays a central role in the regulation of these activities. Here, we show that pGlcNAc treatment of cutaneous wounds results in a smaller scar that has increased tensile strength and elasticity. pGlcNAc treated wounds exhibit decreased collagen content, increased collagen organization and decreased myofibroblast content. A fibrin gel assay was used to assess the regulation of fibroblast alignment in vitro. In this assay, fibrin lattice is formed with two pins that provide focal points upon which the gel can exert force as the cells align from pole to pole. pGlcNAc stimulation of embedded fibroblasts results in cellular alignment as compared to untreated controls, by a process that is Akt1 dependent. We show that Akt1 is required in vivo for the pGlcNAc-induced increased tensile strength and elasticity. Taken together, our findings suggest that pGlcNAc nanofibers stimulate an Akt1 dependent pathway that results in the proper alignment of fibroblasts, decreased scarring, and increased tensile strength during cutaneous wound healing.     

Curcumin treatment enhances the repair and regeneration of wounds in mice exposed to hemibody gamma-irradiation

Hemibody irradiation in multiple fractionated doses is frequently used for the treatment of various neoplastic disorders. It produces both acute and late effects on the skin and subcutaneous tissues that have profound implications in the healing of surgical wounds. Because of the crucial practical importance of hemibody radiation exposure associated with skin wounds, it is imperative to investigate the efficacy of cost-effective herbal products in the reconstruction of irradiated wounds. Therefore, the effect of pretreatment of curcumin was studied on the healing of excision wound in mice exposed to 2, 4, 6, or 8 Gy of hemibody gamma-radiation. A full-thickness skin wound was created by removing the skin flap of the dorsum of 8- to 10-week-old Swiss albino mice partially (lower half, below the rib cage) exposed to 2, 4, 6, or 8 Gy of gamma-radiation. The progression of wound contraction was monitored periodically by capturing video images of the wound, where the first image of each wound from different groups was obtained 1 day after wounding and that day was considered as day 0. Eight animals were used in each group at each exposure dose for wound contraction studies. Furthermore, the effect of curcumin on mean healing time after exposure of mice to 2, 4, 6, or 8Gy of hemibody gamma-radiation was also evaluated, where eight animals were used in each group at each exposure dose. Collagen, hexosamine, DNA, nitric oxide, and histologic profiles were also evaluated during the course of healing of excision wounds at days 4, 8, and 12 after irradiation treated or not with curcumin before exposure to 0 or 6 Gy of gamma-radiation. Six animals were used in each group at each interval for each biochemical parameter studied, except for histologic evaluations, where four animals were used in each group at each interval. Exposure of mice to different doses of gamma-radiation resulted in a dose-dependent delay in contraction and wound-healing time of excision wound, whereas curcumin pretreatment caused a significant elevation in the rate of wound contraction and a decrease in the mean wound-healing time. Treatment with curcumin before irradiation enhanced the synthesis of collagen, hexosamine, DNA, nitrite, and nitrate, and histologic assessment of wound biopsy specimens revealed improved collagen deposition and an increase in fibroblast and vascular densities. The authors' study demonstrates that curcumin pretreatment has a conducive effect on the irradiated wound and could be a substantial therapeutic strategy for ameliorating radiation-induced delay in wound repair in cases of radiation-induced skin injuries.     

Application of platelet derived growth factor-BB and diabetic wound healing: the relationship with oxidative events

The reasons that cause delay in wound healing in diabetes are a decrease in the level of growth factors secretion, an increase in the destruction of growth factors and in oxidative stress. Platelet derived growth factor (PDGF) is one of the important growth factors that play a role in all phases of wound healing. This study investigates time-dependent effects of topically PDGF-BB administration on oxidative events on the healing of dorsolateral-excisional wounds in diabetic rats. Forty-two female Wistar-albino rats with streptozotocin-induced diabetes were divided into four groups: control group, untreated group, chitosan-treated group, chitosan?+?PDGF-BB-treated group. Two identical full-thickness excisional skin wounds were made under anaesthesia in all rats except for the control group. In the PDGF-BB-treated and chitosan-treated groups, the wounds were treated topically PDGF-BB (7?ng/mL, single daily dose) and blank chitosan gel (equal amount) after wounding, respectively. After these administrations, on day 3 and day 7 of wound healing, rats were sacrificed. Thiobarbituric acid reactive substances, glutathione, nitric oxide, ascorbic acid levels, and superoxide dismutase activity in wound tissues were spectrophotometrically measured. PDGF-BB administration significantly increased TBARS levels and non-enzymatic antioxidant levels in early phase of diabetic wound healing. PDGF-BB dramatically reduced NO_x levels on day 3 and sharply increased NO_x levels on day 7 of wound healing. Consequently, PDGF-BB administration can be effective on oxidative balance in the early phase of diabetic wound healing.     

Wound healing activity of ethanolic extract of Shorea robusta Gaertn. f. resin

The ethanolic extract of S. robusta resin (10 and 30 % w/w applied locally in excised and incised wounds) produced a dose-dependent acceleration in wound contraction and increased hydroxyproline content and tensile strength of wounds in rats. The results demonstrate wound healing activity of ethanolic extract of S. robusta resin.