Demonstration of the Rat Ischemic Skin Wound Model

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and unrelieved pressure due to immobility. Animal models have been developed that attempt to mimic these conditions for the purpose of furthering our understanding of the complexity of chronic wounds. The model described herein is a rat ischemic skin flap model that permits a prolonged reduction of blood flow resulting in wounds that become ischemic and resemble a chronic wound phenotype (reduced vascularization, increased inflammation and delayed wound closure). It consists of a bipedicled dorsal flap with 2 ischemic wounds placed centrally and 2 non-ischemic wounds lateral to the flap as controls. A novel addition to this ischemic skin flap model is the placement of a silicone sheet beneath the flap that functions as a barrier and a splint to prevent revascularization and reduce contraction as the wounds heal. Despite the debate of using rats for wound healing studies due to their quite distinct anatomic and physiologic differences compared to humans (i.e., the presence of a panniculus carnosus muscle, short life-span, increased number of hair follicles, and their ability to heal infected wounds) the modifications employed in this model make it a valuable alternative to previously developed ischemic skin flap models.     

The cardiac neural stem cell phenotype is compromised in streptozotocin‐induced diabetic cardiomyopathy

Neural stem cells were identified in the rat heart and during scar formation and healing participated in sympathetic fiber sprouting and angiogenesis. In the setting of diabetes, impaired wound healing represents a typical pathological feature. These findings provided the impetus to test the hypothesis that experimental diabetes adversely influenced the phenotype of cardiac neural stem cells. Streptozotocin (STZ)‐induced diabetic rats were associated with elevated plasma glucose levels, significant loss of body weight and left ventricular contractile dysfunction. In the heart of STZ‐diabetic rats, the density of nestin immunoreactive processes emanating from cardiac neural stem cells were reduced. The latter finding was reaffirmed as nestin protein expression was significantly decreased in the heart of STZ‐diabetic rats and associated with a concomitant reduction of nestin mRNA. Employing the TUNEL assay, the loss of nestin expression in STZ‐diabetic rats was not attributed to widespread cardiac neural stem cell apoptosis. Insulin administration to STZ‐diabetic rats with established hyperglycaemia led to a modest recovery of nestin protein expression in cardiac neural stem cells. By contrast, the administration of insulin immediately after STZ injection improved plasma glucose levels and significantly attenuated the loss of nestin protein expression. These data highlight the novel observation that nestin protein expression in cardiac neural stem cells was significantly reduced in STZ‐induced type I diabetic rats. The aberrant cardiac neural stem cell phenotype may compromise their biological role and predispose the diabetic heart to maladaptive healing following ischemic injury. J. Cell. Physiol. 220: 440–449, 2009. © 2009 Wiley‐Liss, Inc.     

中医病证相符的糖尿病慢性皮肤溃疡大鼠造模方法初探

目的观察不同造模方法致糖尿病大鼠慢性皮肤溃疡创面形态及愈合时间的影响。方法 50只SD大鼠随机分为5组:皮肤缺损组(QS组:剪皮),糖尿病组(DM组:STZ+剪皮),糖尿病加金黄色葡萄球菌组(DMJJ组:STZ+剪皮+金葡菌),糖尿病加激素组(DMJS组:STZ+剪皮+激素注射),糖尿病加激素加异物组(DMYW组:STZ+剪皮+激素注射+异物埋置)。糖尿病模型稳定后每周测量血糖1次,每日称量体重、观察疮面情况、测量创面面积。12 d后处死,石蜡包埋肉芽组织观察其组织病理形态。结果 DMJJ组前5d愈合速度快于其余组(P0.01);DMYW组的愈合时间延长,DMYW组愈合率显著偏低,与其余组比较有统计学意义(P0.01)。造模12 d其余组愈合率无统计学差异,DMYW组愈合率显著低于其余组(P0.01)。结论注射激素大鼠表现出中医"阴证"证型特点,糖尿病加激素注射加异物埋置复合因素造模法能使大鼠创面表现出与临床相似的"阴证"证型特点。     

Application of Electron Paramagnetic Resonance (EPR) Oximetry to Monitor Oxygen in Wounds in Diabetic Models

A lack of oxygen is classically described as a major cause of impaired wound healing in diabetic patients. Even if the role of oxygen in the wound healing process is well recognized, measurement of oxygen levels in a wound remains challenging. The purpose of the present study was to assess the value of electron paramagnetic resonance (EPR) oximetry to monitor pO₂ in wounds during the healing process in diabetic mouse models. Kinetics of wound closure were carried out in streptozotocin (STZ)-treated and db/db mice. The pO₂ was followed repeatedly during the healing process by 1 GHz EPR spectroscopy with lithium phthalocyanine (LiPc) crystals used as oxygen sensor in two different wound models: a full-thickness excisional skin wound and a pedicled skin flap. Wound closure kinetics were dramatically slower in 12-week-old db/db compared to control (db/+) mice, whereas kinetics were not statistically different in STZ-treated compared to control mice. At the center of excisional wounds, measurements were highly influenced by atmospheric oxygen early in the healing process. In pedicled flaps, hypoxia was observed early after wounding. While reoxygenation occurred over time in db/+ mice, hypoxia was prolonged in the diabetic db/db model. This observation was consistent with impaired healing and microangiopathies observed using intravital microscopy. In conclusion, EPR oximetry using LiPc crystals as the oxygen sensor is an appropriate technique to follow wound oxygenation in acute and chronic wounds, in normal and diabetic animals. Nevertheless, the technique is limited for measurements in pedicled skin flaps and cannot be applied to excisional wounds in which diffusion of atmospheric oxygen significantly affects the measurements.     

The plasmid encoding HSP47 enhances collagen expression and promotes skin wound healing in an alloxan-induced diabetic model

The objective of this work was initially to investigate the effects on skin wound healing process by local injection of HSP47 recombinant plasmid in an alloxan-induced diabetic rat model and assess the possibility and utility of gene therapy based on HSP47 plasmid to improve the diabetic skin wound healing. Rats were injected intraperitoneally with alloxan (120 mg/kg) to induce diabetes. The fragment containing the rat 47 kDa heat shock protein (HSP47) gene lacking its own promoter was cloned into plasmids containing a promoter and green fluorescent protein (GFP). The resulting gene constructs were first tested in vitro using 3T3 fibroblast cell line and subsequently in vivo after inducing wounds with alloxan in diabetic rats. Immunohistochemistry, quantitative fluorescent RT-PCR, and Western blotting 3-5 days after plasmid injection were performed to measure the expression changes of HSP47 and collagen I. The results demonstrate an increase of HSP47 levels in vitro in 3T3 fibroblast cells and in vivo in diabetic rat after treatment with plasmids expressing HSP47. The level of collagen I around the wound during the repair process was higher in the treated group than that in the control group, indicating that the constructs may have use in human gene therapy in cases of impaired skin wound healing in diabetes.     

Protective effects of Pycnogenol® on hyperglycemia-induced oxidative damage in the liver of type 2 diabetic rats

Abnormal regulation of glucose and impaired carbohydrate utilization that result from a defective or deficient insulin are the key pathogenic events in type 2 diabetes mellitus (T2DM). Experimental and clinical studies have shown the antidiabetic effects of Pycnogenol^® (PYC). However, the protective effects of PYC on the liver, a major metabolic organ which primarily involves in glucose metabolism and maintains the normal blood glucose level in T2DM model have not been studied. The present study evaluated the beneficial effect of PYC, French maritime pine bark extract, on hyperglycemia and oxidative damage in normal and diabetic rats. Diabetes was induced by feeding rats with a high-fat diet (HFD; 40%) for 2 weeks followed by an intraperitoneal (IP) injection of streptozotocin (STZ; 40 mg/kg; body weight). An IP dose of 10 mg/kg PYC was given continually for 4 weeks after diabetes induction. At the end of the 4-week period, blood was drawn and the rats were then sacrificed, and their livers dissected for biochemical and histopathological assays. In the HFD/STZ group, levels of glycosylated hemoglobin (HbA1c), significantly increased, while hepatic glycogen level decreased. PYC supplementation significantly reversed these parameters. Moreover, supplementation with PYC significantly ameliorated thiobarbituric reactive substances, malonaldehyde, protein carbonyl, glutathione and antioxidant enzymes [glutathione-S-transferase, catalase, superoxide dismutase, glutathione peroxidase and glutathione reductase] in the liver of HFD/STZ rats. These results were supported with histopathological examinations. Although detailed studies are required for the evaluation of the exact protective mechanism of PYC against diabetic complications, these preliminary experimental findings demonstrate that PYC exhibits antidiabetic effects in a rat model of type 2 DM by potentiating the antioxidant defense system. These finding supports the efficacy of PYC for diabetes management.     

A simplified technique for producing an ischemic wound model

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work well in non-ischemic wounds may not work in human diabetic wounds because vasculopathy is one major factor that hinders healing of these wounds. We published an article in 2007 describing a rabbit ear ischemic wound model created by a minimally invasive surgical technique. Since then, we have further simplified the procedure for easier operation. On one ear, three small skin incisions were made on the vascular pedicles, 1-2 cm from the ear base. The central artery was ligated and cut along with the nerve. The whole cranial bundle was cut and ligated, leaving only the caudal branch intact. A circumferential subcutaneous tunnel was made through the incisions, to cut subcutaneous tissues, muscles, nerves, and small vessels. The other ear was used as a non-ischemic control. Four wounds were made on the ventral side of each ear. This technique produces 4 ischemic wounds and 4 non-ischemic wounds in one animal for paired comparisons. After surgery, the ischemic ear was cool and cyanotic, and showed reduced movement and a lack of pulse in the ear artery. Skin temperature of the ischemic ear was 1-10 °C lower than that on the normal ear and this difference was maintained for more than one month. Ear tissue high-energy phosphate contents were lower in the ischemic ear than the control ear. Wound healing times were longer in the ischemic ear than in the non-ischemic ear when the same treatment was used. The technique has now been used on more than 80 rabbits in which 23 were diabetic (diabetes time ranging from 2 weeks to 2 years). No single rabbit has developed any surgical complications such as bleeding, infection, or rupture in the skin incisions. The model has many advantages, such as little skin disruption, longer ischemic time, and higher success rate, when compared to many other models. It can be safely used in animals with reduced resistance, and can also be modified to meet different testing requirements.     

Wound Administration of M2-Polarized Macrophages Does Not Improve Murine Cutaneous Healing Responses

Macrophages play a crucial role in all stages of cutaneous wound healing responses and dysregulation of macrophage function can result in derailed wound repair. The phenotype of macrophages is influenced by the wound microenvironment and evolves during healing from a more pro-inflammatory (M1) profile in early stages, to a less inflammatory pro-healing (M2) phenotype in later stages of repair. The aim of the current study was to investigate the potential of exogenous administration of M2 macrophages to promote wound healing in an experimental mouse model of cutaneous injury. Bone marrow derived macrophages were stimulated in-vitro with IL-4 or IL-10 to obtain two different subsets of M2-polarized cells, M2a or M2c respectively. Polarized macrophages were injected into full-thickness excisional skin wounds of either C57BL/6 or diabetic db/db mice. Control groups were injected with non-polarized (M0) macrophages or saline. Our data indicate that despite M2 macrophages exhibit an anti-inflammatory phenotype in-vitro, they do not improve wound closure in wild type mice while they delay healing in diabetic mice. Examination of wounds on day 15 post-injury indicated delayed re-epithelialization and persistence of neutrophils in M2 macrophage treated diabetic wounds. Therefore, topical application of ex-vivo generated M2 macrophages is not beneficial and contraindicated for cell therapy of skin wounds.     

Neurotensin-loaded collagen dressings reduce inflammation and improve wound healing in diabetic mice

Impaired wound healing is an important clinical problem in diabetes mellitus and results in failure to completely heal diabetic foot ulcers (DFUs), which may lead to lower extremity amputations. In the present study, collagen based dressings were prepared to be applied as support for the delivery of neurotensin (NT), a neuropeptide that acts as an inflammatory modulator in wound healing. The performance of NT alone and NT–loaded collagen matrices to treat wounds in streptozotocin (STZ) diabetic induced mice was evaluated. Results showed that the prepared dressings were not-cytotoxic up to 72 h after contact with macrophages (Raw 264.7) and human keratinocyte (HaCaT) cell lines. Moreover, those cells were shown to adhere to the collagen matrices without noticeable change in their morphology. NT–loaded collagen dressings induced faster healing (17% wound area reduction) in the early phases of wound healing in diabetic wounded mice. In addition, they also significantly reduced inflammatory cytokine expression namely, TNF-α (p < 0.01) and IL-1β (p < 0.01) and decreased the inflammatory infiltrate at day 3 post-wounding (inflammatory phase). After complete healing, metalloproteinase 9 (MMP-9) is reduced in diabetic skin (p < 0.05) which significantly increased fibroblast migration and collagen (collagen type I, alpha 2 (COL1A2) and collagen type III, alpha 1 (COL3A1)) expression and deposition. These results suggest that collagen-based dressings can be an effective support for NT release into diabetic wound enhancing the healing process. Nevertheless, a more prominent scar is observed in diabetic wounds treated with collagen when compared to the treatment with NT alone.     

Effects of umbilical cord blood stem cells on healing factors for diabetic foot injuries

The use of stem or progenitor cells from bone marrow, or peripheral or umbilical cord blood is becoming more common for treatment of diabetic foot problems. These cells promote neovascularization by angiogenic factors and they promote epithelium formation by stimulating cell replication and migration under certain pathological conditions. We investigated the role of CD34 + stem cells from human umbilical cord blood in wound healing using a rat model. Rats were randomly divided into a control group and two groups with diabetes induced by a single dose of 55 mg/kg intraperitoneal streptozocin. Scarred areas 5 mm in diameter were created on the feet of all rats. The diabetic rats constituted the diabetes control group and a diabetes + stem cell group with local injection into the wound site of 0.5 × 106 CD34 + stem cells from human umbilical cord blood. The newly formed skin in the foot wounds following CD34 + stem cell treatment showed significantly improvement by immunohistochemistry and TUNEL staining, and were closer to the wound healing of the control group than the untreated diabetic animals. The increase in FGF expression that accompanied the local injection of CD34 + stem cells indicates that FGF stimulation helped prevent apoptosis. Our findings suggest a promising new treatment approach to diabetic wound healing.     

不同性别大鼠在2型糖尿病造模过程中的稳定性

目的研究不同性别大鼠在2型糖尿病造模过程中的成功率及模型的稳定性。方法高糖高脂饮食联合腹腔注射小剂量链脲佐菌素诱导建立雄、雌性大鼠2型糖尿病模型。成模后所有大鼠每周固定时间测血糖和体重。观察24周后,心脏穿刺取血,测定空腹血糖（FPG）、血清胰岛素（FINS）、HbA1c、甘油三脂（TG）、胆固醇（TC）、高密度脂蛋白-胆固醇（HDL-C）、低密度脂蛋白-胆固醇（LDL-C）。结果单纯高糖高脂饮食喂养,雄、雌性大鼠血糖与正常组无显著差异;STZ注射后,雄性大鼠血糖升高并逐渐平稳,而雌性需两次STZ注射,模型才比较稳定。实验结束时,雄、雌性糖尿病大鼠FPG、FINS、HOMA-IR以及TG、TC、LDL-C均显著升高,说明模型存在胰岛素抵抗和脂代谢紊乱。结论高糖高脂饲料加一次性小剂量链脲佐菌素腹腔注射,可成功建立雄性大鼠2型糖尿病模型,而同等剂量,雌性模型需两次STZ。雄、雌性糖尿病大鼠模型具有高血糖、脂质代谢紊乱和胰岛素抵抗特点。造模成功率及稳定性与性别有关,雄性大鼠较雌性大鼠成模率高,稳定性好,且耗时更短。     

Supplementation with undenatured whey protein during diabetes mellitus improves the healing and closure of diabetic wounds through the rescue of functional long-lived wound macrophages

Long and persistent uncontrolled diabetes tends to degenerate the immune system and increase the incidence of infections in diabetic patients. A serious complication of diabetes is impaired healing, which diminishes physical activity and, in some cases, leads to chronic wounds and limb amputation. Whey proteins (WPs) enhance immunity during early development and have a protective role in some immune disorders. The effect of camel WPs on wound healing in a streptozotocin-induced type 1 diabetic mice model was investigated. Sixty male mice were equally distributed into 3 experimental groups: group 1, non-diabetic control mice; group 2, diabetic mice; and group 3, diabetic mice that were orally supplemented with undenatured WP (100 mg/kg body weight/day for 1 month through oral gavage). We observed that the diabetic mice exhibited delayed wound closure characterized by a significant reduction in collagen deposition, prolonged elevation in inflammatory cytokines, aberrant activation of STAT3 and reduction in the activation of Akt and NF-κB when compared with the control mice. Moreover, in the diabetic mice, the wound-resident macrophages were dysfunctional and demonstrated increased apoptosis, a significant reduction in their phagocytotic ability, aberrant activation of STAT3 and a marked reduction in the activation of Akt. Interestingly, the supplementation of diabetic mice with WP significantly enhanced the collagen deposition, limited the inflammatory stimuli, restored the activation of STAT3, Akt and NF-κB and greatly improved the closure of diabetic wounds compared with the control mice. Most important, the supplementation of diabetic mice with WP rescued functional, long-lived wound-resident macrophages. Our data reveal the benefits of WP supplementation in improving the healing and closure of diabetic wounds.     

Granzyme B degrades extracellular matrix and contributes to delayed wound closure in apolipoprotein E knockout mice

Chronic inflammation and excessive protease activity have a major role in the persistence of non-healing wounds. Granzyme B (GzmB) is a serine protease expressed during chronic inflammation that, in conjunction with perforin, has a well-established role in initiating apoptotic cell death. GzmB is also capable of acting extracellularly, independent of perforin and can degrade several extracellular matrix (ECM) proteins that are critical during wound healing. We used apolipoprotein E (ApoE) knockout (AKO) mice as a novel model of chronic inflammation and impaired wound healing to investigate the role of GzmB in chronic wounds. Wild-type and AKO mice were grown to 7 weeks (young) or 37 weeks (old) of age on a regular chow or high-fat diet (HFD), given a 1-cm diameter full thickness wound on their mid dorsum and allowed to heal for 16 days. Old AKO mice fed a HFD exhibited reduced wound closure, delayed contraction, chronic inflammation and altered ECM remodeling. Conversely, GzmB/ApoE double knockout mice displayed improved wound closure and contraction rates. In addition, murine GzmB was found to degrade both fibronectin and vitronectin derived from healthy mouse granulation tissue. In addition, GzmB-mediated degradation of fibronectin generated a fragment similar in size to that observed in non-healing mouse wounds. These results provide the first direct evidence that GzmB contributes to chronic wound healing in part through degradation of ECM.     

Redox treatment ameliorates diabetes mellitus-induced skin flap necrosis via inhibiting apoptosis and promoting neoangiogenesis

Intractable wound healing is the habitual problem of diabetes mellitus. High blood glucose limits wound healing by interrupting inflammatory responses and inhibiting neoangiogenesis. Oxidative stress is commonly thought to be a major pathogenic cause of diabetic complications. Edaravone (3-methyl-1-phenyl-2-pyrazolin-5-one, EDV) is a free radical scavenger which suppress oxidative stress. This study investigates whether EDV can reduce oxidative stress in wound healing HaCaT/human dermal fibroblasts cells (HDFs) in vitro and in vivo animal model. Cell viability and wound healing assays, FACS flow cytometry, and Hoechst 33342 staining were performed to confirm apoptosis and cytotoxicity in H₂O₂ and EDV-treated HaCaT and HDFs. A streptozotocin-induced hyperglycemic animal model was made in adult C57BL6 mice. Full-thickness skin flap was made on dorsomedial back and re-sutured to evaluate the wound healing process. EDV was delivered slowly in the skin flap with degradable fibrin glue. The flap was monitored and analyzed on postoperative days 1, 3, and 5. CD31/DAPI staining was done to detect newly formed blood vessels. The expression levels of NF-κB, bcl-2, NOX3, and STAT3 proteins in C57BL6 mouse tissues were also examined. The wound healing process in hyper- and normoglycemic mice showed a difference in protein expression, especially in oxidative stress management and angiogenesis. Exogenous H₂O₂ reduced cell viability in a proportion to the concentration via apoptosis. EDV protected HaCaT cells and HDFs from H₂O₂ induced reactive oxygen species cell damage and apoptosis. In the mouse model, EDV with fibrin resulted in less necrotic areas and increased angiogenesis on postoperative day 5, compared to sham-treated mice. Our results indicate that EDV could protect H₂O₂-induced cellular injury via inhibiting early apoptosis and inflammation and also increasing angiogenesis. EDV might be valuable in the treatment of diabetic wounds that oxidative stress has been implicated.     

14S,21R-dihydroxydocosahexaenoic acid remedies impaired healing and mesenchymal stem cell functions in diabetic wounds

Treatment of diabetes-impaired wound healing remains a major unresolved medical challenge. Here, we identified suppressed formation of a novel reparative lipid mediator 14S,21R-dihydroxydocosa-4Z,7Z,10Z,12E,16Z,19Z-hexaenoic acid (14S,21R-diHDHA) in cutaneous wounds of diabetic db/db mice. These results indicate that diabetes impedes the biosynthetic pathways of 14S,21R-diHDHA in skin wounds. Administration of exogenous 14S,21R-diHDHA to wounds in diabetic animals rescued healing and angiogenesis. When db/db mesenchymal stem cells (MSCs) were administered together with 14S,21R-diHDHA to wounds in diabetic animals, they coacted to accelerate wound re-epithelialization, granulation tissue formation, and synergistically improved vascularization. In the pivotal cellular processes of angiogenesis, 14S,21R-diHDHA enhanced VEGF release, vasculature formation, and migration of db/db dermal microvascular endothelial cells (DMVECs), as well as remedied paracrine angiogenic functions of db/db MSCs, including VEGF secretion and the promotion of DMVEC migration and vasculature formation. Our results show that 14S,21R-diHDHA activates the p38 MAPK pathway in wounds, db/db MSCs, and DMVECs. Overall, the impeded formation of 14S,21R-diHDHA described in this study suggests that diabetes could affect the generation of pro-healing lipid mediators in wound healing. By restoring wound healing and MSC functions, 14S,21R-diHDHA is a new lead for the development of better therapeutics used in treating wounds of diabetics.     

Streptozotocin, Type I Diabetes Severity and Bone

As many as 50% of adults with type I (T1) diabetes exhibit bone loss and are at increased risk for fractures. Therapeutic development to prevent bone loss and/or restore lost bone in T1 diabetic patients requires knowledge of the molecular mechanisms accounting for the bone pathology. Because cell culture models alone cannot fully address the systemic/metabolic complexity of T1 diabetes, animal models are critical. A variety of models exist including spontaneous and pharmacologically induced T1 diabetic rodents. In this paper, we discuss the streptozotocin (STZ)-induced T1 diabetic mouse model and examine dose-dependent effects on disease severity and bone. Five daily injections of either 40 or 60 mg/kg STZ induce bone pathologies similar to spontaneously diabetic mouse and rat models and to human T1 diabetic bone pathology. Specifically, bone volume, mineral apposition rate, and osteocalcin serum and tibia messenger RNA levels are decreased. In contrast, bone marrow adiposity and aP2 expression are increased with either dose. However, high-dose STZ caused a more rapid elevation of blood glucose levels and a greater magnitude of change in body mass, fat pad mass, and bone gene expression (osteocalcin, aP2). An increase in cathepsin K and in the ratio of RANKL/OPG was noted in high-dose STZ mice, suggesting the possibility that severe diabetes could increase osteoclast activity, something not seen with lower doses. This may contribute to some of the disparity between existing studies regarding the role of osteoclasts in diabetic bone pathology. Examination of kidney and liver toxicity indicate that the high STZ dose causes some liver inflammation. In summary, the multiple low-dose STZ mouse model exhibits a similar bone phenotype to spontaneous models, has low toxicity, and serves as a useful tool for examining mechanisms of T1 diabetic bone loss.     

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.     

Antimycotic ciclopirox olamine in the diabetic environment promotes angiogenesis and enhances wound healing

Diabetic wounds remain a major medical challenge with often disappointing outcomes despite the best available care. An impaired response to tissue hypoxia and insufficient angiogenesis are major factors responsible for poor healing in diabetic wounds. Here we show that the antimycotic drug ciclopirox olamine (CPX) can induce therapeutic angiogenesis in diabetic wounds. Treatment with CPX in vitro led to upregulation of multiple angiogenic genes and increased availability of HIF-1α. Using an excisional wound splinting model in diabetic mice, we showed that serial topical treatment with CPX enhanced wound healing compared to vehicle control treatment, with significantly accelerated wound closure, increased angiogenesis, and increased dermal cellularity. These findings offer a promising new topical pharmacologic therapy for the treatment of diabetic wounds.