Formulações de entrega de peptídeos antimicrobianos para a cicatrização de feridas

National Patent (INPI)A presente divulgação diz respeito a formulações de entrega de peptídeos antimicrobianos para a cicatrização de feridas, em particular diz respeito a uma composição que compreende pelo menos um polímero, pelo menos um péptido antimicrobiano compreendendo pelo menos uma sequência 95% idêntica à sequência seq. id 1, em particular idêntica à seq id 1. e com uma quantidade terapeuticamente eficaz de todos componentes anteriores.info:eu-repo/semantics/publishedVersio

Release of insulin from PLGA-alginate dressing stimulates regenerative healing of burn wounds in rats

Burn wound healing involves a complex set of overlapping processes in an environment conducive to ischemia, inflammation, and infection costing $7.5 billion/year in the US alone, in addition to the morbidity and mortality that occur when the burns are extensive. We previously showed that insulin, when topically applied to skin excision wounds, accelerates re-epithelialization, and stimulates angiogenesis. More recently, we developed an alginate sponge dressing (ASD) containing insulin encapsulated in PLGA microparticles that provides a sustained release of bioactive insulin for >20days in a moist and protective environment. We hypothesized that insulin-containing ASD accelerates burn healing and stimulates a more regenerative, less scarring, healing. Using a heat-induced burn injury in rats, we show that burns treated with dressings containing 0.04mg insulin/cm2, every three days for 9 days, have faster closure, faster rate of disintegration of dead tissue, and decreased oxidative stress.In addition, in insulin-treated wounds the pattern of neutrophil inflammatory response suggests faster clearing of the burn dead tissue. We also observe faster resolution of the pro-inflammatory macrophages. We also found that insulin stimulates collagen deposition and maturation with the fibers organized more like a basket weave (normal skin) than aligned and crosslinked (scar tissue). In summary , application of ASD-containing insulin-loaded PLGA particles on burns every three days stimulates faster and more regenerative healing. These results suggest insulin as a potential therapeutic agent in burn healing and, because of its long history of safe use in humans, insulin could become one of the treatments of choice when repair and regeneration are critical for proper tissue function.This work was supported by the National Natural Science Fund of China [grant numbers 81170761 and 81270909 (to Y.L.)]; the Natural Sciences and Engineering Research Council of Canada [grant numbers 204794-2011 (to M.H.) and private donor (to M.M.-G.)]

Generating and Reversing Chronic Wounds in Diabetic Mice by Manipulating Wound Redox Parameters

By 2025, more than 500 M people worldwide will suffer from diabetes; 125 M will develop foot ulcer(s) and 20 M will undergo an amputation, creating a major health problem. Understanding how these wounds become chronic will provide insights to reverse chronicity. We hypothesized that oxidative stress (OS) in wounds is a critical component for generation of chronicity. We used the db/db mouse model of impaired healing and inhibited, at time of injury, two major antioxidant enzymes, catalase and glutathione peroxidase, creating high OS in the wounds. This was necessary and sufficient to trigger wounds to become chronic. The wounds initially contained a polymicrobial community that with time selected for specific biofilm-forming bacteria. To reverse chronicity we treated the wounds with the antioxidants α-tocopherol and N-acetylcysteine and found that OS was highly reduced, biofilms had increased sensitivity to antibiotics, and granulation tissue was formed with proper collagen deposition and remodeling. We show for the first time generation of chronic wounds in which biofilm develops spontaneously, illustrating importance of early and continued redox imbalance coupled with the presence of biofilm in development of wound chronicity. This model will help decipher additional mechanisms and potentially better diagnosis of chronicity and treatment of human chronic wounds

Cigarette Smoke Toxins Deposited on Surfaces: Implications for Human Health

Cigarette smoking remains a significant health threat for smokers and nonsmokers alike. Secondhand smoke (SHS) is intrinsically more toxic than directly inhaled smoke. Recently, a new threat has been discovered – Thirdhand smoke (THS) – the accumulation of SHS on surfaces that ages with time, becoming progressively more toxic. THS is a potential health threat to children, spouses of smokers and workers in environments where smoking is or has been allowed. The goal of this study is to investigate the effects of THS on liver, lung, skin healing, and behavior, using an animal model exposed to THS under conditions that mimic exposure of humans. THS-exposed mice show alterations in multiple organ systems and excrete levels of NNAL (a tobacco-specific carcinogen biomarker) similar to those found in children exposed to SHS (and consequently to THS). In liver, THS leads to increased lipid levels and non-alcoholic fatty liver disease, a precursor to cirrhosis and cancer and a potential contributor to cardiovascular disease. In lung, THS stimulates excess collagen production and high levels of inflammatory cytokines, suggesting propensity for fibrosis with implications for inflammation-induced diseases such as chronic obstructive pulmonary disease and asthma. In wounded skin, healing in THS-exposed mice has many characteristics of the poor healing of surgical incisions observed in human smokers. Lastly, behavioral tests show that THS-exposed mice become hyperactive. The latter data, combined with emerging associated behavioral problems in children exposed to SHS/THS, suggest that, with prolonged exposure, they may be at significant risk for developing more severe neurological disorders. These results provide a basis for studies on the toxic effects of THS in humans and inform potential regulatory policies to prevent involuntary exposure to THS

Strategies for Development of Chronic Wounds Using the LIGHT -/- Mouse and a Diabetic Mouse: From Mechanism to Treatment

Wound healing involves many cellular and molecular processes that are integrated in several sequential and overlapping phases, hemostasis, controlled oxidative stress, inflammation, granulation tissue formation, and remodeling. Impaired-healing and chronic wounds exhibit defective regulation of one or more of these processes that leads to conditions such as diabetic foot ulcers, and other similar chronic wounds that impact ∼6.5M people and cost ∼$25B/year in the US alone. Great efforts have been made to stimulate healing of these wounds, including the development of animal models mimicking chronic wounds in order to understand how they develop but success has been limited. Recently, we developed a mouse model of impaired healing that became chronic in presence of biofilm-forming bacteria. I took an integrated approach by using various cellular and molecular approaches to study the wound microenvironment. Using the LIGHT-/- model of impaired healing, I showed that the wounds in these mice, very early during the process of healing have elevated levels of reactive oxygen species (ROS) and reactive nitrogen species (RNS), increased lactate levels and reduced pH that could potentially damage the healing tissue. With the use of luminol I showed, for the very first time, real time monitoring of increase in oxidative stress levels. In addition, I showed that the detrimental effects of increases in ROS and RNS significantly increased damage to DNA, lipid peroxidation and protein nitrosylation. Furthermore, using a lipidomics approach I showed an increase in inflammatory lipids and lipids involved in platelet function. The findings were confirmed by the increases in inflammation, platelet aggregation and reduced bleeding time post wounding. I then showed that by exacerbating the levels of ROS by inhibition major antioxidant enzymes, glutathione peroxidase and catalase, and introducing previously isolated biofilm forming bacteria on the wound bed, led to the development of chronic wounds. The wounds remained open and persistent inflammation was marked by the clear presence of neutrophils and macrophages in the wound tissue. The granulation tissue was poorly formed and there was loss of collagen bundles. Furthermore, I also showed that the bacteria were capable of forming biofilms and were resistant to antibiotics. These results confirmed that redox imbalance and presence of bacteria were crucial elements for chronic wound formation. I then tested the possibility that exacerbated oxidative stress was critical for chronic wound development by performing similar experiment in a diabetic mouse model, the db/db. I showed that only one dose of inhibitors to the antioxidant enzymes at the time of wounding was sufficient to cause the wound to become chronic by 20 days and spontaneously harbor biofilm-forming bacteria. The chronic wounds in these mice did not heal for as long as 90 days. I also showed that the bacteria was resistant to antibiotics and that there were embedded in the extrapolysaccharide (EPS) matrix. To confirm the importance of redox stress, I reversed the stress levels by treating the wounds with antioxidants, N-acetyl cysteine and α-tocopherol, and showed that the wounds healed with decreased levels of oxidative stress, reduction in biofilm forming bacteria, reduction in biofilm on the wound bed, better tissue structure with proper collagen bundles and proper differentiating epithelial cells. The studies presented here provide an in vivo model of chronic wounds that captures many of the clinical aspects of human chronic wounds and that may provide insights into the mechanisms involved in chronic wound development, including the natural growth of biofilm in situ. These findings, taken together suggest the robustness of the LIGHT-/- and db/db models of impaired and chronic wounds respectively that warrants further research to capture the underlying mechanisms in the development of human chronic wounds and hence uncovering potential targets to treat such wounds

Strategies for Development of Chronic Wounds Using the LIGHT-/- Mouse and a Diabetic Mouse: From Mechanism to Treatment

Wound healing involves many cellular and molecular processes that are integrated in several sequential and overlapping phases, hemostasis, controlled oxidative stress, inflammation, granulation tissue formation, and remodeling. Impaired-healing and chronic wounds exhibit defective regulation of one or more of these processes that leads to conditions such as diabetic foot ulcers, and other similar chronic wounds that impact ~6.5M people and cost ~$25B/year in the US alone. Great efforts have been made to stimulate healing of these wounds, including the development of animal models mimicking chronic wounds in order to understand how they develop but success has been limited. Recently, we developed a mouse model of impaired healing that became chronic in presence of biofilm-forming bacteria. I took an integrated approach by using various cellular and molecular approaches to study the wound microenvironment. Using the LIGHT-/- model of impaired healing, I showed that the wounds in these mice, very early during the process of healing have elevated levels of reactive oxygen species (ROS) and reactive nitrogen species (RNS), increased lactate levels and reduced pH that could potentially damage the healing tissue. With the use of luminol I showed, for the very first time, real time monitoring of increase in oxidative stress levels. In addition, I showed that the detrimental effects of increases in ROS and RNS significantly increased damage to DNA, lipid peroxidation and protein nitrosylation. Furthermore, using a lipidomics approach I showed an increase in inflammatory lipids and lipids involved in platelet function. The findings were confirmed by the increases in inflammation, platelet aggregation and reduced bleeding time post wounding. I then showed that by exacerbating the levels of ROS by inhibition major antioxidant enzymes, glutathione peroxidase and catalase, and introducing previously isolated biofilm forming bacteria on the wound bed, led to the development of chronic wounds. The wounds remained open and persistent inflammation was marked by the clear presence of neutrophils and macrophages in the wound tissue. The granulation tissue was poorly formed and there was loss of collagen bundles. Furthermore, I also showed that the bacteria were capable of forming biofilms and were resistant to antibiotics. These results confirmed that redox imbalance and presence of bacteria were crucial elements for chronic wound formation.I then tested the possibility that exacerbated oxidative stress was critical for chronic wound development by performing similar experiment in a diabetic mouse model, the db/db. I showed that only one dose of inhibitors to the antioxidant enzymes at the time of wounding was sufficient to cause the wound to become chronic by 20 days and spontaneously harbor biofilm-forming bacteria. The chronic wounds in these mice did not heal for as long as 90 days. I also showed that the bacteria was resistant to antibiotics and that there were embedded in the extrapolysaccharide (EPS) matrix. To confirm the importance of redox stress, I reversed the stress levels by treating the wounds with antioxidants, N-acetyl cysteine and α-tocopherol, and showed that the wounds healed with decreased levels of oxidative stress, reduction in biofilm forming bacteria, reduction in biofilm on the wound bed, better tissue structure with proper collagen bundles and proper differentiating epithelial cells. The studies presented here provide an in vivo model of chronic wounds that captures many of the clinical aspects of human chronic wounds and that may provide insights into the mechanisms involved in chronic wound development, including the natural growth of biofilm in situ. These findings, taken together suggest the robustness of the LIGHT-/- and db/db models of impaired and chronic wounds respectively that warrants further research to capture the underlying mechanisms in the development of human chronic wounds and hence uncovering potential targets to treat such wounds