RESUMO
The sphingolipid, ceramide-1-phosphate (C1P), has been shown to promote the inflammatory phase and inhibit the proliferation and remodeling stages of wound repair via direct interaction with group IVA cytosolic phospholipase A2, a regulator of eicosanoid biosynthesis that fine-tunes the behaviors of various cell types during wound healing. However, the anabolic enzyme responsible for the production of C1P that suppresses wound healing as well as bioactive eicosanoids and target receptors that drive enhanced wound remodeling have not been characterized. Herein, we determined that decreasing C1P activity via inhibitors or genetic ablation of the anabolic enzyme ceramide kinase (CERK) significantly enhanced wound healing phenotypes. Importantly, postwounding inhibition of CERK enhanced the closure rate of acute wounds, improved the quality of healing, and increased fibroblast migration via a "class switch" in the eicosanoid profile. This switch reduced pro-inflammatory prostaglandins (e.g., prostaglandin E2) and increased levels of 5-hydroxyeicosatetraenoic acid and the downstream metabolite 5-oxo-eicosatetraenoic acid (5-oxo-ETE). Moreover, dermal fibroblasts from mice with genetically ablated CERK showed enhanced wound healing markers, while blockage of the murine 5-oxo-ETE receptor (oxoeicosanoid receptor 1) inhibited the enhanced migration phenotype of these cell models. Together, these studies reinforce the vital roles eicosanoids play in the wound healing process and demonstrate a novel role for CERK-derived C1P as a negative regulator of 5-oxo-ETE biosynthesis and the activation of oxoeicosanoid receptor 1 in wound healing. These findings provide foundational preclinical results for the use of CERK inhibitors to shift the balance from inflammation to resolution and increase the wound healing rate.
Assuntos
Ceramidas , Fosfotransferases (Aceptor do Grupo Álcool) , Animais , Ácidos Araquidônicos , Movimento Celular , Ceramidas/metabolismo , Eicosanoides , Camundongos , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Cicatrização/genéticaRESUMO
The normal wound healing response is characterized by a progression from clot formation, to an inflammatory phase, to a repair phase, and finally, to remodeling. In many chronic wounds there is an extended inflammatory phase that stops this progression. In order to understand the inflammatory phase in more detail, we developed an ordinary differential equation model that accounts for two systemic mediators that are known to modulate this phase, estrogen (a protective hormone during wound healing) and cortisol (a hormone elevated after trauma that slows healing). This model describes the interactions in the wound between wound debris, pathogens, neutrophils and macrophages and the modulation of these interactions by estrogen and cortisol. A collection of parameter sets, which qualitatively match published data on the dynamics of wound healing, was chosen using Latin Hypercube Sampling. This collection of parameter sets represents normal healing in the population as a whole better than one single parameter set. Including the effects of estrogen and cortisol is a necessary step to creating a patient specific model that accounts for gender and trauma. Utilization of math modeling techniques to better understand the wound healing inflammatory phase could lead to new therapeutic strategies for the treatment of chronic wounds. This inflammatory phase model will later become the inflammatory subsystem of our full wound healing model, which includes fibroblast activity, collagen accumulation and remodeling.
Assuntos
Mediadores da Inflamação/metabolismo , Inflamação/patologia , Modelos Biológicos , Cicatrização , Estrogênios/farmacologia , Humanos , Hidrocortisona/farmacologia , Imunidade/efeitos dos fármacos , Cinética , Ativação de Macrófagos/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Reprodutibilidade dos Testes , Fatores de Tempo , Cicatrização/efeitos dos fármacosRESUMO
In these studies, the role of ceramide-1-phosphate (C1P) in the wound-healing process was investigated. Specifically, fibroblasts isolated from mice with the known anabolic enzyme for C1P, ceramide kinase (CERK), ablated (CERK(-/-) mice) and their wild-type littermates (CERK(+/+)) were subjected to in vitro wound-healing assays. Simulation of mechanical trauma of a wound by scratching a monolayer of fibroblasts from CERK(+/+) mice demonstrated steadily increasing levels of arachidonic acid in a time-dependent manner in stark contrast to CERK(-/-) fibroblasts. This observed difference was reflected in scratch-induced eicosanoid levels. Similar, but somewhat less intense, changes were observed in a more complex system utilizing skin biopsies obtained from CERK-null mice. Importantly, C1P levels increased during the early stages of human wound healing correlating with the transition from the inflammatory stage to the peak of the fibroplasia stage (e.g., proliferation and migration of fibroblasts). Finally, the loss of proper eicosanoid response translated into an abnormal migration pattern for the fibroblasts isolated from CERK(-/-) As the proper migration of fibroblasts is one of the necessary steps of wound healing, these studies demonstrate a novel requirement for the CERK-derived C1P in the proper healing response of wounds.
Assuntos
Movimento Celular/efeitos dos fármacos , Eicosanoides/farmacologia , Fibroblastos/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Cicatrização/efeitos dos fármacos , Animais , Movimento Celular/genética , Ceramidas/genética , Ceramidas/metabolismo , Fibroblastos/citologia , Camundongos , Camundongos Knockout , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Cicatrização/genéticaRESUMO
Uncontrolled inflammation is linked to poor outcomes in sepsis and wound healing, both of which proceed through distinct inflammatory and resolution phases. Eicosanoids are a class of bioactive lipids that recruit neutrophils and other innate immune cells. The interaction of ceramide 1-phosphate (C1P) with the eicosanoid biosynthetic enzyme cytosolic phospholipase A2 (cPLA2) reduces the production of a subtype of eicosanoids called oxoeicosanoids. We investigated the effect of shifting the balance in eicosanoid biosynthesis on neutrophil polarization and function. Knockin mice expressing a cPLA2 mutant lacking the C1P binding site (cPLA2αKI/KI mice) showed enhanced and sustained neutrophil infiltration into wounds and the peritoneum during the inflammatory phase of wound healing and sepsis, respectively. The mice exhibited improved wound healing and reduced susceptibility to sepsis, which was associated with an increase in anti-inflammatory N2-type neutrophils demonstrating proresolution behaviors and a decrease in proinflammatory N1-type neutrophils. The N2 polarization of cPLA2αKI/KI neutrophils resulted from increased oxoeicosanoid biosynthesis and autocrine signaling through the oxoeicosanoid receptor OXER1 and partially depended on OXER1-dependent inhibition of the pentose phosphate pathway (PPP). Thus, C1P binding to cPLA2α suppresses neutrophil N2 polarization, thereby impairing wound healing and the response to sepsis.
Assuntos
Neutrófilos , Sepse , Animais , Camundongos , Sepse/genética , Comunicação Autócrina , Fosfolipases A2 do Grupo IV/genética , InflamaçãoRESUMO
Wound healing is a complex biological process which involves many cell types and biochemical signals and which progresses through multiple, overlapping phases. In this manuscript, we develop a model of collagen accumulation as a marker of wound healing. The mathematical model is a system of ordinary differential equations which tracks fibroblasts, collagen, inflammation and pathogens. The model was validated by comparison to the normal time course of wound healing where appropriate activity for the inflammatory, proliferative and remodeling phases was recorded. Further validation was made by comparison to collagen accumulation experiments by Madden and Peacock (Ann. Surg. 174(3):511-520, 1971). The model was then used to investigate the impact of local oxygen levels on wound healing. Finally, we present a comparison of two wound healing therapies, antibiotics and increased fibroblast proliferation. This model is a step in developing a comprehensive model of wound healing which can be used to develop and test new therapeutic treatments.
Assuntos
Colágeno/metabolismo , Inflamação/metabolismo , Modelos Biológicos , Cicatrização/fisiologia , Fibroblastos/metabolismo , Humanos , Inflamação/patologia , Oxigênio/metabolismoRESUMO
The complex interactions that characterize acute wound healing have stymied the development of effective therapeutic modalities. The use of computational models holds the promise to improve our basic approach to understanding the process. By modifying an existing ordinary differential equation model of systemic inflammation to simulate local wound healing, we expect to improve the understanding of the underlying complexities of wound healing and thus allow for the development of novel, targeted therapeutic strategies. The modifications in this local acute wound healing model include: evolution from a systemic model to a local model, the incorporation of fibroblast activity, and the effects of tissue oxygenation. Using these modifications we are able to simulate impaired wound healing in hypoxic wounds with varying levels of contamination. Possible therapeutic targets, such as fibroblast death rate and rate of fibroblast recruitment, have been identified by computational analysis. This model is a step toward constructing an integrative systems biology model of human wound healing.
Assuntos
Biologia Computacional , Modelos Biológicos , Cicatrização/fisiologia , Fibroblastos/fisiologia , Humanos , Inflamação/fisiopatologia , Oxigênio/sangue , Pele/lesões , Fenômenos Fisiológicos da Pele , Infecção dos Ferimentos/fisiopatologiaRESUMO
Chronic wounds present a unique therapeutic challenge to heal. Chronic wounds are colonized with bacteria and the presence of a biofilm that further inhibits the normal wound healing processes, and are locked into a very damaging proinflammatory response. The treatment of chronic wounds requires a coordinated approach, including debridement of devitalized tissue, minimizing bacteria and biofilm, control of inflammation, and the use of specialized dressings to address the specific aspects of the particular nonhealing ulcer.
Assuntos
Angiopatias Diabéticas/fisiopatologia , Úlcera Cutânea/fisiopatologia , Cicatrização/fisiologia , Anti-Infecciosos/uso terapêutico , Biofilmes/efeitos dos fármacos , Doença Crônica , Citocinas/fisiologia , Angiopatias Diabéticas/imunologia , Angiopatias Diabéticas/terapia , Farmacorresistência Bacteriana/fisiologia , Quimioterapia Combinada , Humanos , Imunidade Celular/fisiologia , Peptídeo Hidrolases/fisiologia , Úlcera Cutânea/imunologia , Úlcera Cutânea/terapia , Cicatrização/imunologia , Infecção dos Ferimentos/imunologia , Infecção dos Ferimentos/fisiopatologia , Infecção dos Ferimentos/terapiaRESUMO
It is well recognized that stress of any nature will cause a delay in the wound healing response. This delayed healing response appears closely associated with immune regulators. In this study, CD-1 mice were injected with a long acting form of methyl prednisolone to cause a steroid-induced immune suppression. After 24 hours, two 6-mm full thickness wounds were placed on the animals' backs and one group of animals received the immune-regulating hormone, androstenediol. Wound contraction was quantified by planimetry for the subsequent 14 days. Animals that were stressed with methyl prednisolone but receiving androstenediol contracted their open wounds at faster rates compared with methyl prednisolone-stressed animals treated with the vehicle alone. These findings suggest that restoration of immune regulation by androstenediol can reverse the delayed open wound contraction secondary to steroid stress.
Assuntos
Anabolizantes/farmacologia , Androstenodiol/farmacologia , Glucocorticoides/farmacologia , Metilprednisolona/farmacologia , Cicatrização/efeitos dos fármacos , Ferimentos e Lesões/tratamento farmacológico , Animais , Modelos Animais de Doenças , Masculino , Camundongos , Pele/efeitos dos fármacos , Pele/lesões , Cicatrização/imunologia , Ferimentos e Lesões/imunologiaRESUMO
The sphingolipid ceramide 1-phosphate (C1P) directly binds to and activates group IVA cytosolic phospholipase A2 (cPLA2α) to stimulate the production of eicosanoids. Because eicosanoids are important in wound healing, we examined the repair of skin wounds in knockout (KO) mice lacking cPLA2α and in knock-in (KI) mice in which endogenous cPLA2α was replaced with a mutant form having an ablated C1P interaction site. Wound closure rate was not affected in the KO or KI mice, but wound maturation was enhanced in the KI mice compared to that in wild-type controls. Wounds in KI mice displayed increased infiltration of dermal fibroblasts into the wound environment, increased wound tensile strength, and a higher ratio of type I:type III collagen. In vitro, primary dermal fibroblasts (pDFs) from KI mice showed substantially increased collagen deposition and migration velocity compared to pDFs from wild-type and KO mice. KI mice also showed an altered eicosanoid profile of reduced proinflammatory prostaglandins (PGE2 and TXB2) and an increased abundance of certain hydroxyeicosatetraenoic acid (HETE) species. Specifically, an increase in 5-HETE enhanced dermal fibroblast migration and collagen deposition. This gain-of-function role for the mutant cPLA2α was also linked to the relocalization of cPLA2α and 5-HETE biosynthetic enzymes to the cytoplasm and cytoplasmic vesicles. These findings demonstrate the regulation of key wound-healing mechanisms in vivo by a defined protein-lipid interaction and provide insights into the roles that cPLA2α and eicosanoids play in orchestrating wound repair.
Assuntos
Ceramidas/metabolismo , Fosfolipases A2 do Grupo IV/genética , Fosfolipases A2 do Grupo IV/metabolismo , Cicatrização , Animais , Movimento Celular , Núcleo Celular/metabolismo , Proliferação de Células , Colágeno/metabolismo , Citoplasma/metabolismo , Citosol/metabolismo , Dinoprostona/metabolismo , Eicosanoides/metabolismo , Fibroblastos/metabolismo , Genótipo , Ácidos Hidroxieicosatetraenoicos/farmacologia , Inflamação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia de Fluorescência , Fenótipo , Pele/metabolismo , Resistência à Tração , Tromboxano B2/metabolismoRESUMO
BACKGROUND: Multiple physiologic impairments are responsible for chronic wounds. A cell line grown which retains its phenotype from patient wounds would provide means of testing new therapies. Clinical information on patients from whom cells were grown can provide insights into mechanisms of specific disease such as diabetes or biological processes such as aging. The objective of this study was 1) To culture human cells derived from patients with chronic wounds and to test the effects of putative therapies, Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) on these cells. 2) To describe a methodology to create fibroblast cell lines from patients with chronic wounds. METHODS: Patient biopsies were obtained from 3 distinct locations on venous ulcers. Fibroblasts derived from different wound locations were tested for their migration capacities without stimulators and in response to GM-CSF. Another portion of the patient biopsy was used to develop primary fibroblast cultures after rigorous passage and antimicrobial testing. RESULTS: Fibroblasts from the non-healing edge had almost no migration capacity, wound base fibroblasts were intermediate, and fibroblasts derived from the healing edge had a capacity to migrate similar to healthy, normal, primary dermal fibroblasts. Non-healing edge fibroblasts did not respond to GM-CSF. Six fibroblast cell lines are currently available at the National Institute on Aging (NIA) Cell Repository. CONCLUSION: We conclude that primary cells from chronic ulcers can be established in culture and that they maintain their in vivo phenotype. These cells can be utilized for evaluating the effects of wound healing stimulators in vitro.
Assuntos
Técnicas de Cultura de Células/métodos , Linhagem Celular , Fibroblastos , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Cicatrização/efeitos dos fármacos , Movimento Celular/fisiologia , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Fenótipo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Úlcera Varicosa/patologia , Cicatrização/fisiologiaRESUMO
Non-healing wounds represent a significant cause of morbidity and mortality for a large portion of the adult population. Wounds that fail to heal are entrapped in a self-sustaining cycle of chronic inflammation leading to the destruction of the extracellular matrix. Among cancer patients, malnutrition, radiation, physical dehabilitation, chemotherapy, and the malignancy itself increase the likelihood of chronic wound formation, and these co-morbidity factors inhibit the normal wound healing process. Current wound treatments are aimed at some, but not all, of the underlying causes responsible for delayed healing of wounds. These impediments block the normal processes that allow normal progression through the specific stages of wound healing. This review summarizes the current information regarding the management and treatment of complex wounds that fail to heal normally and offers some insights into novel future therapies [Menke N, Ward KR, Diegelmann R. Non-healing wounds. Emerg Med Rep 2007; 28(4).,Menke NB, Ward KR, Witten TM, Bonchev DG, Diegelmann RF. Impaired wound healing. Clin Dermatol 2007;25:19-25].
Assuntos
Cicatrização/fisiologia , Ferimentos e Lesões/terapia , Doença Crônica , Humanos , Infecção dos Ferimentos/patologia , Infecção dos Ferimentos/terapia , Ferimentos e Lesões/microbiologia , Ferimentos e Lesões/patologiaRESUMO
OBJECTIVE: The healing of wounds is critical in protecting the human body against environmental factors. The mechanisms involving protein expression during this complex physiological process have not been fully elucidated. APPROACH: Here, we use reverse-phase protein microarrays (RPPA) involving 94 phosphoproteins to study tissue samples from tubes implanted in healing dermal wounds in seven human subjects tracked over two weeks. We compare the proteomic profiles to proteomes of controls obtained from skin biopsies from the same subjects. MAIN RESULTS: Compared to previous proteomic studies of wound healing, our approach focuses on wound tissue instead of wound fluid, and has the sensitivity to go beyond measuring only highly abundant proteins. To study the temporal dynamics of networks involved in wound healing, we applied two network analysis methods that integrate the experimental results with prior knowledge about protein-protein physical and regulatory interactions, as well as higher-level biological processes and associated pathways. SIGNIFICANCE: We uncovered densely connected networks of proteins that are up- or down-regulated during human wound healing, as well as their relationships to microRNAs and to proteins outside of our set of targets that we measured with proteomic microarrays.
Assuntos
Proteômica , Fenômenos Fisiológicos da Pele , Pele/metabolismo , Cicatrização , Regulação para Baixo , Humanos , Fosfoproteínas/metabolismo , Análise Serial de Proteínas , Regulação para CimaRESUMO
OBJECTIVE: This pilot study tested the potential of puromycin (PUR) to inhibit protein synthesis and reduce oxygen utilization in a non-hibernating, whole animal preparation. METHODS: After anesthesia and instrumentation, male rats received a single dose of PUR or 0.9% saline (control), followed 60 min later with [(35)S] methionine/cysteine radiolabeling. Thirty minutes after isotope injection, organ biopsies were taken for quantification of de novo protein synthesis. Arterial and central venous blood gases were obtained at baseline and 60 min after injection of PUR or 0.9% saline. Temperature, mean arterial pressure (MAP), and heart rate were recorded continuously. RESULTS: Animals receiving PUR demonstrated significant reductions in protein synthesis in all organ systems sampled (p<0.05). The overall reduction averaged 67.8%. Central venous oxygen saturations (S(cv)O(2)) were higher in the PUR group than the controls at 60 min (90+/-2% versus 80+/-4%, p<0.05). The oxygen extraction ratio (O(2)ER) decreased from 16.1+/-1.7% to 6.8+/-1.2% in the PUR group (p<0.05) and increased from 12.5+/-3.2% to 16.0+/-4.2% in the controls (p=0.44). There was no difference in temperature, MAP, heart rate or blood gas variables, other than S(cv)O(2), at baseline or 60 min between groups. CONCLUSIONS: These results demonstrate that PUR is capable of reducing whole body protein synthesis significantly within a relatively short duration of time. This appears to decrease whole body oxygen utilization as evidenced by an increase in S(cv)O(2) and a decrease in O(2)ER. Protein synthesis inhibition may reduce metabolic demands and should be tested for its potential to improve outcomes where oxygen demands exceed oxygen delivery.
Assuntos
Regulação para Baixo/efeitos dos fármacos , Proteínas/metabolismo , Puromicina/farmacologia , Animais , Gasometria , Masculino , Consumo de Oxigênio/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Choque/fisiopatologiaRESUMO
Nonhealing wounds represent a significant cause of morbidity and mortality for a large portion of the population. One of the underlying mechanisms responsible for the failure of chronic wounds to heal is an out-of-control inflammatory response that is self-sustaining. Underappreciation of the inherent complexity of the healing wound has led to the failure of monotherapies, with no significant reduction in wound healing times. A model of the inflammatory profile of a nonhealing wound is one in which the equilibrium between synthesis and degradation has been shifted toward degradation. This review summarizes the current information regarding acute wound healing responses as contrasted to the delayed response characteristic of chronic wounds. In addition, some initial complexity theoretical models are proposed to define and explain the underlying pathophysiology.
Assuntos
Úlcera Cutânea/fisiopatologia , Pele/lesões , Pele/fisiopatologia , Cicatrização , Ferimentos Penetrantes/fisiopatologia , Doença Aguda , Doença Crônica , Humanos , Inflamação/complicações , Modelos Teóricos , Pele/imunologia , Úlcera Cutânea/imunologia , Úlcera Cutânea/terapia , Falha de Tratamento , Cicatrização/fisiologia , Ferimentos Penetrantes/imunologia , Ferimentos Penetrantes/terapiaRESUMO
BACKGROUND: Gaining hemostatic control of lethal vascular injuries sustained in combat using topical agents remains a challenge. Recent animal testing using a lethal arterial injury model has demonstrated that QuikClot zeolite granules (QCG) and the HemCon chitosan bandage (HC) are not capable of providing hemostasis and improving survival over the Army gauze field bandage (AFB). We have developed a new hemostatic agent consisting of a granular combination of a smectite mineral and a polymer (WoundStat) capable of producing hemostasis in the face of high-pressure arterial bleeding. We compared the performance of WoundStat (WS) to QCG, HC, AFB, and the new QuikClot zeolite Advance Clotting Sponge (ACS) in a lethal vascular injury model. METHODS: Hemostatic agents were tested using a lethal femoral artery vascular injury model. Twenty-five (5 per group) male swine (42 kg +/- 3 kg) were anesthetized, instrumented, and splenectomized. A lethal femoral artery injury was produced by creating a 6-mm arteriotomy in the vessel. After 45 seconds of hemorrhage, animals were randomized to be treated with AFB (control group), HC, QCG, ACS, or WS. Pressure (200 mm Hg) was applied over the product in the wound for 3 minutes. A second application and 3 additional minutes of pressure was provided if hemostasis was not achieved. Fluid resuscitation was begun at the time of application with 500 mL of Hextend, followed by lactated Ringer's solution at 100 mL/min to achieve and maintain a postapplication mean arterial blood pressure of 65 mm Hg. Animals were observed for 180 minutes or until death. Primary endpoints were survival, survival time, post-treatment blood loss, and amount of resuscitation fluid. RESULTS: All animals treated with WS survived to 180 minutes and required only a single application. No animal in the AFB, QCG, or ACS group survived. One animal in the HC group survived. Survival (p < 0.05) and survival times (p < 0.0001) for WS animals were significantly greater than for all other groups. No significant difference in survival or survival time existed between the AFB, QCG, ACS, or HC groups. Post-treatment blood loss (p = 0.0099) and postresuscitation fluid volume (p = 0.006) was significantly less for animals treated with WS than for all other groups. No significant difference in these parameters existed between the AFB, QCG, ACS, and HC groups. CONCLUSION: WS was superior to the other hemostatic agents tested in this study of lethal arterial vascular injury. Additional study is warranted on this agent to determine its potential for use in combat and civilian trauma.
Assuntos
Bandagens , Adesivo Tecidual de Fibrina/uso terapêutico , Hemorragia/terapia , Hemostáticos/uso terapêutico , Administração Tópica , Animais , Modelos Animais de Doenças , Artéria Femoral/lesões , Hidratação , Hemorragia/fisiopatologia , Masculino , Probabilidade , Distribuição Aleatória , Valores de Referência , Sensibilidade e Especificidade , Taxa de Sobrevida , SuínosRESUMO
BACKGROUND: Traumatic shock activates the hypothalamic-pituitary-adrenal axis (HPA) to mediate a cascade of defensive mechanisms that often include overwhelming inflammatory response and immunosuppression, which may lead to multiple organ failure. Androstenetriol (5 androstene, 3beta, 7beta, 17beta triol-AET) is a metabolite of dehydroepiandrosterone that markedly up regulates host immune response, prevents immune suppression, modulates inflammation and improves survival after lethal infections by pathogens and lethal radiation. HYPOTHESIS: AET-induced immune modulation will improve survival in a conscious rodent model of traumatic shock. METHODS: A relevant traumatic shock rodent model that applies to both combat and civilian sectors was used. After creation of a midline laparotomy (soft tissue trauma), animals were hemorrhaged to a mean arterial pressure of 35-40 mm Hg. Resuscitation was initiated sixty minutes later with crystalloid fluid and packed red blood cells and animals were observed for two days. In a randomized and blinded fashion, AET or vehicle was administered subcutaneously at the beginning of resuscitation. RESULTS: In the vehicle group 5 out of 16 animals survived, (31%). In contrast, 9 out of 13 animals who received AET survived (69%), (Fisher Exact Test p < 0.05). Survival in the AET treatment group was associated with reduced levels of IL-6, IL-10, and IL-18, and enhanced IFN-gamma and IL-2 levels. CONCLUSION: : The results indicate that AET provides a significant protective effect and improves survival in a clinically relevant model of traumatic hemorrhagic shock. AET protective effects are associated with an elevation of Th1 and reduction of Th2 cytokines.
Assuntos
Androstenóis/farmacologia , Fatores Imunológicos/farmacologia , Ressuscitação , Choque Traumático/tratamento farmacológico , Animais , Modelos Animais de Doenças , Interleucina-10/sangue , Interleucina-6/sangue , Interleucina-8/sangue , Masculino , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Choque Traumático/fisiopatologiaRESUMO
UNLABELLED: Trauma results in activation of the hypothalamic-pituitary-adrenal axis to mediate a cascade of neurohormonal changes as a defensive mechanism. Its prolongation, however, leads to a hypermetabolic, hypoperfused, and immunosuppressed state, setting the stage for subsequent sepsis and organ failure. Androstenetriol (5-androstene-3beta, 7beta, 17betatriol - AET), a metabolite of dehydroepiandrosterone, up-regulates the host immune response markedly, prevents immune suppression and controls inflammation, leading to improved survival after lethal infections by several diverse pathogens and lethal radiation. Such actions may be useful in improving survival from traumatic shock. HYPOTHESIS: The neurosteroid AET will increase survival following traumatic shock. METHODS: A combat relevant model of traumatic shock was used. Male Sprague-Dawley rats were anesthetized, catheterized and subjected to soft tissue injury (laparotomy). Animals were allowed to regain consciousness over the next 0.5 h and then bled 40% of their blood volume over 15 min. Forty-five minutes after the onset of hemorrhage animals were randomized to receive either a single subcutaneous dose of AET (40 mg/kg, sc) or vehicle (methylcellulose). Volume resuscitation consisted of l-lactated Ringer's (three times the shed blood volume), followed by packed red blood cells (one-third shed red cell volume). Animals were observed for three days. RESULTS: A total of 24 animals were studied. Of the 12 animals randomized to receive AET, all (100%) survived compared to 9 of 12 animals (75%) randomized to receive the vehicle (p < 0.05). CONCLUSION: AET significantly improved survival when administered subcutaneously in a single dose in this rodent model of traumatic shock. Further survival and mechanism studies are warranted.
Assuntos
Androstenóis/farmacologia , Fatores Imunológicos/farmacologia , Ressuscitação , Choque Traumático/tratamento farmacológico , Androstenóis/uso terapêutico , Animais , Quimiocina CCL2/sangue , Modelos Animais de Doenças , Fatores Imunológicos/uso terapêutico , Interleucina-1alfa/sangue , Masculino , Projetos Piloto , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Projetos de Pesquisa , Choque Traumático/sangue , Choque Traumático/fisiopatologia , Choque Traumático/terapiaRESUMO
PURPOSE OF REVIEW: Educate the reader of the multiple roles undertaken by the human epidermal lipidome and the experimental techniques of measuring them. RECENT FINDINGS: Damage to skin elicits a wound healing process that is capped by the recreation of the lipid barrier. In addition to barrier function, lipids also undertake an active signaling role during wound healing. Achievement of these multiple functions necessitates a significant complexity and diversity in the lipidome resulting in a composition that is unique to the human skin. As such, any attempts to delineate the function of the lipidome during the wound healing process in humans need to be addressed via studies undertaken in humans. SUMMARY: The human cutaneous lipidome is unique and play a functionally significant role in maintaining barrier and regulating wound healing. Modern mass spectrometry and Raman spectroscopy based methods enable the investigation epidermal lipidome with respect to those functions.
RESUMO
Acute wounds normally heal in a very orderly and efficient manner characterized by four distinct, but overlapping phases: hemostasis, inflammation, proliferation and remodeling. Specific biological markers characterize healing of acute wounds. Likewise, unique biologic markers also characterize pathologic responses resulting in fibrosis and chronic non-healing ulcers. This review describes the major biological processes associated with both normal and pathologic healing. The normal healing response begins the moment the tissue is injured. As the blood components spill into the site of injury, the platelets come into contact with exposed collagen and other elements of the extracellular matrix. This contact triggers the platelets to release clotting factors as well as essential growth factors and cytokines such as platelet-derived growth factor (PDGF) and transforming growth factor beta (TGF-beta). Following hemostasis, the neutrophils then enter the wound site and begin the critical task of phagocytosis to remove foreign materials, bacteria and damaged tissue. As part of this inflammatory phase, the macrophages appear and continue the process of phagocytosis as well as releasing more PDGF and TGF beta. Once the wound site is cleaned out, fibroblasts migrate in to begin the proliferative phase and deposit new extracellular matrix. The new collagen matrix then becomes cross-linked and organized during the final remodeling phase. In order for this efficient and highly controlled repair process to take place, there are numerous cell-signaling events that are required. In pathologic conditions such as non-healing pressure ulcers, this efficient and orderly process is lost and the ulcers are locked into a state of chronic inflammation characterized by abundant neutrophil infiltration with associated reactive oxygen species and destructive enzymes. Healing proceeds only after the inflammation is controlled. On the opposite end of the spectrum, fibrosis is characterized by excessive matrix deposition and reduced remodeling. Often fibrotic lesions are associated with increased densities of mast cells. By understanding the functional relationships of these biological processes of normal compared to abnormal wound healing, hopefully new strategies can be designed to treat the pathological conditions.
Assuntos
Fibrose/etiologia , Transdução de Sinais/fisiologia , Úlcera/etiologia , Cicatrização/fisiologia , Ferimentos e Lesões/fisiopatologia , Animais , Humanos , Ferimentos e Lesões/patologiaRESUMO
OBJECTIVE: The wound healing process is well-understood on the cellular and tissue level; however, its complex molecular mechanisms are not yet uncovered in their entirety. Viewing wounds as perturbed molecular networks provides the tools for analyzing and optimizing the healing process. It helps to answer specific questions that lead to better understanding of the complexity of the process. What are the molecular pathways involved in wound healing? How do these pathways interact with each other during the different stages of wound healing? Is it possible to grasp the entire mechanism of regulatory interactions in the healing of a wound? APPROACH: Networks are structures composed of nodes connected by links. A network describing the state of a cell taking part in the healing process may contain nodes representing genes, proteins, microRNAs, metabolites, and drug molecules. The links connecting nodes represent interactions such as binding, regulation, co-expression, chemical reaction, and others. Both nodes and links can be weighted by numbers related to molecular concentration and the intensity of intermolecular interactions. Proceeding from data and from molecular profiling experiments, different types of networks are built to characterize the stages of the healing process. Network nodes having a higher degree of connectivity and centrality usually play more important roles for the functioning of the system they describe. RESULTS: We describe here the algorithms and software packages for building, manipulating and analyzing networks proceeding from information available from a literature or database search or directly extracted from experimental gene expression, metabolic, and proteomic data. Network analysis identifies genes/proteins most differentiated during the healing process, and their organization in functional pathways or modules, and their distribution into gene ontology categories of biological processes, molecular functions, and cellular localization. We provide an example of how network analysis can be used to reach better understanding of regulation of key wound healing mediators and microRNAs that regulate them. INNOVATION: Univariate statistical tests widely used in clinical studies are not enough to improve understanding and optimize the processes of wound healing. Network methods of analysis of patients "omics" data, such as transcriptoms, proteomes, and others can provide a better insight into the healing processes and help in development of better treatment practices. We review several articles that are examples of this emergent approach to the study of wound healing. CONCLUSION: Network analysis has the potential to considerably contribute to the better understanding of the molecular mechanisms of wound healing and to the discovery of means to control and optimize that process.