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BACKGROUND: Ischemia-reperfusion injury is a common problem in liver surgery and transplantation. Although ischemia-reperfusion injury is known to be more pronounced in fatty livers, the underlying mechanisms for this difference remain poorly understood. We hypothesized that ferroptosis plays a significant role in fatty liver ischemia-reperfusion injury due to increased lipid peroxidation in the presence of stored iron in the fatty liver. To test this hypothesis, the ferroptosis pathway was evaluated in a murine fatty liver ischemia-reperfusion injury model. METHODS: C57BL6 mice were fed with a normal diet or a high fat, high sucrose diet for 12 weeks. At 22 weeks of age, liver ischemia-reperfusion injury was induced through partial (70%) hepatic pedicle clamping for 60 minutes, followed by 24 hours of reperfusion before tissue harvest. Acyl-coenzyme A synthetase long-chain family member 4 and 4-hydroxynonenal were quantified in the liver tissues. In separate experiments, liproxstatin-1 or vehicle control was administered for 7 consecutive days before liver ischemia-reperfusion injury. RESULTS: Exacerbated ischemia-reperfusion injury was observed in the livers of high fat, high sucrose diet fed mice. High fat, high sucrose diet + ischemia-reperfusion injury (HDF+IRI) livers had a significantly greater abundance of acyl-coenzyme A synthetase long-chain family member 4 and 4-hydroxynonenal compared with normal diet + ischemia-reperfusion injury (ND+IRI) livers or sham fatty livers, which indicated an increase of ferroptosis. HFD fed animals receiving liproxstatin-1 injections had a significant reduction in serum aspartate transaminase and alanine transaminase after ischemia-reperfusion injury, consistent with attenuation of ischemia-reperfusion injury in the liver. CONCLUSION: Ferroptosis plays a significant role in ischemia-reperfusion injury in fatty livers. Inhibiting ferroptotic pathways in the liver may serve as a novel therapeutic strategy to protect the fatty liver in the setting of ischemia-reperfusion injury.
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Ferroptosis , Peroxidación de Lípido , Hígado , Ratones Endogámicos C57BL , Daño por Reperfusión , Animales , Daño por Reperfusión/metabolismo , Daño por Reperfusión/etiología , Daño por Reperfusión/patología , Ratones , Masculino , Hígado/metabolismo , Hígado/irrigación sanguínea , Hígado/patología , Hígado Graso/metabolismo , Hígado Graso/etiología , Hígado Graso/patología , Modelos Animales de Enfermedad , Aldehídos/metabolismo , Coenzima A Ligasas/metabolismo , Dieta Alta en Grasa/efectos adversos , Quinoxalinas , Compuestos de EspiroRESUMEN
Diabetes mellitus (DM) is a chronic metabolic disorder recognized as a major health problem globally. A defective insulin activity contributes to the prevalence and expansion of DM. Treatment of DM is often hampered by limited options of conventional therapies and adverse effects associated with existing procedures. This has led to a spike in the exploration for potential therapeutic agents from various natural resources for clinical applications. The marine environment is a huge store of unexplored diversity of chemicals produced by a multitude of organisms. To date, marine microorganisms, microalgae, macroalgae, corals, sponges, and fishes have been evaluated for their anti-diabetic properties. The structural diversity of bioactive metabolites discovered has shown promising hypoglycaemic potential through in vitro and in vivo screenings via various mechanisms of action, such as PTP1B, α-glucosidase, α-amylase, ß-glucosidase, and aldose reductase inhibition as well as PPAR alpha/gamma dual agonists activities. On the other hand, hypoglycaemic effect is also shown to be exerted through the balance of antioxidants and free radicals. This review highlights marine-derived chemicals with hypoglycaemic effects and their respective mechanisms of action in the management of DM in humans.
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The increasing prevalence of allergic diseases is of great public health concern. Environmental and food allergens are the major triggers of allergic diseases via respiratory or gastrointestinal routes, respectively. A major setback in the clinical management of allergies is the unavailability of purified allergens required for diagnostic purposes. Furthermore, manipulation of allergen sequences and structures by employing protein-engineering approaches is needed to design immunotherapeutic vaccines. All these approaches rely upon the sequence, structure, and epitope location of allergens. A number of databases have therefore been developed that serve as repositories of molecular information of allergens. In this chapter, we discuss the five most important widely used allergen databases that might be helpful for the research community working on molecular allergology.
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Alérgenos , Hipersensibilidad a los Alimentos , Humanos , Epítopos , Bases de Datos FactualesRESUMEN
In this chapter, the steps of designing candidate vaccine molecules for allergen-specific immunotherapy (AIT) using immunoinformatics are described. The most modern approach of AIT deals with carrier-bound B cell epitope and multi-epitope vaccine molecules. The strategy for designing these molecules and the bioinformatics tools and servers used for that are discussed in detail here.
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Alérgenos , Vacunas , Desensibilización Inmunológica , Epítopos de Linfocito B , Biología Computacional , Epítopos de Linfocito TRESUMEN
Repair of epithelial defect is complicated by infection and related metabolites. Pyocyanin (PYO) is one such metabolite that is secreted during Pseudomonas aeruginosa infection. Keratinocyte (KC) migration is required for the closure of skin epithelial defects. This work sought to understand PYO-KC interaction and its significance in tissue repair. Stable Isotope Labeling by Amino acids in Cell culture proteomics identified mitochondrial dysfunction as the top pathway responsive to PYO exposure in human KCs. Consistently, functional studies showed mitochondrial stress, depletion of reducing equivalents, and adenosine triphosphate. Strikingly, despite all stated earlier, PYO markedly accelerated KC migration. Investigation of underlying mechanisms revealed, to our knowledge, a previously unreported function of keratin 6A in KCs. Keratin 6A was PYO inducible and accelerated closure of epithelial defect. Acceleration of closure was associated with poor quality healing, including compromised expression of apical junction proteins. This work recognizes keratin 6A for its role in enhancing KC migration under conditions of threat posed by PYO. Qualitatively deficient junctional proteins under conditions of defensive acceleration of KC migration explain why an infected wound close with deficient skin barrier function as previously reported.
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Queratina-6 , Piocianina , Humanos , Piocianina/química , Piocianina/metabolismo , Queratina-6/metabolismo , Piel/metabolismo , Mitocondrias/metabolismoRESUMEN
Tissue injury to skin diminishes miR-200b in dermal fibroblasts. Fibroblasts are widely reported to directly reprogram into endothelial-like cells and we hypothesized that miR-200b inhibition may cause such changes. We transfected human dermal fibroblasts with anti-miR-200b oligonucleotide, then using single cell RNA sequencing, identified emergence of a vasculogenic subset with a distinct fibroblast transcriptome and demonstrated blood vessel forming function in vivo. Anti-miR-200b delivery to murine injury sites likewise enhanced tissue perfusion, wound closure, and vasculogenic fibroblast contribution to perfused vessels in a FLI1 dependent manner. Vasculogenic fibroblast subset emergence was blunted in delayed healing wounds of diabetic animals but, topical tissue nanotransfection of a single anti-miR-200b oligonucleotide was sufficient to restore FLI1 expression, vasculogenic fibroblast emergence, tissue perfusion, and wound healing. Augmenting a physiologic tissue injury adaptive response mechanism that produces a vasculogenic fibroblast state change opens new avenues for therapeutic tissue vascularization of ischemic wounds.
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Fibroblastos , Piel , Cicatrización de Heridas , Animales , Humanos , Ratones , Antagomirs/farmacología , Antagomirs/uso terapéutico , Fibroblastos/metabolismo , Fibroblastos/fisiología , Oligonucleótidos/farmacología , Piel/metabolismo , Cicatrización de Heridas/genética , Cicatrización de Heridas/fisiologíaRESUMEN
Fetal cutaneous wound closure and repair differ from that in adulthood. In this work, we identify an oxidant stress sensor protein, nonselenocysteine-containing phospholipid hydroperoxide glutathione peroxidase (NPGPx), that is abundantly expressed in normal fetal epidermis (and required for fetal wound closure), though not in adult epidermis, but is variably re-induced upon adult tissue wounding. NPGPx is a direct target of the miR-29 family. Following injury, abundance of miR-29 is lowered, permitting a prompt increase in NPGPx transcripts and protein expression in adult wound-edge tissue. NPGPx expression was required to mediate increased keratinocyte migration induced by miR-29 inhibition in vitro and in vivo. Increased NPGPx expression induced increased SOX2 expression and ß-catenin nuclear localization in keratinocytes. Augmenting physiologic NPGPx expression via experimentally induced miR-29 suppression, using cutaneous tissue nanotransfection or targeted lipid nanoparticle delivery of anti-sense oligonucleotides, proved to be sufficient to overcome the deleterious effects of diabetes on this specific pathway to enhance tissue repair.
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MicroARNs , Cicatrización de Heridas , Embarazo , Humanos , Femenino , Cicatrización de Heridas/genética , Piel/metabolismo , Queratinocitos/metabolismo , Movimiento Celular , MicroARNs/metabolismoRESUMEN
OBJECTIVE: This work addressing complexities in wound infection, seeks to test the reliance of bacterial pathogen Pseudomonas aeruginosa (PA) on host skin lipids to form biofilm with pathological consequences. BACKGROUND: PA biofilm causes wound chronicity. Both CDC as well as NIH recognizes biofilm infection as a threat leading to wound chronicity. Chronic wounds on lower extremities often lead to surgical limb amputation. METHODS: An established preclinical porcine chronic wound biofilm model, infected with PA or Pseudomonas aeruginosa ceramidase mutant (PA ∆Cer ), was used. RESULTS: We observed that bacteria drew resource from host lipids to induce PA ceramidase expression by three orders of magnitude. PA utilized product of host ceramide catabolism to augment transcription of PA ceramidase. Biofilm formation was more robust in PA compared to PA ∆Cer . Downstream products of such metabolism such as sphingosine and sphingosine-1-phosphate were both directly implicated in the induction of ceramidase and inhibition of peroxisome proliferator-activated receptor (PPAR)δ, respectively. PA biofilm, in a ceram-idastin-sensitive manner, also silenced PPARδ via induction of miR-106b. Low PPARδ limited ABCA12 expression resulting in disruption of skin lipid homeostasis. Barrier function of the wound-site was thus compromised. CONCLUSIONS: This work demonstrates that microbial pathogens must co-opt host skin lipids to unleash biofilm pathogenicity. Anti-biofilm strategies must not necessarily always target the microbe and targeting host lipids at risk of infection could be productive. This work may be viewed as a first step, laying fundamental mechanistic groundwork, toward a paradigm change in biofilm management.
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PPAR delta , Pseudomonas aeruginosa , Animales , Ceramidasas , Extremidad Inferior , PorcinosRESUMEN
Diabetes is an endocrinological disorder with a rapidly increasing number of patients globally. Over the last few years, the alarming status of diabetes has become a pivotal factor pertaining to morbidity and mortality among the youth as well as middle-aged people. Current developments in our understanding related to autoimmune responses leading to diabetes have developed a cause for concern in the prospective usage of immunomodulatory agents to prevent diabetes. The mechanism of action of vaccines varies greatly, such as removing autoreactive T cells and inhibiting the interactions between immune cells. Currently, most developed diabetes vaccines have been tested in animal models, while only a few human trials have been completed with positive outcomes. In this review, we investigate the undergoing clinical trial studies for the development of a prototype diabetes vaccine.
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Diabetes Mellitus Tipo 2 , Vacunas , Adolescente , Animales , Autoinmunidad , Diabetes Mellitus Tipo 2/prevención & control , Humanos , Persona de Mediana Edad , Estudios Prospectivos , Linfocitos T , Vacunas/uso terapéuticoRESUMEN
Next-generation allergy vaccines refer to allergen-derived attenuated molecules that can boost allergen-blocking IgG response. These IgG antibodies are specifically directed toward the IgE epitope of allergens and interfere in allergen-IgE interaction. Our study is a computational approach to design such vaccines against four widespread pan-allergens families. Pan-allergens display extensive immunological cross-reactivity due to the presence of conserved IgE epitope and T cell epitope. In this study, the vaccine design is based on hapten-carrier concept in which the carrier protein is an immunogenic component providing T cell help. Either PreS protein of hepatitis B or cholera enterotoxin B (CTB) fused with three tetanus toxoid fragments (TTFrC) was used here as the carrier. The hapten components are nonanaphylactic peptides (NAPs) derived from experimentally determined antigenic regions of the allergens. The charged residues of NAPs are selectively modified to obliterate IgE, as well as T cell reaction, and hence, are safe to apply in allergy patients. Various combinations of vaccine constructs (PreS/CTB+TTFrC and NAPs) were designed with intermediate linker motifs. Screening of constructs was performed through a three-step method such as physicochemical parameters, secondary structures, and tertiary structures using various bioinformatic tools. The final construct with best quality and stability was selected for each allergen family. Suitability of these constructs for being expressed in recombinant form was checked at DNA, RNA, and protein level. Presence of putative epitopes inducing tolerogenic interleukin-10 was also predicted for these constructs. The present work led to the design of putative vaccines with immunotherapeutic potential and broad applicability for allergic diseases caused by a wide array of cross-reactive allergens.
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Hipersensibilidad , Vacunas , Humanos , Alérgenos/genética , Alérgenos/química , Anticuerpos Monoclonales , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/química , Haptenos , Inmunoglobulina E/metabolismo , Inmunoglobulina G , PéptidosRESUMEN
Chronic wounds infected by Pseudomonas aeruginosa (Pa) are characterized by disease progression and increased mortality. We reveal Pf, a bacteriophage produced by Pa that delays healing of chronically infected wounds in human subjects and animal models of disease. Interestingly, impairment of wound closure by Pf is independent of its effects on Pa pathogenesis. Rather, Pf impedes keratinocyte migration, which is essential for wound healing, through direct inhibition of CXCL1 signaling. In support of these findings, a prospective cohort study of 36 human patients with chronic Pa wound infections reveals that wounds infected with Pf-positive strains of Pa are more likely to progress in size compared with wounds infected with Pf-negative strains. Together, these data implicate Pf phage in the delayed wound healing associated with Pa infection through direct manipulation of mammalian cells. These findings suggest Pf may have potential as a biomarker and therapeutic target in chronic wounds.
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Inovirus , Infecciones por Pseudomonas , Infección de Heridas , Animales , Biopelículas , Humanos , Mamíferos , Estudios Prospectivos , Pseudomonas , Infecciones por Pseudomonas/terapia , Pseudomonas aeruginosa , Cicatrización de Heridas , Infección de Heridas/terapiaRESUMEN
SCOPE: Reactive oxygen species production by innate immune cells plays a central role in host defense against invading pathogens at wound-site. A weakened host-defense results in persistent infection leading to wound chronicity. Fermented Papaya Preparation (FPP), a complex sugar matrix, bolsters respiratory burst activity and improves wound healing outcomes in chronic wound patients. The objective of the current study was to identify underlying molecular factor/s responsible for augmenting macrophage host defense mechanisms following FPP supplementation. METHODS AND RESULTS: In depth LC-MS/MS analysis of cells supplemented with FPP led to identification of myo-inositol as a key determinant of FPP activity towards improving macrophage function. Myo-inositol, in quantities that is present in FPP, significantly improved macrophage respiratory burst and phagocytosis via de novo synthesis pathway of ISYNA1. In addition, myo-inositol transporters, HMIT and SMIT1, played a significant role in such activity. Blocking these pathways using siRNA attenuated FPP-induced improved macrophage host defense activities. FPP supplementation emerged as a novel approach to increase intracellular myo-inositol levels. Such supplementation also modified wound microenvironment in chronic wound patients to augment myo-inositol levels in wound fluid. CONCLUSION: These observations indicate that myo-inositol in FPP influences multiple aspects of macrophage function critical for host defense against invading pathogens.
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Azúcares , Espectrometría de Masas en Tándem , Cromatografía Liquida , Humanos , Inositol/farmacología , Macrófagos/metabolismoRESUMEN
Impaired re-epithelialization characterized by hyperkeratotic nonmigratory wound epithelium is a hallmark of nonhealing diabetic wounds. In chronic wounds, the copious release of oncostatin M (OSM) from wound macrophages is evident. OSM is a potent keratinocyte (KC) activator. This work sought to understand the signal transduction pathway responsible for wound re-epithelialization, the primary mechanism underlying wound closure. Daily topical treatment of full-thickness excisional wounds of C57BL/6 mice with recombinant murine OSM improved wound re-epithelialization and accelerated wound closure by bolstering KC proliferation and migration. OSM activated the Jak-signal transducer and activator of transcription pathway as manifested by signal transducer and activator of transcription 3 phosphorylation. Such signal transduction in the human KC induced TP63, the master regulator of KC function. Elevated TP63 induced ITGB1, a known effector of KC migration. In diabetic wounds, OSM was more abundant than the level in nondiabetic wounds. However, in diabetic wounds, OSM activity was compromised by glycation. Aminoguanidine, a deglycation agent, rescued the compromised KC migration caused by glycated OSM. Finally, topical application of recombinant OSM improved KC migration and accelerated wound closure in db/db mice. This work recognizes that despite its abundance at the wound site, OSM is inactivated by glycation, and topical delivery of exogenous OSM is likely to be productive in accelerating diabetic wound closure.
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Diabetes Mellitus , Repitelización , Animales , Ratones , Ratones Endogámicos C57BL , Oncostatina M , Cicatrización de Heridas/fisiologíaRESUMEN
Healing of cutaneous wounds requires the collective migration of epithelial keratinocytes to seal the wound bed from the environment. However, the signaling events that coordinate this collective migration are unclear. In this report, we address the role of phosphorylation of eukaryotic initiation factor 2 (eIF2) and attendant gene expression during wound healing. Wounding of human keratinocyte monolayers in vitro led to the rapid activation of the eIF2 kinase GCN2. We determined that deletion or pharmacological inhibition of GCN2 significantly delayed collective cell migration and wound closure. Global transcriptomic, biochemical, and cellular analyses indicated that GCN2 is necessary for maintenance of intracellular free amino acids, particularly cysteine, as well as coordination of RAC1-GTP-driven reactive oxygen species (ROS) generation, lamellipodia formation, and focal adhesion dynamics following keratinocyte wounding. In vivo experiments using mice deficient for GCN2 validated the role of the eIF2 kinase during wound healing in intact skin. These results indicate that GCN2 is critical for appropriate induction of collective cell migration and plays a critical role in coordinating the re-epithelialization of cutaneous wounds.
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Movimiento Celular , Queratinocitos/enzimología , Proteínas Serina-Treonina Quinasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Cicatrización de Heridas , Aminoácidos/metabolismo , Animales , Línea Celular Transformada , Adhesiones Focales/genética , Adhesiones Focales/metabolismo , Humanos , Queratinocitos/patología , Ratones , Ratones Noqueados , Proteínas Serina-Treonina Quinasas/genética , Seudópodos/genética , Seudópodos/metabolismo , Piel/enzimología , Piel/lesiones , Piel/patologíaRESUMEN
BACKGROUND: Plantation of road-side avenue trees has become a major part of urbanization programme for megacity beautification and environmental management. Due to evergreen habit and vibrant flower colour, Delonix regia (Gulmohor/Flamboyant) is frequently selected as ornamental tree for plantation programme. However, D. regia pollen is related to IgE mediated allergic reactions and no allergen has been reported so far. OBJECTIVE: Measuring the prevalence of D. regia pollen sensitivity among the local atopic individuals and identifying the allergens using immunoproteomic tools. METHODS: Aerobiological study was conducted for a period of two years to record the D. regia pollen concentration in the outdoor ambient air. Clinico-immunological tests were performed on atopic individuals to check the prevalence of sensitivity against D. regia pollen. Allergens were detected in the pollen proteome, fractionated in 1D and 2D gel by IgE serology and finally identified by mass spectrometry. RESULT: In the study area D. regia pollen grains were present in ample amount in the air during May to July. About 38% of atopic individuals displayed positive Skin Prick Test (SPT) against D. regia pollen along with elevated level of specific IgE and histamine in the sera. Immunoproteomic analyses revealed the presence of 14 IgE reactive proteins in the 2D pollen proteome, of which 8 IgE reactive proteins were identified by MALDI TOF/TOF using homology driven proteomic approach. CONCLUSION: This study demonstrated pollen related allergy symptoms by D. regia and gave significant message regarding the plantation programme to avoid the unnecessary load of allergic pollen. Also, a panel of 8 allergens were identified for the first time from D. regia pollen. Detailed study of these allergens would help to design immunotherapeutic strategies for pollinosis management.
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BACKGROUND: Currently, the novel coronavirus or COVID-19 pandemic poses the greatest global health threat worldwide, and India is no exception. As an overpopulated developing country, it is very difficult to maintain social distancing to restrict the spread of the disease in India. Under these circumstances, it is necessary to examine India's interstate performances to combat COVID-19. This study aims to explore twin objectives: to investigate the comparative efficiency of Indian states to combat COVID-19 and to unfold the factors responsible for interstate disparities in the efficiency in combatting COVID-19. METHODS: The stochastic production frontier model was utilized for data analysis. The empirical analysis was facilitated by the inefficiency effects model, revealing the factors that influence interstate variability in disease management efficiency. Three types of variables, namely, output, inputs, and exogenous, were used to measure health system efficiency. The relevant variables were compiled from secondary sources. The recovery rate from COVID-19 was the output variable and health infrastructures were considered as the input variable. On the contrary, the non-health determinants considered to have a strong influence on the efficiency of states' disease management, but could not be considered as input variables, were recognised as exogenous variables. These exogenous variables were specifically used for the inefficiency analysis. RESULTS: The empirical results demonstrated the existence of disparities across Indian states in the level of efficiency in combatting COVID-19. A non-trivial outcome of this study was that Tamil Nadu was the best performer and Manipur was the worst performer of the investigated states. Variables such as elderly people, sex ratio, literacy rate, population density, influenced the efficiency of states, and thus, affected the recovery rate. CONCLUSION: This study argues for the efficient utilisation of the existing health infrastructures in India. Simultaneously, the study suggests improving the health infrastructure to achieve a long-run benefit.
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COVID-19/epidemiología , Control de Enfermedades Transmisibles/estadística & datos numéricos , Utilización de Instalaciones y Servicios/estadística & datos numéricos , Disparidades en Atención de Salud/estadística & datos numéricos , COVID-19/prevención & control , Geografía , Humanos , India/epidemiología , Distanciamiento Físico , Densidad de Población , SARS-CoV-2 , Procesos EstocásticosRESUMEN
Urolithin A (UA) is a natural compound that is known to improve muscle function. In this work we sought to evaluate the effect of UA on muscle angiogenesis and identify the underlying molecular mechanisms. C57BL/6 mice were administered with UA (10 mg/body weight) for 12-16 weeks. ATP levels and NAD+ levels were measured using in vivo 31P NMR and HPLC, respectively. UA significantly increased ATP and NAD+ levels in mice skeletal muscle. Unbiased transcriptomics analysis followed by Ingenuity Pathway Analysis (IPA) revealed upregulation of angiogenic pathways upon UA supplementation in murine muscle. The expression of the differentially regulated genes were validated using quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC). Angiogenic markers such as VEGFA and CDH5 which were blunted in skeletal muscles of 28 week old mice were found to be upregulated upon UA supplementation. Such augmentation of skeletal muscle vascularization was found to be bolstered via Silent information regulator 1 (SIRT1) and peroxisome proliferator-activated receptor-gamma coactivator-1-alpha (PGC-1α) pathway. Inhibition of SIRT1 by selisistat EX527 blunted UA-induced angiogenic markers in C2C12 cells. Thus this work provides maiden evidence demonstrating that UA supplementation bolsters skeletal muscle ATP and NAD+ levels causing upregulated angiogenic pathways via a SIRT1-PGC-1α pathway.
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Cumarinas/farmacología , Músculo Esquelético/efectos de los fármacos , NAD/metabolismo , Sirtuina 1/metabolismo , Adenosina Trifosfato/metabolismo , Administración Oral , Animales , Cumarinas/administración & dosificación , Perfilación de la Expresión Génica , Masculino , Ratones Endogámicos C57BL , Músculo Esquelético/irrigación sanguínea , Músculo Esquelético/metabolismo , Neovascularización Fisiológica/efectos de los fármacos , Neovascularización Fisiológica/genética , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/metabolismo , Reproducibilidad de los ResultadosRESUMEN
Sunflower pollen was reported to contain respiratory allergens responsible for occupational allergy and pollinosis. The present study describes the comprehensive characterization of a major sunflower allergen Hel a 6. Natural Hel a 6 was purified from sunflower pollen by anion exchange and gel filtration chromatography. Hel a 6 reacted with IgE-antibodies from 57% of 39 sunflower-sensitized patient sera suggesting it to be a major allergen. The patients were of Indian origin and suffering from pollinosis and allergic rhinitis. Hel a 6 exhibited allergenic activity by stimulating mediator release from basophils. Monomeric Hel a 6 displayed pectate lyase activity. The effect of various physicochemical parameters such as temperature, pH, and calcium ion on the functional activity of Hel a 6 revealed a stable nature of the protein. Hel a 6 was folded, and its melting curve showed reversible denaturation in which it refolded back to its native conformation from a denatured state. Hel a 6 displayed a high degree of sequence conservation with the pectate lyase allergens from related taxonomic families such as Amb a 1 (67%) and Art v 6 (57%). The IgE-cross reactivity was observed between Hel a 6 and its ragweed and mugwort homologs. The cross-reactivity was further substantiated by the mediator release when Hel a 6-sensitized effector cells were cross-stimulated with Art v 6 and Amb a 1. Several putative B cell epitopes were predicted and mapped on these 3 allergens. Two antigenic regions were found to be commonly shared by these 3 allergens, which could be crucial for cross-reactivity. In conclusion, Hel a 6 serves as a candidate molecule for diagnosis and immunotherapy for weed allergy.
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Alérgenos/química , Alérgenos/inmunología , Helianthus/química , Hipersensibilidad/inmunología , Polisacárido Liasas/inmunología , Alérgenos/aislamiento & purificación , Alérgenos/metabolismo , Ambrosia/inmunología , Dicroismo Circular , Reacciones Cruzadas , Epítopos/inmunología , Granjas , Helianthus/inmunología , Histamina/metabolismo , Humanos , Concentración de Iones de Hidrógeno , Sueros Inmunes , Espectrometría de Masas , Proteínas de Plantas/química , Proteínas de Plantas/inmunología , Proteínas de Plantas/aislamiento & purificación , Polen/enzimología , Polen/inmunología , Polisacárido Liasas/química , Polisacárido Liasas/aislamiento & purificación , Polisacárido Liasas/metabolismo , Pliegue de Proteína , Pruebas Cutáneas , TemperaturaRESUMEN
OBJECTIVE: The objective of this work was to causatively link biofilm properties of bacterial infection to specific pathogenic mechanisms in wound healing. BACKGROUND: Staphylococcus aureus is one of the four most prevalent bacterial species identified in chronic wounds. Causatively linking wound pathology to biofilm properties of bacterial infection is challenging. Thus, isogenic mutant stains of S. aureus with varying degree of biofilm formation ability was studied in an established preclinical porcine model of wound biofilm infection. METHODS: Isogenic mutant strains of S. aureus with varying degree (ΔrexBâ>âUSA300â>âΔsarA) of biofilm-forming ability were used to infect full-thickness porcine cutaneous wounds. RESULTS: Compared with that of ΔsarA infection, wound biofilm burden was significantly higher in response to ΔrexB or USA300 infection. Biofilm infection caused degradation of cutaneous collagen, specifically collagen 1 (Col1), with ΔrexB being most pathogenic in that regard. Biofilm infection of the wound repressed wound-edge miR-143 causing upregulation of its downstream target gene matrix metalloproteinase-2. Pathogenic rise of collagenolytic matrix metalloproteinase-2 in biofilm-infected wound-edge tissue sharply decreased collagen 1/collagen 3 ratio compromising the biomechanical properties of the repaired skin. Tensile strength of the biofilm infected skin was compromised supporting the notion that healed wounds with a history of biofilm infection are likely to recur. CONCLUSION: This study provides maiden evidence that chronic S. aureus biofilm infection in wounds results in impaired granulation tissue collagen leading to compromised wound tissue biomechanics. Clinically, such compromise in tissue repair is likely to increase wound recidivism.
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Biopelículas , Colágeno/metabolismo , Tejido de Granulación/metabolismo , Staphylococcus aureus/aislamiento & purificación , Cicatrización de Heridas/fisiología , Infección de Heridas/microbiología , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Tejido de Granulación/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Infecciones Estafilocócicas/microbiología , Porcinos , Infección de Heridas/diagnósticoRESUMEN
Collagen containing wound-care dressings are extensively used. However, the mechanism of action of these dressings remain unclear. Earlier studies utilizing a modified collagen gel (MCG) dressing demonstrated improved vascularization of ischemic wounds and better healing outcomes. Wound macrophages are pivotal in facilitating wound angiogenesis and timely healing. The current study was designed to investigate the effect of MCG on wound macrophage phenotype and function. MCG augmented recruitment of macrophage at the wound-site, attenuated pro-inflammatory and promoted anti-inflammatory macrophage polarization. Additionally, MCG increased anti-inflammatory IL-10, IL-4 and pro-angiogenic VEGF production, indicating a direct role of MCG in resolving wound inflammation and improving angiogenesis. At the wound-site, impairment in clearance of apoptotic cell bioburden enables chronic inflammation. Engulfment of apoptotic cells by macrophages (efferocytosis) resolves inflammation via a miR-21-PDCD4-IL-10 pathway. MCG-treated wound macrophages exhibited a significantly bolstered efferocytosis index. Such favorable outcome significantly induced miR-21 expression. MCG-mediated IL-10 production was dampened under conditions of miR-21 knockdown pointing towards miR-21 as a causative factor. Pharmacological inhibition of JNK attenuated IL-10 production by MCG, implicating miR-21-JNK pathway in MCG-mediated IL-10 production by macrophages. This work provides direct evidence demonstrating that a collagen-based wound-care dressing may influence wound macrophage function and therefore modify wound inflammation outcomes.