RESUMEN

There is a great effort from numerous research groups in the development of materials and therapeutic strategies for the functional recovery of patients who have suffered peripheral nerve injuries (PNI). In an article in vivo, the formation of a nerve bridge was observed, reconnecting the distal and proximal stumps, in the sciatic nerve of rats, indicating the effective participation of the biomaterial in the recovery of peripheral nerve injuries. For the current pilot study, 15 cases of multiple fractures of the mandible, with involvement of the inferior alveolar nerve (IAN) were selected and studied: JC (control cases) n = 6 with conventional treatment, and JT (treated cases) n = 9, with the use of biomimetic biomaterial. The evaluation of the return to sensitivity was measured through a self-assessment, where the patients assigned scores from 0 to 10, where zero (0) represented the complete absence of sensitivity and ten (10) the normality of the perception of local sensitivity. Patients were evaluated from the preoperative period to the 360th day. The statistical results obtained by the t-Student, Shapiro-Wilk normality and non-parametric One-Way ANOVA tests indicated statistically significant differences (p < 0.005; 0.005 e 0.5 respectively), between the two treatments, which were reflected in the clinical results observed, we also calculate the size of the effect represented by ϵ2, calculated by Cohen's d. The results indicate a great difference between the treatments performed,ϵ2 = 1.00. In the 6 cases followed up in the JC group, four remained with a significant deficit until the end of the evaluations and two indicated the remission of the lack of sensitivity in this period. In the JT group, in 28 days, all cases indicated complete remission of the lack of sensitivity with healing concentration. In one of the cases where there was a complete rupture of the mental nerve, the (score-10) was observed in 60 days. The observed results indicate the existence of a statistical significance between the groups and an important relationship when using the biomimetic biomaterial during the recovery of the perception of sensitivity in polytraumatized patients, compatible with the results observed in laboratory animals, which may indicate its clinical feasibility in the reduction of sequelae in PNI.

RESUMEN

NADH and NADPH are labile coenzymes that undergo hydration by enzymatic reaction or by heat at 5,6 double bond, and convert into non-functional hydrates, NADHX and NADPHX, respectively. The NAD(P)H hydrate dehydratase enzyme catalyzes the dehydration of S-NADHX/S-NADPHX at the expense of ATP, and thus contributes in the nicotinamide nucleotide repair process. This enzyme is also known as "metabolite-proofreading enzyme". Herein, we report the molecular cloning and expression of this highly conserved enzyme of vancomycin-resistant Staphylococcus aureus (VRSA). Its functional and inhibition studies were performed for the first time by NMR spectroscopy. NMR studies showed the dehydration of S epimer of NADHX, in the presence of R-NADHX and cyc-NADHX, by NAD(P)H hydrate dehydratase. In addition, by employing the STD-NMR approach, a library of drugs and natural products (total 79) were evaluated for their binding interactions with the NAD(P)H hydrate dehydratase enzyme. Among them, seven compounds showed ligand-like interactions with the enzyme, and thus functional activity of the enzyme was again checked in the presence of each ligand. Compound 2 (Thiamine HCl) was found to fully inhibit the enzyme's function, and recognized as a potential inhibitor. Current study demonstrates that this enzyme deserves further studies as a potential drug target, as its inhibition can disrupt the normal metabolism of pathogenic VRSA.

Asunto(s)

Productos Biológicos , Staphylococcus aureus Resistente a Meticilina , Adenosina Trifosfato , Deshidratación , Humanos , Hidroliasas/química , Ligandos , Espectroscopía de Resonancia Magnética , NAD/metabolismo , NADP/metabolismo , Niacinamida , Tiamina , Staphylococcus aureus Resistente a Vancomicina

RESUMEN

Emerging evidence indicates that the accretion of senescent cells is linked to metabolic disorders. However, the underlying mechanisms and metabolic consequences of cellular senescence in obesity remain obscure. In this study, we found that obese adipocytes are senescence-susceptible cells accompanied with genome instability. Additionally, we discovered that SREBP1c may play a key role in genome stability and senescence in adipocytes by modulating DNA-damage responses. Unexpectedly, SREBP1c interacted with PARP1 and potentiated PARP1 activity during DNA repair, independent of its canonical lipogenic function. The genetic depletion of SREBP1c accelerated adipocyte senescence, leading to immune cell recruitment into obese adipose tissue. These deleterious effects provoked unhealthy adipose tissue remodeling and insulin resistance in obesity. In contrast, the elimination of senescent adipocytes alleviated adipose tissue inflammation and improved insulin resistance. These findings revealed distinctive roles of SREBP1c-PARP1 axis in the regulation of adipocyte senescence and will help decipher the metabolic significance of senescence in obesity.

Asunto(s)

Resistencia a la Insulina , Adipocitos/metabolismo , Tejido Adiposo/metabolismo , Humanos , Resistencia a la Insulina/fisiología , Obesidad/metabolismo , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/genética , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo

RESUMEN

In this paper, the Inconel 625 laser clads characterized by microstructural homogeneity due to the application of the Laser Engineered Net Shaping (LENS, Optomec, Albuquerque, NM, USA) technology were studied in detail. The optimized LENS process parameters (laser power of 550 W, powder flow rate of 19.9 g/min, and heating of the substrate to 300 °C) enabled to deposit defect-free laser cladding. Additionally, the laser clad was applied in at least three layers on the repairing place. The deposited laser clads were characterized by slightly higher mechanical properties in comparison to the Inconel 625 substrate material. Microscopic observations and X-ray Tomography (XRT, Nikon Corporation, Tokyo, Japan) confirmed, that the substrate and cladding interface zone exhibited a defect-free structure. Mechanical properties and flexural strength of the laser cladding were examined using microhardness and three-point bending tests. It was concluded, that the LENS technology could be successfully applied for the repair since a similar strain distribution was found after Digital Image Correlation measurements during three-point bending tests.

RESUMEN

The effects of environmental stresses on microorganisms have been well-studied, and cellular responses to stresses such as heat, cold, acids, and salts have been extensively discussed. Although high pressure processing (HPP) is becoming more popular as a preservation method in the food industry, the characteristics of the cellular damage caused by high pressure are unclear, and the microbial response to this stress has not yet been well-explored. We exposed the pathogen Listeria monocytogenes to HPP (400 MPa, 8 min, 8°C) and found that the high pressure created plasma membrane pores. Using a common staining technique involving propidium iodide (PI) combined with high-frequency fluorescence microscopy, we monitored the rate of diffusion of PI molecules into hundreds of bacterial cells through these pores on days 0, 1, 2, 3, and 4 after pressurization. We also developed a mathematical dynamic model based on mass transfer and passive diffusion laws, calibrated using our microscopy experiments, to evaluate the response of bacteria to HPP. We found that the rate of diffusion of PI into the cells decreased over the 4 consecutive days after exposure to HPP, indicating repair of the pressure-created membrane pores. The model suggested a temporal change in the size of pores until closure. To the best of our knowledge, this is the first time that pressure-created membrane pores have been quantitatively described and shown to diminish with time. In addition, we found that the membrane repair rate in response to HPP was linear, and growth was temporarily arrested at the population level during the repair period. These results support the existence of a progressive repair process in some of the cells that take up PI, which can therefore be considered as being sub-lethally injured rather than dead. Hence, we showed that a subgroup of bacteria survived HPP and actively repaired their membrane pores.

RESUMEN

The influence of the repair process on the electrical properties of the normally off p-GaN high-electron-mobility transistor (HEMT) is studied in detail in this paper. We find that the etching process will cause the two-dimensional electron gas (2DEG) and the mobility of the p-GaN HEMT to decrease. However, the repair process will gradually recover the electrical properties. We study different repair methods and different repair conditions, propose the best repair conditions, and further fabricate the p-GaN HEMTs devices. The threshold voltage of the fabricated device is 1.6 V, the maximum gate voltage is 7 V, and the on-resistance is 23 Ω·mm. The device has a good performance, which proves that the repair conditions can be successfully applied to the fabricate of the p-GaN HEMT devices.

RESUMEN

Human apurinic/apyrimidinic endonuclease 1 (APE1) is known to be a critical player of the base excision repair (BER) pathway. In general, BER involves consecutive actions of DNA glycosylases, AP endonucleases, DNA polymerases, and DNA ligases. It is known that these proteins interact with APE1 either at upstream or downstream steps of BER. Therefore, we may propose that even a minor disturbance of protein-protein interactions on the DNA template reduces coordination and repair efficiency. Here, the ability of various human DNA repair enzymes (such as DNA glycosylases OGG1, UNG2, and AAG; DNA polymerase Polß; or accessory proteins XRCC1 and PCNA) to influence the activity of wild-type (WT) APE1 and its seven natural polymorphic variants (R221C, N222H, R237A, G241R, M270T, R274Q, and P311S) was tested. Förster resonance energy transfer-based kinetic analysis of abasic site cleavage in a model DNA substrate was conducted to detect the effects of interacting proteins on the activity of WT APE1 and its single-nucleotide polymorphism (SNP) variants. The results revealed that WT APE1 activity was stimulated by almost all tested DNA repair proteins. For the SNP variants, the matters were more complicated. Analysis of two SNP variants, R237A and G241R, suggested that a positive charge in this area of the APE1 surface impairs the protein-protein interactions. In contrast, variant R221C (where the affected residue is located near the DNA-binding site) showed permanently lower activation relative to WT APE1, whereas neighboring SNP N222H did not cause a noticeable difference as compared to WT APE1. Buried substitution P311S had an inconsistent effect, whereas each substitution at the DNA-binding site, M270T and R274Q, resulted in the lowest stimulation by BER proteins. Protein-protein molecular docking was performed between repair proteins to identify amino acid residues involved in their interactions. The data uncovered differences in the effects of BER proteins on APE1, indicating an important role of protein-protein interactions in the coordination of the repair pathway.

Asunto(s)

ADN Glicosilasas/química , Reparación del ADN , ADN-(Sitio Apurínico o Apirimidínico) Liasa/química , ADN/química , Sustitución de Aminoácidos , Sitios de Unión , ADN/genética , ADN/metabolismo , Daño del ADN , ADN Glicosilasas/genética , ADN Glicosilasas/metabolismo , ADN Polimerasa beta/química , ADN Polimerasa beta/genética , ADN Polimerasa beta/metabolismo , ADN-(Sitio Apurínico o Apirimidínico) Liasa/genética , ADN-(Sitio Apurínico o Apirimidínico) Liasa/metabolismo , Expresión Génica , Humanos , Cinética , Simulación del Acoplamiento Molecular , Conformación de Ácido Nucleico , Oligodesoxirribonucleótidos/química , Oligodesoxirribonucleótidos/genética , Oligodesoxirribonucleótidos/metabolismo , Polimorfismo de Nucleótido Simple , Antígeno Nuclear de Célula en Proliferación/química , Antígeno Nuclear de Célula en Proliferación/genética , Antígeno Nuclear de Célula en Proliferación/metabolismo , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Especificidad por Sustrato , Proteína 1 de Reparación por Escisión del Grupo de Complementación Cruzada de las Lesiones por Rayos X/química , Proteína 1 de Reparación por Escisión del Grupo de Complementación Cruzada de las Lesiones por Rayos X/genética , Proteína 1 de Reparación por Escisión del Grupo de Complementación Cruzada de las Lesiones por Rayos X/metabolismo

RESUMEN

Nanostructured hydroxyapatite (HAp) has been applied widely as a scaffold material for bone tissue engineering for its good osteoinduction and biodegradability. However, the degradation process and the distribution of degraded HAp within the bone-defect cavity is still not clear. To visually study the behavior of HAp in bone repair process, a membrane of HAp/terbium (Tb)-HAp nanowires (NWs) was prepared with a concentric circle structure (CCS), of which the inner circle and the outer ring were constructed with Tb-HAp and HAp NWs, respectively. HAp/Tb-HAp CCS membrane possessed good osteogenic capacity and efficient fluorescence in the center for visualization. The in vitro experimental results proved that the Tb-HAp and HAp NWs membranes both presented high cytocompatibility and adequate efficiency to induce osteogenic differentiation of bone marrow stem cells (BMSCs). HAp/Tb-HAp CCS membranes were then implanted into a rat calvarial bone-defect model to study the behavior of HAp in bone repair process in vivo by tracking the fluorescence distribution. The results showed that the fluorescence of Tb-HAp diffused gradually from the inner circle to the outer ring, which suggested that the HAp was first degraded, and then the degraded product was diffused and finally reconstructed. Further, the histological results proved that the doping of Tb did not impair the promotive effect of HAp on bone repair process. Therefore, this study provided a visual method to observe the degradation-diffusion-reconstruction behavior of HAp nanomaterials in bone repair process. STATEMENT OF SIGNIFICANCE: The study of dynamic degradation process of implanted hydroxyapatite (HAp) materials in bone-defect cavity is of great significance to bone tissue engineering applications. Here, we designed a HAp/Tb-HAp nanowires (NWs) membrane with concentric circle structure (CCS) to visibly observe the behavior of HAp during bone repair process. HAp/Tb-HAp CCS membrane possessed both osteoinduction ability and fluorescence property. Calvarial bone-defect repair experiments in vivo showed that the fluorescence of Tb-HAp diffused gradually from inner circle to outer ring, which suggested that HAp was first degraded, then diffused and finally reconstructed. Therefore, this invention provides not only a visible method to observe the degradation-diffusion-reconstruction behavior of HAp-based biomaterials, but also a basic understanding of the dynamic change of HAp-based biomaterials.

Asunto(s)

Regeneración Ósea/efectos de los fármacos , Durapatita/farmacología , Ingeniería de Tejidos/métodos , Animales , Forma de la Célula/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Difusión , Fluorescencia , Masculino , Membranas Artificiales , Nanocables/química , Osteogénesis/efectos de los fármacos , Ratas Wistar , Terbio/química , Microtomografía por Rayos X

RESUMEN

A growing body of evidence suggests that elevated levels of reactive oxygen species (ROS) in the airways caused by exposure to gas phase pollutants or particulate matter are able to activate dendritic cells (DCs); however, the exact mechanisms are still unclear. When present in excess, ROS can modify macromolecules including DNA. One of the most abundant DNA base lesions is 7,8-dihydro-8-oxoguanine (8-oxoG), which is repaired by the 8-oxoguanine DNA glycosylase 1 (OGG1)-initiated base excision repair (BER) (OGG1-BER) pathway. Studies have also demonstrated that in addition to its role in repairing oxidized purines, OGG1 has guanine nucleotide exchange factor activity when bound to 8-oxoG. In the present study, we tested the hypothesis that exposure to 8-oxoG, the specific product of OGG1-BER, induces functional changes of DCs. Supporting our hypothesis, transcriptome analysis revealed that in mouse lungs, out of 95 genes associated with DCs' function, 22 or 42 were significantly upregulated after a single or multiple intranasal 8-oxoG challenges, respectively. In a murine model of allergic airway inflammation, significantly increased serum levels of ovalbumin (OVA)-specific IgE antibodies were detected in mice sensitized via nasal challenges with OVA+8-oxoG compared to those challenged with OVA alone. Furthermore, exposure of primary human monocyte-derived DCs (moDC) to 8-oxoG base resulted in significantly enhanced expression of cell surface molecules (CD40, CD86, CD83, HLA-DQ) and augmented the secretion of pro-inflammatory mediators IL-6, TNF and IL-8, whereas it did not considerably influence the production of the anti-inflammatory cytokine IL-10. The stimulatory effects of 8-oxoG on human moDCs were abolished upon siRNA-mediated OGG1 depletion. Collectively, these data suggest that OGG1-BER-generated 8-oxoG base-driven cell signaling activates DCs, which may contribute to initiation of both the innate and adaptive immune responses under conditions of oxidative stress.

Asunto(s)

Reparación del ADN , ADN/química , Células Dendríticas/inmunología , Guanina/análogos & derivados , Inmunidad Adaptativa , Animales , Quimiocinas/metabolismo , Citocinas/metabolismo , ADN Glicosilasas/metabolismo , Células Dendríticas/efectos de los fármacos , Femenino , Perfilación de la Expresión Génica , Guanina/farmacología , Factores de Intercambio de Guanina Nucleótido/metabolismo , Humanos , Inmunoglobulina E/inmunología , Inmunoglobulina M/inmunología , Inflamación , Ratones , Ratones Endogámicos BALB C , Monocitos/inmunología , Estrés Oxidativo , ARN Interferente Pequeño/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal

RESUMEN

We report a nano-technological method of creating a micrometer sized hole on the live cell membrane using atomic force microscope (AFM) and its resealing process at the single cellular level as a model of molecular level wound healing. First, the cell membrane was fluorescently labeled with Kusabira Orange (KO) which was tagged to a lipophilic membrane-sorting peptide. Then a glass bead glued on an AFM cantilever and modified with phospholipase A2 was made to contact the cell membrane. A small dark hole (4-14 µm2 in area) was created on the otherwise fluorescent cell surface often being accompanied by bleb formation. Refilling of holes with KO fluorescence proceeded at an average rate of ~0.014µm2s-1. The fluorescent lumps which initially surrounded the hole were gradually lost. We compared the present result with our previous ones on the repair processes of artificially damaged stress fibers (Graphical Abstract: Figure S2).

Asunto(s)

Membrana Celular/patología , Fibras de Estrés/patología , Cicatrización de Heridas , Membrana Celular/metabolismo , Membrana Celular/ultraestructura , Células HeLa , Humanos , Microscopía de Fuerza Atómica , Paxillin/análisis , Paxillin/metabolismo , Análisis de la Célula Individual , Fibras de Estrés/metabolismo , Fibras de Estrés/ultraestructura

RESUMEN

Neste trabalho desenhou-se bases de desenvolvimento, caracterização e avaliação dos aspectos biológicos relacionados ao Sistema BoneLithium, idealizado a partir da associação de partículas de carbonato de lítio dispersas em matriz gel de carbopol®, com capacidade de atuar como um sistema liberador de fármacos. Metodologicamente este estudo se dividiu em quatro partes: Na primeira delas, o objetivo central foi o desenvolvimento e a caracterização do biomaterial através da manipulação farmacológica. Na segunda etapa, avaliou-se a reação tecidual em subcutâneo de ratos, na terceira a influência das partículas de lítio liberadas pelo Sistema BoneLithium no reparo ósseo através de modelos experimentais utilizando coelhos, e por ultimo, a capacidade de cicatrização de defeitos ósseos criados cirurgicamente em calvária de ratos, tratados com o biomaterial e diferentes opções de enxertos ósseos com o objetivo de comparar a eficiência do Sistema BoneLithium aos protocolos pré-existentes. Experimentalmente, avaliou-se a reação tecidual onde se utilizou 15 ratos machos divididos aleatatoriamente em 5 grupos onde implantouse no subcutâneo tubos de butterfly contendo o biomaterial por períodos de preservação recomendados pela norma ADA 10993 para teste de reação tecidual. Os resultados demonstram que o Sistema BoneLithium apresenta reação tecidual normal. Para a avaliação do comportamento biológico do Sistema BoneLithium foram utilizados coelhos brancos adultos da raça New Zealand nos quais defeitos ósseos bilaterais de 1 cm de diâmetro foram confeccionados cirurgicamente na calvária e foram Tratados com o Sistema Bone Lithium do (lado Direito) e somente o Gel de Carbopol (lado esquerdo)/Coágulo sanguíneo (controle). A Histomorfometria demonstrou comportamento favorável ao reparo ósseo e adicionalmente através de Microtomografia Computadorizada (CT SKYSCAN), foi possível constatar diferenças significativas considerando p> 0.05 (ANOVA, Tukey) para o processo de reparo ósseo. A avaliação da performance do Sistema BoneLithium utilizando ratos Wistar nos quais foram criados defeitos críticos no centro da calvária e tratados com diferentes modalidades de enxertos ósseos (controle, autógeno, osso de banco (Unioss®, Marília Brasil), Bio-Oss® e associações com o Sistema BoneLithium. A histomorfometria mostrou diferenças significativas considerando p> 0.05 (ANOVA, Tukey) para avaliação de tecido conjuntivo pré-osteogênico e tecido ósseo neoformado, e quando avaliado qualitativamente por tomografia computadorizada de feixe cônico (I cat Cone Beam FOV 0.05 Xoran Tecnology, LLC, EUA e E-vol, CDT, Brasil), observaram-se áreas de neoformação óssea compatíveis com hiperdensidade óssea em toda a extensão do defeito quando apuradas em analises de paridade em escala Hounsfield. Dessa forma, conclui-se que no contexto deste estudo é possível concluir que Sistema BoneLithium representa uma alternativa com potencial viabilidade clínica e necessita seguimento de aplicação em novas metodologias.(AU)

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In this work, bases for the development, characterization and evaluation of the biological aspects related to the BoneLithium System were designed, based on the association of lithium carbonate particles dispersed in carbopol® gel matrix, capable of acting as a drug-releasing system. Methodologically this study was divided in four parts: In the first one, the central objective was the development and characterization of the biomaterial through the pharmacological manipulation. In the second step, the tissue reaction was evaluated in subcutaneous of rats, in the third the influence of the lithium particles released by BoneLithium System in the bone repair through experimental models using rabbits, and finally, the capacity of healing of bone defects created surgically in Calvaria of rats, treated with the biomaterial and different options of bone grafts with the objective to compare the efficiency of the BoneLithium System to the preexisting protocols. Experimentally, the tissue reaction was evaluated in which 15 male rats were randomly divided into 5 groups, where butterfly tubes containing the biomaterial were implanted in the subcutaneous tubes for preservation periods recommended by the ADA 10993 standard for biocompatibility test. The results demonstrate that the BoneLithium System is tissue reaction positive. To evaluate the biological behavior of the BoneLithium System, adult New Zealand white rabbits were used in which bilateral bone defects of 1 cm in diameter were surgically made on calvaria and treated with the Bone Lithium System (right side) and only Gel Of Carbopol (left side) / blood clot (control). Histomorphometry showed a favorable behavior to bone repair and, in addition, through Computerized Microtomography (CT SKYSCAN), it was possible to verify significant differences considering p> 0.05 (ANOVA, Tukey) for the bone repair process. The evaluation of the performance of the BoneLithium System using Wistar rats in which critical defects were created at the calvarial center and treated with different bone graft modalities (control, autogenous, bone bank (Unioss®, Marília Brazil), Bio-Oss® and associations (ANOVA, Tukey) for evaluation of pre osteogenic connective tissue and neoformed bone tissue, and when assessed qualitatively by cone beam computed tomography (I cat - Cone Beam - FOV 0.05 - Xoran Tecnology, LLC, USA and E-vol, CDT, Brazil), areas of bone neoformation compatible with bone hyperdensity throughout the extent of the defect were ascertained in Hounsfield scale parity analyzes, It is concluded that in the context of this study it is possible to conclude that the BoneLithium System represents an alternative with potential clinical feasibility And requires follow-up of application in new methodologies.(AU)

Asunto(s)

Animales , Masculino , Conejos , Ratas , Regeneración Ósea/efectos de los fármacos , Regeneración Ósea/fisiología , Sustitutos de Huesos/química , Sustitutos de Huesos/uso terapéutico , Carbonato de Litio/química , Carbonato de Litio/uso terapéutico , Materiales Biocompatibles/química , Materiales Biocompatibles/uso terapéutico , Trasplante Óseo/métodos , Tomografía Computarizada de Haz Cónico , Ratas Wistar , Reproducibilidad de los Resultados , Factores de Tiempo

RESUMEN

BACKGROUND: DNA glycosylases remove the modified, damaged or mismatched bases from the DNA by hydrolyzing the N-glycosidic bonds. Some enzymes can further catalyze the incision of a resulting abasic (apurinic/apyrimidinic, AP) site through ß- or ß,δ-elimination mechanisms. In most cases, the incision reaction of the AP-site is catalyzed by special enzymes called AP-endonucleases. METHODS: Here, we report the kinetic analysis of the mechanisms of modified DNA transfer from some DNA glycosylases to the AP endonuclease, APE1. The modified DNA contained the tetrahydrofurane residue (F), the analogue of the AP-site. DNA glycosylases AAG, OGG1, NEIL1, MBD4(cat) and UNG from different structural superfamilies were used. RESULTS: We found that all DNA glycosylases may utilise direct protein-protein interactions in the transient ternary complex for the transfer of the AP-containing DNA strand to APE1. CONCLUSIONS: We hypothesize a fast "flip-flop" exchange mechanism of damaged and undamaged DNA strands within this complex for monofunctional DNA glycosylases like MBD4(cat), AAG and UNG. Bifunctional DNA glycosylase NEIL1 creates tightly specific complex with DNA containing F-site thereby efficiently competing with APE1. Whereas APE1 fast displaces other bifunctional DNA glycosylase OGG1 on F-site thereby induces its shifts to undamaged DNA regions. GENERAL SIGNIFICANCE: Kinetic analysis of the transfer of DNA between human DNA glycosylases and APE1 allows us to elucidate the critical step in the base excision repair pathway.

Asunto(s)

Daño del ADN , ADN Glicosilasas/química , Reparación del ADN , ADN-(Sitio Apurínico o Apirimidínico) Liasa/química , Fluorescencia , Humanos

RESUMEN

We previously reported that bone morphogenetic proteins (BMPs) and their endogenous antagonist noggin are expressed in the brain weeks after an ischemic insult. Here, to define their roles in ischemic brain tissue repair and remodeling, we infused recombinant BMP7 or noggin into the ipsilateral ventricle of mice for 2weeks starting 2weeks after transient middle cerebral artery occlusion (MCAO). Four weeks after MCAO, we measured ischemic brain volume, functional recovery, and molecules related to neurogenesis and angiogenesis such as synaptophysin, GAP-43, and VEGF. Noggin-treated mice but not BMP7-treated mice showed preserved ipsilateral brain volume and reduced neurological deficits compared with artificial cerebrospinal fluids (aCSF)-treated mice. Noggin treatment also decreased glial scar thickness, increased levels of GAP-43 and VEGF protein, and increased the number of Iba1-positive activated microglia in the ipsilateral brain. Furthermore, noggin treatment decreased M1 markers (IL-1ß, TNF-α, IL-12, CCL2 and CD86) and increased M2 markers (IL-1ra, IL-10, arginase 1, CD206 and Ym1) of activated microglia, suggesting a shift from M1 to M2 phenotypes. These results suggest that noggin improves functional recovery from ischemic stroke and enhances alternatively activated microglia, thereby promoting tissue repair and remodeling.

Asunto(s)

Isquemia Encefálica/tratamiento farmacológico , Encéfalo/efectos de los fármacos , Proteínas Portadoras/uso terapéutico , Microglía/efectos de los fármacos , Fármacos Neuroprotectores/uso terapéutico , Animales , Proteína Morfogenética Ósea 7/uso terapéutico , Encéfalo/metabolismo , Isquemia Encefálica/etiología , Proteínas Portadoras/administración & dosificación , Infarto de la Arteria Cerebral Media/complicaciones , Masculino , Ratones , Ratones Endogámicos C57BL , Microglía/metabolismo , Fármacos Neuroprotectores/administración & dosificación , Proteínas Recombinantes/uso terapéutico

RESUMEN

Modeling of the RadA family mechanism is crucial to understanding the DNA SOS repair process. In a 2007 report, the archaeal RadA proteins function as rotary motors (linker region: I71-K88) such as shown in Figure 1. Molecular simulations approaches help to shed further light onto this phenomenon. We find 11 rotary residues (R72, T75-K81, M84, V86 and K87) and five zero rotary residues (I71, K74, E82, R83 and K88) in the simulations. Inclusion of our simulations may help to understand the RadA family mechanism.

Asunto(s)

Proteínas Arqueales/química , Proteínas de Unión al ADN/química , Rec A Recombinasas/química , Adenosina Trifosfato/química , Algoritmos , Secuencia de Aminoácidos , Simulación por Computador , Citoesqueleto/metabolismo , Reparación del ADN , Simulación de Dinámica Molecular , Datos de Secuencia Molecular , Unión Proteica , Estructura Terciaria de Proteína , Homología de Secuencia de Aminoácido

RESUMEN

Linear focal elastosis (LFE) is a rare dermal elastosis characterized by hypertrophic yellowish linear plaques and increased abnormal elastic tissues in the lumbosacral area. Although the pathogenesis of this disorder remains unknown, it may be associated with keloidal repair process (KRP) of elastic tissues in striae distensae (SD), because there have been some reported cases of LFE accompanied by SD. We herein report a 14-year-old boy with LFE following SD in the lumbar region. Our case supports the hypothesis of KRP in the pathogenesis of LFE. Immunohistochemical study for transforming growth factor-beta (TGF-ß) was negative. Therefore, we assume that the pathogenesis of KRP in LFE is different from that of keloid development, which is the TGF-ß signaling pathway.

RESUMEN

Linear focal elastosis (LFE) is a rare dermal elastosis characterized by hypertrophic yellowish linear plaques and increased abnormal elastic tissues in the lumbosacral area. Although the pathogenesis of this disorder remains unknown, it may be associated with keloidal repair process (KRP) of elastic tissues in striae distensae (SD), because there have been some reported cases of LFE accompanied by SD. We herein report a 14-year-old boy with LFE following SD in the lumbar region. Our case supports the hypothesis of KRP in the pathogenesis of LFE. Immunohistochemical study for transforming growth factor-beta (TGF-beta) was negative. Therefore, we assume that the pathogenesis of KRP in LFE is different from that of keloid development, which is the TGF-beta signaling pathway.

Asunto(s)

Adolescente , Humanos , Tejido Elástico , Queloide , Región Lumbosacra , Estrías de Distensión , Factor de Crecimiento Transformador beta

RESUMEN

After bone damage of the fetal rat femurs induced by administrating cyclophosphamide(CP),(1/8 LD50) to the pregnant rat on 13th day of gestation, the effects of serum and ascorbic acid on the repair process of the bone during organ culture were studied, histologically and scanning electron microscopically. CP-damaged fetal femurs harvested at 20 days of gestation were cultured fro 2, 5 and 7 days in the waymouth media(WM) with or without fetal bovine serum(FBS) and ascorbic acid, and were observed with light microscope and JSM-35C scanning electron microscope. The results were as follows:1. CP-damaged bone tissue cultured in WM with 10% FBS showed relatively enhanced activities in the differentiation of chondrocytes and ossificstion as compared to that cultured in WM. 2. CP-damaged bone tissue cultured in WM with 10% FBS and 100µg/ml ascorbic acid, showed increase in the length of the bone marrow cavity, and active formation of new osteoid and collagen bundles. 3. The bone tissues cultured in WM with 10% FBS and 400µg/ml ascorbic acid revealed active deposition of bone matrix, thickening of periosteum and marked elongation of the bone marrow cavity. 4. Bone trabeculae of CP-damaged femurs cultured for 2 days in WM showed poor cell proliferation and insignificant bone matix formation. 5. The number of new cells and the amount of the collagen fibrils increased on the bone trabeculae of the bone cultured in WM with 10% FBS as compared to that cultured in WM and this increase was enhanced as the culture time progressed. 6. A remarkable increase was noted in the number of cells and collagen fibrils in the bone tissues cultured in WM with 10% FBS and ascorbic acid than in those cultured in WM with 10% FBS. 7. The number of the spherules formed by cellular component with collagen fibrils is more numerous than that formed by calcospherites associated with collagen fibrils.

Asunto(s)

Animales , Embarazo , Ratas , Ácido Ascórbico , Huesos , Médula Ósea , Matriz Ósea , Proliferación Celular , Condrocitos , Colágeno , Ciclofosfamida , Fémur , Técnicas In Vitro , Técnicas de Cultivo de Órganos , Periostio