RESUMO
BACKGROUND: Novel picosecond lasers using a diffractive optical element (P-DOE) have been available for skin resurfacing with distinct mechanisms. However, there are limited data directly comparing P-DOE and conventional fractional lasers for the treatment of atrophic acne scarring. OBJECTIVES: We sought to compare the efficacy and safety of a 1064-nm neodymium-doped yttrium aluminium garnet P-DOE and a non-ablative fractional laser (NAFL) in the treatment of acne scarring. METHODS: A prospective, randomized, split-face, controlled trial was performed. One randomly assigned half-side of each patient's face (n = 25) was treated with four consecutive sessions of P-DOE at 3-week intervals and the other side with NAFL, with subsequent follow-up for 8 weeks after the final sessions. The efficacy and safety of the two lasers were determined by the Echelle d'Evaluation Clinique des Cicatrices d'acné (Scale of Clinical Evaluation of Acne Scars; ECCA) grading scale, Investigator's Global Assessment (IGA) score and patients' reports at the final visit. Histologic analysis was also performed. RESULTS: The P-DOE-treated side achieved a significantly better improvement in acne appearance (ECCA per cent reduction: 55% vs. 42%) with less severe pain (4.3 vs. 5.6) (P < 0.05). The IGA score and subjective satisfaction were consistent with ECCA score results. Occurrences of treatment-related side-effects were also lower in the group treated with P-DOE (P < 0.05). Histologic analysis revealed elongation and increased density of neocollagen fibres, elastic fibres and mucin throughout the dermis from both sides. CONCLUSIONS: Compared with NAFL, P-DOE afforded better clinical outcomes and fewer side-effects in the treatment of acne scarring in Asian patients.
Assuntos
Acne Vulgar , Lasers de Estado Sólido , Acne Vulgar/complicações , Alumínio , Cicatriz/etiologia , Cicatriz/patologia , Érbio , Humanos , Lasers de Estado Sólido/uso terapêutico , Neodímio , Satisfação do Paciente , Estudos Prospectivos , Resultado do Tratamento , ÍtrioRESUMO
BACKGROUND: While device-based acne treatments are widely applied for patients not tolerating conventional medications, related controlled studies have been still limited. Recently, non-ablative 1450-nm diode laser (DL) and fractional microneedling radiofrequency (FMR) have been effectively used for acne, in addition to well-recognized dermal remodelling effects. OBJECTIVE: To compare the clinical course of acne treatment between DL and FMR. METHODS: Twenty-five Korean patients with mild-to-moderate facial acne completed treatments with DL and FMR through a 20-week, randomized split-face study. One randomly assigned half side of each patient's face received DL and the other side by FMR. Treatments were scheduled to receive three consecutive sessions at 4-week intervals. Objective assessments including revised Leeds grades, lesion counts, sebum output measurements, and patients' subjective satisfaction were investigated. RESULTS: Both DL and FMR demonstrated steady improvement of acne and seborrhoea during treatment sessions. While results between two devices were similar during treatment sessions, FMR was superior to DL in the 12-week follow-up. Patients' subjective assessments for seborrhoea improvement were similar between two devices, while those for acne, skin texture, and acne scars were more satisfactory for FMR. For safety profile, no significant difference was observed between two regimens, while mild postinflammatory hyperpigmentation was observed only in DL side. CONCLUSION: Both DL and FMR demonstrated efficacies for acne and seborrhoea, with reasonable safety profile. FMR was more effective than DL for the long-term maintenance, and subjective assessments for texture and scar improvements. Therefore, a few sessions of these devices would be a viable option for acne treatments.
Assuntos
Acne Vulgar/terapia , Dermatite Seborreica/terapia , Terapia a Laser/instrumentação , Agulhas , Acne Vulgar/patologia , Acne Vulgar/radioterapia , Adolescente , Adulto , Dermatite Seborreica/patologia , Dermatite Seborreica/radioterapia , Feminino , Humanos , Terapia a Laser/efeitos adversos , Masculino , Agulhas/efeitos adversos , Satisfação do Paciente , Estudos Prospectivos , República da Coreia , Sebo/metabolismo , Índice de Gravidade de Doença , Resultado do Tratamento , Adulto JovemRESUMO
BACKGROUND: There are insufficient data on the long-term outcome of a combination therapy that comprises phototherapy and topical administration of tacrolimus. AIM: To evaluate the clinical efficacy according to the duration of treatment and in vitro results of a combination therapy involving topical tacrolimus and an excimer laser in the treatment of vitiligo. METHODS: In total, 276 patients with nonsegmental vitiligo were treated with an excimer laser twice weekly, or with tacrolimus ointment twice daily, or both. The melanin contents and levels of melanogenic enzymes were measured in cultured human melanocytes treated with tacrolimus and/or excimer laser. RESULTS: After adjusting for potential confounders, the combination of tacrolimus plus excimer laser was significantly more effective than either tacrolimus or excimer laser alone (P < 0.001 and P < 0.01, respectively) for the first 6 months. However, this superiority was not observed after the initial 6 months of treatment. In vitro, the combination of tacrolimus plus excimer laser led to a higher level of melanogenesis than with either treatment alone. CONCLUSIONS: A combination treatment with topical tacrolimus and an excimer laser may be useful as an induction therapy for up to 6 months, but continuation of this therapy for > 6 months might not provide a better final outcome than monotherapy.
Assuntos
Imunossupressores/uso terapêutico , Fototerapia/métodos , Tacrolimo/uso terapêutico , Vitiligo/terapia , Administração Tópica , Adolescente , Adulto , Idoso , Análise de Variância , Western Blotting , Criança , Pré-Escolar , Terapia Combinada , Feminino , Humanos , Lactente , Oxirredutases Intramoleculares/metabolismo , Modelos Logísticos , Masculino , Melaninas/metabolismo , Melanócitos/metabolismo , Pessoa de Meia-Idade , Monofenol Mono-Oxigenase/metabolismo , Fatores de Tempo , Tripsina/metabolismo , Vitiligo/tratamento farmacológico , Vitiligo/metabolismo , Adulto JovemRESUMO
Last year, the KSTAR divertor material was changed from carbon to tungsten tiles. An optimized collection optic design for divertor Thomson scattering diagnostics in KSTAR was conducted for electron temperature (1-100 eV) and electron density (1 × 1018-1 × 1019 m-3) profile diagnostics. This diagnostic system will utilize a 1064 nm Nd:YAG laser directly from the K-top port toward the beam dump located at K-bottom, while collecting scattered light from five spatial points in the divertor area via collection optics situated in the j-middle port. Given spatial limitations, the solid angle of measurement points is limited, and the collection optic design facing the tungsten divertor is susceptible to stray light. So, the design of the collection optic is important for divertor Thomson scattering diagnostics. For optimal performance, we performed two types of collection optic designs: Cooke-triplet and double-Gaussian. We present performance ray tracing analysis results for both designs and derive the optimal design.
Assuntos
Melanoma/patologia , Doenças da Unha/patologia , Unhas/patologia , Idoso , Feminino , HumanosRESUMO
The objective of this experiment was to investigate the effect of feeding corn distillers dried grains with solubles (DDGS) naturally contaminated with deoxynivalenol (DON) on growth performance, meat quality, intestinal permeability, and utilization of energy and nutrients in broiler chickens. Two trials (growth and metabolism trials) were conducted. In the growth trial, a total of four hundred 7-day-old Ross 308 broiler chicks were allotted to 1 of 5 dietary treatments with 8 replicates in a completely randomized design. The diets were formulated to contain 5 inclusion levels of 0, 5, 10, 15, or 20% DON-contaminated DDGS in diets and were fed to birds for 21 d. Results indicated that increasing inclusion levels of DON-contaminated DDGS decreased (linear, P < 0.01) BW gain and feed efficiency of broiler chickens. The relative organ weights of the liver and breast were decreased (linear and quadratic, P < 0.05) by increasing inclusion levels of DON-contaminated DDGS in diets. The transepithelial electrical resistance values as a measure of intestinal permeability were decreased (linear, P < 0.05) by increasing inclusion levels of DON-contaminated DDGS in diets. In the metabolism trial, a total of twenty four 22-day-old Ross 308 broiler chickens were allotted to 1 of 3 dietary treatments consisting of 0, 10, or 20% inclusion of DON-contaminated DDGS in diets. Each treatment had 8 replicates. Increasing inclusion levels of DON-contaminated DDGS in diets decreased (linear and quadratic, P < 0.05) MEn (AMEn and TMEn) and apparent total tract retention of nitrogen and acid-hydrolyzed ether extract in diets. In conclusion, feeding diets containing more than 10% DON-contaminated DDGS to broiler chickens has negative effects on growth performance, intestinal permeability, and utilization of energy and nutrients in diets. Therefore, it is suggested that if DDGS is contaminated with DON, inclusion level of DDGS should be limited, possibly at less than 5.0% in broiler diets.
Assuntos
Ração Animal , Galinhas , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Animais , Dieta/veterinária , Grão Comestível , Carne , Nutrientes , Permeabilidade , Tricotecenos , Zea maysRESUMO
BACKGROUND: Epidermal growth factor receptor (EGFR) critically regulates tumour cell division, survival and metastasis. Agents that inhibit EGFR have been used in the treatment of advanced-stage malignancies, but cause variable cutaneous side-effects, most often papulopustular eruptions and xerosis. OBJECTIVES: We assayed expression of inflammatory cytokines [interleukin (IL)-1alpha, tumour necrosis factor (TNF)-alpha, interferon (IFN)-gamma, human leucocyte antigen (HLA)-DR and intercellular adhesion molecule (ICAM)-1], differentiation markers (filaggrin, involucrin and loricrin) and phosphorylated EGFRs (pEGFRs) in papulopustular eruptions to determine the association between these markers and the eruptions caused by cetuximab. PATIENTS/METHODS: Twelve papulopustular lesion biopsies were selected from patients with colon cancer who had received cetuximab treatment. Immunohistochemistry and immunofluorescence with a confocal laser scanning microscopy were performed. RESULTS: Filaggrin expression decreased and expression of involucrin, various inflammatory markers (IL-1alpha, TNF-alpha, ICAM-1 and HLA-DR) increased and the expression of pEGFR was markedly downregulated in papulopustular eruptions. In perilesions, decreased pEGFR expression was noted in hair follicles compared with interfollicular epidermis. The increase of IL-1alpha and TNF-alpha was observed in perilesions as in the lesions. CONCLUSIONS: The early inflammatory events (IL-1alpha and TNF-alpha expression) seen, and the lack of pEGFR in perilesional follicles, indicate that inflammatory events induced by EGFR inhibition may initiate papulopustular eruptions along with the altered differentiations. The decrease of filaggrin may contribute to the pathogenesis of the xerosis caused by cetuximab.
Assuntos
Anticorpos Monoclonais/efeitos adversos , Antineoplásicos/efeitos adversos , Citocinas/metabolismo , Toxidermias/imunologia , Fator de Crescimento Epidérmico/metabolismo , Epiderme/metabolismo , Anticorpos Monoclonais Humanizados , Biomarcadores/análise , Biomarcadores/metabolismo , Cetuximab , Toxidermias/metabolismo , Receptores ErbB/análise , Receptores ErbB/metabolismo , Feminino , Proteínas Filagrinas , Antígenos HLA-DR/análise , Antígenos HLA-DR/metabolismo , Humanos , Molécula 1 de Adesão Intercelular/análise , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-1alfa/metabolismo , Proteínas de Filamentos Intermediários/análise , Proteínas de Filamentos Intermediários/metabolismo , Masculino , Pessoa de Meia-Idade , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/metabolismoRESUMO
AIMS: The rapidly increasing prevalence of chronic diseases is an important challenge to healthcare systems worldwide. To improve the quality and efficiency of chronic disease care, we investigated the effectiveness and applicability of the Ubiquitous Chronic Disease Care (UCDC) system using cellular phones and the internet for overweight patients with both Type 2 diabetes and hypertension. METHODS: We conducted a randomized, controlled clinical trial over 3 months that included 123 patients at a university hospital and a community public health centre. RESULTS: After 12 weeks, there were significant improvements in HbA(1c) in the intervention group (7.6 +/- 0.9% to 7.1 +/- 0.8%, P < 0.001) compared with the control group (7.4 +/- 0.9% to 7.6 +/- 1.0%, P = 0.03). Furthermore, we observed a significant reduction in systolic and diastolic blood pressure, as well as improvements in total cholesterol, low-density lipoprotein-cholesterol and triglyceride levels in the intervention group. Furthermore, there was a significant increase in adiponectin levels in the intervention group compared with the control group, although high-sensitivity C-reactive protein and interleukin-6 levels did not change in either group. CONCLUSIONS: The novel UCDC system presented in this paper improved multiple metabolic parameters simultaneously in overweight patients with both Type 2 diabetes and hypertension.
Assuntos
Automonitorização da Glicemia/métodos , Monitorização Ambulatorial da Pressão Arterial/métodos , Telefone Celular , Atenção à Saúde/métodos , Internet , Adulto , Idoso , Estudos de Casos e Controles , Diabetes Mellitus Tipo 2/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Sobrepeso , Cooperação do Paciente , Educação de Pacientes como AssuntoRESUMO
The objective of the current experiment was to investigate the effect of stocking density and sex on growth performance, meat quality, and intestinal barrier function in broiler chickens. The experiment was conducted in a completely randomized design with a 2 × 4 factorial arrangement consisting of sex and four different stocking densities in battery cages. A total of 540 1-d-old Ross 308 broiler chickens were allotted to one of eight treatments with five replicates. Within each sex, birds were raised at four different stocking densities of 15.2, 20.2, 25.3, or 30.4 birds/m2 from 1 to 28 d of age. Different stocking densities were achieved by raising a different number of birds per battery cage with identical floor size (0.76 m × 0.78 m). At the end of the experiment, two birds per replicate were euthanized by CO2 asphyxiation to collect tissue samples for further analyses. Results indicated that no interactions between sex and stocking density were observed for all measurements except for serum lipopolysaccharide (LPS) concentrations. Increasing stocking density decreased (linear, P < 0.01) body weight gain and feed intake, but had no negative effects on meat quality. Trans-epithelial electrical resistance values, a measure of intestinal permeability, were decreased (linear, P < 0.01) with increasing stocking density, regardless of sex. Accordingly, serum LPS concentrations were increased (linear, P < 0.01) with increasing stocking density. However, increasing stocking density increased serum LPS concentrations in male broiler chickens, but had no effects on female broiler chickens, showing an interaction (P < 0.01). The expression of zonula occludens-1 (ZO-1) and junctional adhesion molecule B (JAM-2) was decreased (linear, P < 0.05) with increasing stocking density. In conclusion, increasing stocking density decreases broiler performance regardless of sex and this negative effect is likely associated with decreased intestinal barrier function.
Assuntos
Criação de Animais Domésticos/métodos , Galinhas/fisiologia , Intestinos/fisiologia , Carne/análise , Fenômenos Fisiológicos da Nutrição Animal , Animais , Galinhas/crescimento & desenvolvimento , Feminino , Masculino , Densidade Demográfica , Distribuição Aleatória , Fatores SexuaisRESUMO
The objective of the present experiment was to investigate the effect of stocking density (SD) and dietary supplementation of crystalline tryptophan (Trp) on growth performance and intestinal barrier function in broiler chickens raised in a floor pen. The experiment was conducted using a completely randomized design with a 2 × 2 factorial arrangement consisting of 2 different SD and 2 supplemental levels of dietary Trp. A total of 1,140 Ross 308 broiler chickens at 21 d of age were allotted to 1 of 4 treatments with 5 replicates. Low SD (9 birds/m2) and high SD (18 birds/m2) were achieved by raising different number of birds per identical floor pen (2.0 m × 2.4 m). The basal diet was formulated with no supplemental Trp in diets to meet or exceed nutrient recommendation of the Ross 308 manual. The calculated concentrations of total Trp and digestible Trp in the basal diet were 0.19 and 0.16%, respectively. The other diet was prepared by adding 0.16% crystalline Trp to the basal diet. Diets were fed to birds for 21 d. At the end of the experiment, 2 birds per replicate were euthanized to collect tissue samples for further analyses. Results indicated that there were no interactions between SD and dietary Trp for all measurements. For the main effects, birds raised at a low SD had greater (P < 0.01) body weight gain, feed intake, and feed efficiency than those raised at a high SD. However, supplementation of dietary Trp had no effect on broiler performance. Furthermore, there were no main effects of SD and dietary Trp on intestinal barrier functions. In conclusion, broiler chickens raised in a floor pen with a high SD (18 birds/m2) have decreased growth performance with little changes in intestinal barrier functions. Supplementation of dietary Trp at 0.16% has no positive effect on broiler chickens raised in a floor pen with either a low or high SD.
Assuntos
Criação de Animais Domésticos/métodos , Galinhas/fisiologia , Dieta/veterinária , Triptofano/metabolismo , Ração Animal/análise , Fenômenos Fisiológicos da Nutrição Animal , Animais , Galinhas/crescimento & desenvolvimento , Galinhas/imunologia , Suplementos Nutricionais/análise , Abrigo para Animais , Densidade Demográfica , Distribuição Aleatória , Triptofano/administração & dosagemRESUMO
The objectives of the current experiment were to investigate the effect of dietary melamine concentrations on growth performance, excreta characteristics, plasma measurements, and melamine residue in the tissue of male and female broiler chickens. Finally, the safe levels of melamine in broiler diets were determined based on BW gain (BWG) and melamine residue in the breast meat. A total of 1,008 1-d-old Ross 308 male and female broiler chickens were allotted to 1 of 7 dietary treatments within each sex in a completely randomized design. There were 6 replicates per treatment and each replicate consisted of 12 birds. Dietary melamine concentrations were set to 0; 250; 500; 750; 1,000; 5,000; or 10,000 mg/kg by adding a purified form of melamine. Diets were provided to birds on ad libitum basis for 35 d. Results indicated that no significant interaction between sex and dietary melamine concentrations was observed for all measurements. The BW, BWG, and feed intake for birds fed diets containing 10,000 mg/kg melamine were less (P < 0.05) than for those fed other diets. Melamine residues in the kidney and breast for birds fed diets containing 10,000 mg/kg melamine were greater (P < 0.05) than for birds fed other diets. The toxic level of dietary melamine based on BWG was determined by the one-slope broken-line analysis. The resulting equation was Y = 1,851 - 0.0404 × (X - 4,292), which indicated that a greater than 4,292 mg/kg melamine in diets was toxic to broiler chickens. The safe level of dietary melamine to limit melamine residue in the broiler breast was analyzed using the linear regression, which indicated that the safe level of melamine in broiler diets was 814 mg/kg. In conclusion, less than 814 mg/kg melamine in broiler diets should be maintained to satisfy human food safety regulations for melamine residue in the breast meat of broiler chickens.
Assuntos
Galinhas/crescimento & desenvolvimento , Triazinas/toxicidade , Ração Animal/análise , Animais , Peso Corporal/efeitos dos fármacos , Dieta/veterinária , Fezes/química , Feminino , Contaminação de Alimentos , Rim/química , Masculino , Carne/análise , Triazinas/metabolismoRESUMO
Bovine brain myelin basic protein, free of associated proteolytic activity, was found to be a specific inhibitor of histone-specific protein methylase I (S-adenosyl-L-methionine:protein-L-arginine N-methyltransferase, EC 2.1.1.23) purified from bovine brain. 50% of the methyl group incorporation into the histone substrate catalyzed by the methylase I was inhibited by myelin basic protein at a concentration of 0.326 mM. However, neither of the peptide fragments (residues 1-116 and residues 117-170) generated by the chemical cleavage of myelin basic protein at the tryptophan residue retained the inhibitory activity for histone-specific protein methylase I. Proteins such as gamma-globulin, bovine serum albumin, bovine pancreatic ribonuclease and polyarginine did not exhibit significant inhibitory activity toward the enzyme. The Ki value for myelin basic protein was estimated to be 3.42 X 10(-5) M for histone-specific protein methylase I and the nature of the inhibition was uncompetitive toward histone substrate.
Assuntos
Histonas/metabolismo , Proteína Básica da Mielina/farmacologia , Proteínas Metiltransferases/antagonistas & inibidores , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , Aminoácidos/metabolismo , Animais , Encéfalo/enzimologia , Bovinos , Metilação , Proteína-Arginina N-Metiltransferases/isolamento & purificaçãoRESUMO
Myelin is a membrane characteristic of the nervous tissue and functions as an insulator to increase the velocity of the stimuli being transmitted between a nerve cell body and its target. Myelin isolated from human and bovine nervous tissue is composed of approximately 80% lipid and 20% protein, and 30% of the protein fraction constitutes myelin basic protein (MBP). MBP has an unusual amino acid at Res-107 as a mixture of NG-monomethylarginine and NG, N'G-dimethylarginine. The formation of these methylarginine derivatives is catalysed by one of the subtypes of protein methylase I, which specifically methylates Res-107 of this protein. Evidence is presented to demonstrate an involvement of this biological methylation in the integrity and maintenance of myelin.
Assuntos
Proteína Básica da Mielina/metabolismo , Animais , Encéfalo/enzimologia , Humanos , Metilação , Esclerose Múltipla/metabolismo , Bainha de Mielina/fisiologia , Proteína-Arginina N-Metiltransferases/metabolismoRESUMO
O6-Methylguanine-DNA methyltransferase, a DNA repair enzyme which transfers the methyl group of O6-methylguanine residue to a cysteinyl residue in the methyltransferase itself, was examined in rat organs by quantifying the S-methylcysteine formed in the methyl acceptor protein. Among the various organs examined, the spleen exhibited the highest enzyme specific activity followed by the thymus, liver, lung and testis. Brain had the lowest activity. The patterns of subcellular distribution of the methyltransferase in spleen and liver were different: while 75-80% of the activity was present in the nuclear fraction of the spleen, 54% of the activity in the liver was found in the nuclei and 35% in the cytosolic fraction. Forty-five and thirty-five percent of the total nuclear enzyme activity could be extracted with 1 M and 2 M NaCl solutions, respectively, indicating that the repair enzyme is not tightly bound to the nuclear matrix. When isolated nuclei were incubated with [methyl-3H]DNA substrate and subsequently fractionated into histone and non-histone protein fractions, over 90% of the radioactivity was coeluted on a Bio-Rex 70 column with the non-histone fraction and only a negligible amount of radioactivity was found to be associated with the histone fraction. The molecular mass of the [methyl-3H]methyltransferase in the non-histone fraction was determined to be 23,000, and its pI value was found to be 6.6 by two-dimensional polyacrylamide gel electrophoresis.
Assuntos
Reparo do DNA , Guanina/análogos & derivados , Metiltransferases/metabolismo , Animais , Núcleo Celular/enzimologia , Guanina/metabolismo , Fígado/enzimologia , Masculino , O(6)-Metilguanina-DNA Metiltransferase , Ratos , Baço/enzimologia , Frações Subcelulares/enzimologia , Distribuição TecidualRESUMO
C-terminal farnesyl cysteine carboxyl methylation has been known to be the last step in the post-translational modification processes of several important signal transduction proteins in eukaryotes including ras related GTP binding proteins and the gamma-subunit of heterotrimeric G proteins. Protein farnesyl cysteine carboxyl methyltransferase (PFCCMT; EC, 2.1.1.100) catalyzing the reaction is well characterized as being stimulated by guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) and suppressed by N-acetyl-S-farnesyl-L-cysteine (AFC). As an initial step to understand the physiological significance of the process, we attempted to purify the enzyme, which was partially purified 130-fold (specific activity, 143 pmol of methyl group transferred/min/mg of protein) with yield of 1.8% after purification by fast protein liquid chromatography (FPLC) on a Superdex 75 column. The enzyme was further purified with non denaturing polyacrylamide gel electrophoresis (ND-PAGE) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight of PFCCMT was determined to be about 30 kDa based on Superdex 75 FPLC as well as photoaffinity labelling with S-adenosyl-L-[methyl-3H] methionine ([methyl-3H]SAM). The partially purified enzyme (Superdex 75 eluate) was found to be characteristically affected by GTP gamma S, being activated about 40-fold in 2 mM, in contrast to ATP which did not show any effect on enzyme activity. Meanwhile, the enzyme was found to be markedly inhibited by AFC, reaching 0 activity in 2 mM. These observations strongly suggested that the partially purified enzyme was PFCCMT.
Assuntos
Encéfalo/enzimologia , Proteínas Metiltransferases/isolamento & purificação , Acetilcisteína/análogos & derivados , Acetilcisteína/farmacologia , Animais , Bovinos , Cromatografia Líquida , Guanosina 5'-O-(3-Tiotrifosfato)/farmacologia , Peso Molecular , Proteínas Metiltransferases/química , Processamento de Proteína Pós-TraducionalRESUMO
DNA topoisomerase II copurifies with and is phosphorylated by protein kinase CKII. In this study, a yeast two-hybrid system was used to investigate the interaction between human topoisomerase II isozymes and CKII subunits. The two-hybrid test clearly showed that both topoisomerase IIalpha and IIbeta interact with the CKIIbeta, but not the CKIIalpha subunit. The two-hybrid test also demonstrated that topoisomerase IIbeta residues 1099-1263 and topoisomerase IIalpha residues 1078-1182 mediate the interaction with the CKIIbeta subunit, providing evidence that the leucine zipper motif and the major CKII-dependent phosphorylation sites of topoisomerase II are unnecessary for its physical binding to CKIIbeta. Furthermore, a DNA relaxation assay demonstrated that the CKII subunit enhances topoisomerase II activity by physical interaction with topoisomerase II.
Assuntos
DNA Topoisomerases Tipo II/genética , DNA Topoisomerases Tipo II/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Sítios de Ligação/fisiologia , Caseína Quinase II , DNA/metabolismo , Ativação Enzimática/fisiologia , Deleção de Genes , Humanos , Zíper de Leucina/fisiologia , Fosforilação , Proteínas Serina-Treonina Quinases/química , Estrutura Terciária de Proteína , Técnicas do Sistema de Duplo-Híbrido , LevedurasRESUMO
We have developed a polymerase chain reaction (PCR)-based assay that could effectively reduce the time period required to screen and select the cold tolerance gene of rice seedlings under field conditions. The two specific random amplified polymorphic DNA (RAPD) fragments for the assay were identified on the basis of quantitative trait loci (QTL) analysis which were found to be tightly linked to cold sensitivity. The two RAPD fragments, OPT8(600) in the cold sensitivity rice cultivar 'Dular (indica)' and OPU20(1200) in the resistance rice cultivar 'Toyohatamochi (japonica)', were identified after screening 11 RAPD fragments using 2 random primers on the genomic DNAs of 'Dular' and 'Toyohatamochi'. These primers, when used in a multiplexed PCR, specifically amplified a 0.6 kb and a 1.2 kb fragment in the sensitive and resistant rice cultivars, respectively. When this assay was performed on the genomic DNAs of 16 japonica, 3 Tongil (indica/ japonica), and 2 indica rice cultivars, the primers amplified a 0.6 kb fragment in all of the cold sensitivity rice cultivars or 1.2 kb fragment in all of the resistance ones. These markers can be of potential use in the marker-assisted selection (MAS) for cold tolerance in rice seedling. As screening for resistance can now be conducted independent of the availability of low temperature, the breeding of cold tolerance cultivars can be hastened.
Assuntos
Oryza/crescimento & desenvolvimento , Oryza/genética , Técnica de Amplificação ao Acaso de DNA Polimórfico , Aclimatação/genética , Sequência de Bases , Temperatura Baixa , Primers do DNA/genética , DNA de Plantas/genética , Marcadores Genéticos , Oryza/fisiologia , Polimorfismo GenéticoRESUMO
We present the first application of switched angle spinning (SAS) to correlate the first-order dipolar spectrum of a liquid crystalline sample with the isotropic magic angle spinning (MAS) spectrum in a two-dimensional experiment. In this experiment we are able to select the degree of dipolar couplings introduced via mechanical manipulations of the liquid crystal director in a single oriented sample. The (19)F SAS-COSY correlation of iodotrifluoroethylene, an AMX spin system, dissolved in the nematic liquid crystal 4-octylphenyl-2-chloro-4-(4-heptylbenzoyloxy)-benzoate provides assignment of both the J and dipolar couplings in a single experiment. This work demonstrates the use of oriented samples and sample spinning to resolve homonuclear dipolar couplings using isotropic chemical shifts.
RESUMO
Enzymatic methylation of endogenous proteins in several cancer cell lines was investigated to understand a possible relationship between protein-arginine methylation and cellular proliferation. Cytosolic extracts prepared from several cancer cells (HeLa, HCT-48, A549, and HepG2) and incubated with S-adenosyl-L-[methyl-3H]methionine revealed an intensely [methyl-3H]-labeled 20-kDa polypeptide. On the other hand, cytosolic extracts prepared from normal colon cells did not show any methylation of the 20-kDa protein under identical conditions. To identify nature of the 20-kDa polypeptide, purified histones were methylated with HCT-48 cytosolic extracts and analyzed by SDS-PAGE. However, none of the histones comigrated with the methylated 20-kDa polypeptide, indicating that it is unlikely to be any of the histone subclasses. The [methyl-3H]group in the 20-kDa polypeptide was stable at pH 10-11 (37 degrees C for 30 min) and methylation was not stimulated by GTPgammaS (4 mM), thus the reaction is neither carboxyl methylesterification on isoaspartyl residues, nor on C-terminal farnesylated cysteine. The present study together with the previous identification of N(G)-methylated arginine residues in the HCT-48 cytosol fraction suggests that this novel endogenous 20-kDa arginine-methylation is a cellular proliferation-related posttranslational modification reaction.