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1.
Arch. argent. pediatr ; 116(6): 773-777, dic. 2018. ilus, graf
Article in Spanish | LILACS, BINACIS | ID: biblio-973696

ABSTRACT

El síndrome de Sjogren-Larsson se caracteriza por retardo mental, ictiosis congènita y diplejía o cuadriplejía espástica. El defecto primario en este síndrome es la mutación del gen ALDH3A2, que codifica la enzima aldehído deshidrogenasa grasa y causa una deficiencia enzimática que produce una acumulación de alcoholes y aldehídos grasos en los tejidos que comprometen la integridad de la membrana celular, cuyos efectos pueden observarse en la piel, los ojos y el sistema nervioso central. El diagnóstico se realiza por medio de la cuantificación de la actividad de la enzima. Se describe el caso de una paciente con signos clínicos patognomónicos del síndrome de Sjogren-Larsson, cuyo diagnóstico se realizó por medio de la cuantificación de la actividad enzimática en un cultivo de fibroblastos. Además, tomando en cuenta el árbol genealógico de la paciente, se realizó el estudio en los padres y un hermano con signos sugestivos del síndrome de Sjogren-Larsson.


Sjogren-Larsson syndrome is characterized by congenital ichthyosis, mental retardation and spastic diplegia or quadriplegia. The primary defect in this syndrome is mutation of ALDH3A2 gen that codes for the fatty aldehyde dehydrogenase. Deficiency of this enzyme causes an accumulation of fatty alcohols and fatty aldehydes, leading to altered cell-membrane integrity. Skin, eyes, and the central nervous system are affected latter. The diagnosis is carried out through the cuantification of the enzyme activity.


Subject(s)
Humans , Female , Child , Sjogren-Larsson Syndrome/diagnosis , Aldehyde Oxidoreductases/genetics , Sjogren-Larsson Syndrome/genetics , Fibroblasts/enzymology , Mutation
2.
Braz. oral res. (Online) ; 32: e114, 2018. tab, graf
Article in English | LILACS | ID: biblio-974440

ABSTRACT

Abstract Smokers have a risk of developing periodontal disease. Matrix metalloproteinases (MMP) play a significant role in periodontal tissue destruction. In this study possible relationship between smoking and gingival tissue expression of gelatinases in chronic periodontitis patients relative to periodontally healthy subjects was investigated. Forty chronic periodontitis patients (20 smokers and 20 non-smokers) and forty periodontally healthy subjects (20 smokers and 20 non-smokers) were enrolled. The clinical periodontal measurements recorded, and gingival tissues harvested after that. After histologic evaluation, matrix metalloproteinases -2 and -9 expressions were analyzed immunohistochemically. In nonsmokers, higher expression of metalloproteinases -2 and -9 detected in chronic periodontitis group compared to the periodontally healthy group. In the smoker chronic periodontitis group, the expression of metalloproteinases-2 was lower than nonsmoker chronic periodontitis group. Statistically significant differences detected between smoker and nonsmoker periodontally healthy groups in metalloproteinases-2 expression. For metalloproteinases-9 expression, smoker chronic periodontitis group has lower values than nonsmoker chronic periodontitis group. In periodontally healthy group smokers showed higher metalloproteinases -9 expressions than non- smokers. Present findings support the role of gelatinases in chronic periodontitis pathogenesis. Based on the current results we conclude that smoking alters the expression of gelatinases in gingival tissues.


Subject(s)
Humans , Male , Female , Adolescent , Adult , Young Adult , Smoking/adverse effects , Matrix Metalloproteinase 2/analysis , Matrix Metalloproteinase 9/analysis , Chronic Periodontitis/enzymology , Biopsy , Immunohistochemistry , Case-Control Studies , Cross-Sectional Studies , Analysis of Variance , Statistics, Nonparametric , Fibroblasts/enzymology , Gingiva/enzymology , Middle Aged
3.
Clin. biomed. res ; 37(4): 330-333, 2017. ilus, graf
Article in English | LILACS | ID: biblio-876699

ABSTRACT

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder caused by deficiency of alpha-L-iduronidase (IDUA). Limitations such as the need for weekly injections, high morbidity and mortality, and high cost of current treatments show that new approaches to treat this disease are required. In this study, we aimed to correct fibroblasts from a patient with MPS I using non-viral gene therapy. Using a plasmid encoding the human IDUA cDNA, we achieved stable high IDUA levels in transfected fibroblasts up to 6 months of treatment. These results serve as proof of concept that a non-viral approach can correct the enzyme deficiency in cells of patients with lysosomal storage disorders, which can be used as a research tool for a series of disease aspects. Future studies should focus on showing if this approach can be useful in small animals and clinical trials (AU)


Subject(s)
Humans , Fibroblasts/enzymology , Gene Transfer Techniques , Genetic Vectors , Iduronidase/metabolism , Mucopolysaccharidosis I/therapy , DNA, Complementary , Genetic Therapy/methods , Iduronidase/genetics , Mucopolysaccharidosis I/genetics , Plasmids/genetics , Transfection/methods
4.
Biol. Res ; 49: 1-9, 2016. tab
Article in English | LILACS | ID: lil-774431

ABSTRACT

BACKGROUND: Coenzyme Q10 (CoQ10 or ubiquinone) deficiency can be due either to mutations in genes involved in CoQ10 biosynthesis pathway, or to mutations in genes unrelated to CoQ10 biosynthesis. CoQ10 defect is the only oxidative phosphorylation disorder that can be clinically improved after oral CoQ10 supplementation. Thus, early diagnosis, first evoked by mitochondrial respiratory chain (MRC) spectrophotometric analysis, then confirmed by direct measurement of CoQ10 levels, is of critical importance to prevent irreversible damage in organs such as the kidney and the central nervous system. It is widely reported that CoQ10 deficient patients present decreased quinone-dependent activities (segments I + III or G3P + III and II + III) while MRC activities of complexes I, II, III, IV and V are normal. We previously suggested that CoQ10 defect may be associated with a deficiency of CoQ10-independent MRC complexes. The aim of this study was to verify this hypothesis in order to improve the diagnosis of this disease. RESULTS: To determine whether CoQ10 defect could be associated with MRC deficiency, we quantified CoQ10 by LC-MSMS in a cohort of 18 patients presenting CoQ10-dependent deficiency associated with MRC defect. We found decreased levels of CoQ10 in eight patients out of 18 (45 %), thus confirming CoQ10 disease. CONCLUSIONS: Our study shows that CoQ10 defect can be associated with MRC deficiency. This could be of major importance in clinical practice for the diagnosis of a disease that can be improved by CoQ10 supplementation.


Subject(s)
Adolescent , Adult , Aged , Child , Child, Preschool , Female , Humans , Infant , Male , Middle Aged , Young Adult , Ataxia/genetics , Electron Transport/genetics , Mutation , Mitochondrial Diseases/genetics , Muscle Weakness/genetics , Ubiquinone/analogs & derivatives , Ubiquinone/deficiency , Ataxia/diagnosis , Ataxia/metabolism , Biopsy , Cells, Cultured , Chromatography, Liquid , Fibroblasts/enzymology , Mitochondrial Diseases/diagnosis , Mitochondrial Diseases/metabolism , Muscle Weakness/diagnosis , Muscle Weakness/metabolism , Muscles/pathology , Spectrophotometry/methods , Tandem Mass Spectrometry/methods , Ubiquinone/biosynthesis , Ubiquinone/genetics , Ubiquinone/metabolism
5.
Salud colect ; 11(1): 9-21, ene.-mar. 2015.
Article in Spanish | LILACS | ID: lil-746681

ABSTRACT

El fortalecimiento mundial de la patente farmacéutica tensiona el acceso a los medicamentos esenciales. De acuerdo a este trabajo ello ha derivado en la colisión del derecho de propiedad intelectual que impulsa el Acuerdo sobre los Aspectos de los Derechos de Propiedad Intelectual relacionados con el Comercio (ADPIC) de la Organización Mundial del Comercio (OMC), con el derecho a la salud previsto en el Pacto Internacional de Derechos Económicos, Sociales y Culturales (PIDESC). Diversas controversias ventiladas en la OMC ilustran el enfrentamiento entre países con una poderosa industria farmacéutica y los intereses de países en desarrollo. Se concluye que las normas ADPIC-plus suscritas en tratados de libre comercio por países en desarrollo, que confieren al titular de la patente farmacéutica más derechos que los previstos en el propio Acuerdo sobre los ADPIC, son incompatibles con las obligaciones que asumen respecto del acceso a medicamentos esenciales en el marco del derecho a la salud del PIDESC.


The strengthening of pharmaceutical patent protection globally puts strains on access to essential medicines. According to the present paper, this process has led to the collision of the intellectual property rights adopted in the World Trade Organization (WTO) Trade-Related Aspects of Intellectual Property Rights (TRIPS) Agreement and the right to health stated in the International Covenant on Economic, Social and Cultural Rights (ICESCR). Several controversies disputed in the WTO illustrate the confrontation between countries with a powerful pharmaceutical industry and the interests of developing countries. It is concluded that the TRIPS-plus rules subscribed to by developing countries in free trade agreements which give the pharmaceutical patent holder more rights than those stipulated in the original TRIPS Agreement are incompatible with the obligations to provide access to essential medicines under the right to health of the ICESCR.


Subject(s)
Animals , Cricetinae , Humans , Mice , Genetic Vectors , Poly(ADP-ribose) Polymerases/genetics , RNA Interference , Cloning, Molecular , DNA , Fibroblasts/enzymology , Gene Targeting , Mice, Knockout , Poly(ADP-ribose) Polymerases/analysis , Skin/cytology
6.
Biol. Res ; 48: 1-8, 2015. graf
Article in English | LILACS | ID: biblio-950773

ABSTRACT

BACKGROUND: Various health benefits have been attributed to Er-Miao-San (EMS), a traditional Chinese herbal formulation that contains equal amounts of cortex phellodendri (Phellodendron amurense Ruprecht) and rhizoma atractylodis (Atractylodes lancea D.C). However, its effect on the anti-inflammatory activity in human dermal fibroblasts (HDFs) and the mechanism underlying this effect are unknown. RESULTS: This study investigated the effects of EMS on TNF-α-induced MMP-1 expression in HDFs. Our data show that EMS inhibited TNF-α-induced MMP-1 expression in a concentration-dependent manner. Interestingly, EMS maintained IkB content without inhibiting the phosphorylation of MAPKs, which are well-established upstream kinases of NF-kB. Moreover, EMS reduced the level of nuclear p65 protein in HDFs. Luciferase assay revealed that EMS inhibits the transcriptional activity of NF-kBbystabilizing IkB. Our results show that EMS exerts its anti-inflammatory effect by inhibiting NF-kB-regulated genes such as IL-1ß and IL-8. Moreover, EMS effectively inhibited TNF-α-induced expression of MMP-1 via the NF-kBpathway. CONCLUSIONS: Taken together, our data suggest that EMS could potentially be used as an anti-inflammatory and anti-aging treatment.


Subject(s)
Humans , Aging/drug effects , Drugs, Chinese Herbal/pharmacology , Plant Extracts/pharmacology , Dermis/cytology , Matrix Metalloproteinase 1/biosynthesis , Fibroblasts/drug effects , Signal Transduction/drug effects , Cell Line , Cell Survival/drug effects , Enzyme Induction/drug effects , Gene Expression Regulation/drug effects , Interleukin-8/drug effects , Interleukin-8/metabolism , NF-kappa B/metabolism , Tumor Necrosis Factor-alpha/metabolism , Mitogen-Activated Protein Kinases/drug effects , Interleukin-1beta/drug effects , Interleukin-1beta/metabolism , Real-Time Polymerase Chain Reaction , Fibroblasts/enzymology , Anti-Inflammatory Agents/administration & dosage
7.
Salud pública Méx ; 56(6): 631-637, nov.-dic. 2014.
Article in Spanish | LILACS | ID: lil-733342

ABSTRACT

Objetivo. Analizar la percepción que el prestador de servicios de salud y el adulto mayor (AM) tienen sobre el maltrato al AM en los servicios públicos de salud, en ciudades seleccionadas de México. Material y métodos. De 2009 a 2012 se realizó un estudio con diseño cualitativo y estrategia de triangulación de fuentes de datos; se efectuaron entrevistas semiestructuradas a 13 prestadores y a 12 ancianos para recuperar su experiencia en el tema. El análisis utilizó procedimientos de la Teoría Fundamentada. Resultados. El maltrato contra el AM es una práctica naturalizada por el personal y por el anciano, la cual se manifiesta de formas diversas. Conclusiones. La institucionalización, profesionalización histórica y falta de conciencia sobre las necesidades de los AM demandan cambios de planeación, organización y supervisión del Sistema de Salud. El personal requiere intervenciones de formación, capacitación y cambio de actitudes/comportamiento, para otorgar atención integral, digna, humana y de respeto a los Derechos Humanos de los AM.


Objective. To analyze the health care providers (HCP) and elderly patients' perceptions about abuse of the elderly by health personnel of public health services, in selected cities in Mexico. Materials and methods. A qualitative study and a strategy of data triangulation were performed during 2009 and 2012; 13 HCPs and 12 elders were interviewed, in order to obtain their experience regarding elder abuse. Grounded Theory proceedings were used for the analysis. Results. Elder abuse is a naturalized practice, from HCP and elderly people's point of view; these perceptions are showed in different ways. Conclusion. Institutionalization, historical professionalization and lack of consciousness about needs of the elderly (sociocultural and economic), require changes in planning, organization and monitoring process in the Health System; training and educational interventions on staff and exchange attitudes and behavior are necessary in order to offer a health care that is comprehensive, decent, human and with respect for the human rights.


Subject(s)
Animals , Female , Humans , Mice , Antimetabolites, Antineoplastic/pharmacology , Cyclins/metabolism , Enzyme Inhibitors/metabolism , Phenylacetates/pharmacology , Antisense Elements (Genetics) , Breast Neoplasms , Calcium-Calmodulin-Dependent Protein Kinases/metabolism , Cell Division/drug effects , Cyclin-Dependent Kinases/genetics , Cyclin-Dependent Kinases/metabolism , Cyclins/genetics , Fibroblasts/cytology , Fibroblasts/drug effects , Fibroblasts/enzymology , Gene Expression Regulation, Neoplastic/physiology , Mice, Knockout , Phosphorylation , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , RNA, Messenger/metabolism , Retinoblastoma Protein/metabolism , Signal Transduction/physiology , Tumor Cells, Cultured/cytology , Tumor Cells, Cultured/drug effects , Tumor Cells, Cultured/enzymology , Up-Regulation/drug effects
8.
Gac. méd. Caracas ; 122(1): 25-28, ene.-mar. 2014. graf
Article in Spanish | LILACS | ID: lil-772736

ABSTRACT

Se reseña la publicación de un importante trabajo donde se reporta el éxito obtenido por científicos chinos utilizando sustancias de bajo tamaño molecular, que actúan como reguladores de enzimas y de procesos bioquímicos de señalización, en la reprogramación de células diferenciadas de ratón para convertirlas en células madre pluripotentes similares a las células madre embrionarias y a las células madre pluripotentes inducidas por genes de factores de transcripción. Se hace referencia a las publicaciones previas de los investigadores que lograron por primera vez el mismo propósito con diferentes métodos. Se comentan las ventajas del nuevo método.


This is a review of an important publication by Chinese scientists about the successful reprogramming of differentiated murine cells to pluripotent stem cells that resemble embryonic stem cells using small-molecule compounds that act as regulators of enzymes and signaling pathways. Reference is made of previous publications by researchers who achieved for the first time the same goal by different methods. Comments on the advantages of the new method are included.


Subject(s)
Animals , Mice , Induced Pluripotent Stem Cells/chemistry , Fibroblasts/enzymology , Fibroblasts/chemistry , Cellular Reprogramming , Valproic Acid/administration & dosage , China
9.
Acta cir. bras ; 28(3): 216-220, Mar. 2013. ilus
Article in English | LILACS | ID: lil-667933

ABSTRACT

PURPOSE: To describe a method to characterize the gelatinase activity of cultured human periodontal fibroblasts stimulated with Pam3Cys and E. coli LPS, ligands of TLR2 and TLR4 respectively, and by centrifugation of the cultures, simulating an orthodontic force. METHODS: To study MMP-2 activity, primary cultures of human periodontal fibroblasts were stimulated with the addition of TLRs 2 and 4 ligands and the application of mechanical force by centrifugation at 141 x g for 30 min. Supernatant media was collected 24 hours later to perform protein quantification and zymography. RESULTS: MMP-2 activity suffered an increase in cultures co-stimulated with TLRs 2 and 4 ligands alone or with the presence of mechanical force application compared to basal levels. CONCLUSION: Zymography, one of the several methods to study MMPs activities, is a simple, qualitative and efficient method based on electrophoresis of bis-acrylamide gels copolymerized with a protein substrate.


Subject(s)
Humans , Electrophoresis/methods , Fibroblasts/enzymology , /analysis , Cell Survival , Cells, Cultured , Gelatinases/physiology , Lipoproteins , /physiology , Periodontal Ligament/cytology , Reproducibility of Results , Statistics, Nonparametric , Time Factors , Toll-Like Receptors/analysis
10.
Article in English | WPRIM | ID: wpr-35840

ABSTRACT

Apoptosis has an important role in maintaining tissue homeostasis in cellular stress responses such as inflammation, endoplasmic reticulum stress, and oxidative stress. T-cell death-associated gene 51 (TDAG51) is a member of the pleckstrin homology-like domain family and was first identified as a pro-apoptotic gene in T-cell receptor-mediated cell death. However, its pro-apoptotic function remains controversial. In this study, we investigated the role of TDAG51 in oxidative stress-induced apoptotic cell death in mouse embryonic fibroblasts (MEFs). TDAG51 expression was highly increased by oxidative stress responses. In response to oxidative stress, the production of intracellular reactive oxygen species was significantly enhanced in TDAG51-deficient MEFs, resulting in the activation of caspase-3. Thus, TDAG51 deficiency promotes apoptotic cell death in MEFs, and these results indicate that TDAG51 has a protective role in oxidative stress-induced cell death in MEFs.


Subject(s)
Animals , Apoptosis , Embryo, Mammalian/cytology , Fibroblasts/enzymology , Gene Expression Regulation , Intracellular Space/metabolism , Mice , Mitogen-Activated Protein Kinases/metabolism , NF-kappa B/metabolism , Oxidative Stress/genetics , Reactive Oxygen Species/metabolism , Signal Transduction , Transcription Factors/deficiency
11.
Article in English | LILACS | ID: lil-582934

ABSTRACT

The lip vermillion constitutes a transition tissue, between oral mucosa and skin, where oral mucosal cells from epithelial and connective tissue compartments are exposed to ultraviolet (UV) sunlight. Fibroblasts are abundant resident cells of the connective tissue which are key regulators of extracellular matrix composition, as well as, epithelial and endothelial cell function. UVB light, an inherent component of sunlight, causes several alterations in skin fibroblasts, including premature senescence and increased cyclooxygenase (COX)-2 expression. To assess if UVB irradiation had similar effects on fibroblasts derived from human oral mucosa (HOM), primary cultures of HOM fibroblasts were irradiated with a single dose of 30 or 60 mJ/cm²of UVB light or sham-irradiated. Fibroblast proliferation was assessed from 3 to 48 hrs after UVB-irradiation utilizing [³H]-thymidine incorporation and MTT assays. In addition, COX-2 mRNA expression was detected by RT-PCR, and PGE2 production was assessed using enzyme immunoassay from 0.5 to 24 hrs after UVB-irradiation. The results showed a significant decrease in proliferation of UVB-irradiated HOM fibroblasts as compared to controls as measured by both [³H]-thymidine incorporation and MTT assays (p<0.001). HOM fibroblasts had increased COX-2 mRNA expression at 0.5 and 12 hrs after irradiation, and PGE2 production was elevated at 12 and 24 hrs post-irradiation as compared to controls (p<0.05). The results showed an inhibitory effect of a single dose of UVB irradiation on HOM fibroblast proliferation with an increase in COX-2 expression and activation. Therefore, photodamaged fibroblasts may play and important role in the pathogenesis of UV-induced lesions of the lip.


Subject(s)
Humans , /radiation effects , Fibroblasts/radiation effects , Mouth Mucosa/cytology , Ultraviolet Rays , Fibroblasts/enzymology , Mouth Mucosa/radiation effects , Mouth Mucosa/enzymology , Cell Proliferation , Reverse Transcriptase Polymerase Chain Reaction
12.
Braz. j. med. biol. res ; 42(1): 44-52, Jan. 2009. ilus
Article in English | LILACS | ID: lil-505421

ABSTRACT

Focal adhesion kinase (FAK) is a broadly expressed tyrosine kinase implicated in cellular functions such as migration, growth and survival. Emerging data support a role for FAK in cardiac development, reactive hypertrophy and failure. Data reviewed here indicate that FAK plays a critical role at the cellular level in the responses of cardiomyocytes and cardiac fibroblasts to biomechanical stress and to hypertrophic agonists such as angiotensin II and endothelin. The signaling mechanisms regulated by FAK are discussed to provide insight into its role in the pathophysiology of cardiac hypertrophy and failure.


Subject(s)
Animals , Humans , Cardiomegaly/enzymology , Fibroblasts/enzymology , Focal Adhesion Protein-Tyrosine Kinases/physiology , Heart Failure/enzymology , Myocytes, Cardiac/enzymology , Cell Proliferation , Cardiomegaly/etiology , Cardiomegaly/physiopathology , Heart Failure/etiology , Heart Failure/physiopathology , Signal Transduction/physiology
13.
Arq. bras. endocrinol. metab ; 52(8): 1244-1251, Nov. 2008. ilus, graf, tab
Article in English | LILACS | ID: lil-503289

ABSTRACT

Mutations in the vitamin D receptor (VDR) are associated to the hereditary 1,25-dihydroxivitamin D-resistant rickets. The objectives of this work are: search for mutations in the VDR and analyze their functional consequences in four Brazilian children presented with rickets and alopecia. The coding region of the VDR was amplified by PCR e direct sequenced. We identified three mutations: two patients had the same mutation in exon 7 at aminoacid position 259 (p.Q259E); one patient had a mutation in exon 8 at codon 319 (p.G319V) and another one had a mutation in exon 3 leading to a truncated protein at position 73 (p.R73X). Functional studies of the mutant receptors of fibroblast primary culture, from patients' skin biopsy treated with increasing doses of 1,25(OH)2 vitamin D showed that VDR mutants were unable to be properly activated and presented a reduction in 24-hydroxylase expression level.


Mutações no receptor de vitamina D (VDR) são associadas a raquitismo hereditário resistente a 1,25-dihidroxivitamina D. Os objetivos deste trabalho foram procurar mutações no VDR e analisar suas conseqüências funcionais em quatro pacientes com raquitismo e alopécia. A região codificadora do VDR foi amplificada por PCR e seqüenciada diretamente. Identificamos três mutações: dois pacientes apresentavam a mesma mutação no éxon 7 na posição protéica 259 (p.Q259E); um paciente apresentava uma mutação no éxon 8 no códon 319 (p.G319V) e o outro apresentava uma mutação no exon 3 resultando em uma proteína truncada na posição 73 (p.R73X). O estudo funcional dos receptores mutados nos extratos de culturas de fibroblasto primárias obtidas de biópsia de pele dos pacientes, tratados com doses crescentes de 1,25(OH)2 vitamina D demonstraram que os receptores mutantes não apresentam ativação adequada apresentando expressão reduzida de 24-hidroxilase.


Subject(s)
Child , Female , Humans , Male , Young Adult , Alopecia/genetics , Familial Hypophosphatemic Rickets/genetics , Mutation , Receptors, Calcitriol/genetics , Alopecia/drug therapy , Base Sequence , Calcitriol/therapeutic use , Fibroblasts/drug effects , Fibroblasts/enzymology , Familial Hypophosphatemic Rickets/drug therapy , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Calcitriol/metabolism , Sequence Analysis, DNA , Steroid Hydroxylases/genetics , Steroid Hydroxylases/metabolism , Vitamins/therapeutic use , Young Adult
14.
Indian J Ophthalmol ; 2008 Jul-Aug; 56(4): 279-83
Article in English | IMSEAR | ID: sea-69912

ABSTRACT

BACKGROUND: Pterygia are common, benign, fibrovascular, and infiltrative processes of the corneo-conjunctival junction of unknown pathogenesis. Cyclooxygenase-2 (COX-2) mediates the rate-limiting step in arachidonic acid metabolism. Extensive evidence indicates that the COX-2 prostanoid pathway is involved in inflammation. The aim of the study was to document the immunohistochemical expression of COX-2 in primary and recurrent pterygia. MATERIALS AND METHODS: In this study, 21 primary pterygia and 12 recurrent pterygia from subjects undergoing pterygium surgery and six normal corneal-scleral tissue specimens were studied immunohistochemically for COX-2 expression. RESULTS: COX-2 was expressed in primary pterygia and recurrent pterygia specimens. There was a statistically significant difference in COX-2 expressions in fibroblasts between primary and recurrent pterygium cases ( P = 0.001). There were statistically significant differences in COX-2 expressions in surface epithelium ( P = 0.028) and stromal inflammatory cells ( P =0.000) between control tissues and primary pterygia tissues. We also detected statistically significant differences in COX-2 expressions in surface epithelium ( P =0.000), stromal fibroblasts P =0.000 (stromal fibroblasts and inflammatory cells), vessels ( P = 0.027) and inflammatory cells ( P =0.001) between control tissues and recurrent pterygia tissues. CONCLUSIONS: This is the first study to document the expression of COX-2 in primary and recurrent pterygia. In our opinion after excision of pterygia, fibroblastic proliferation continues and this contributes to recurrence.


Subject(s)
Adult , Aged , Aged, 80 and over , Cyclooxygenase 2/metabolism , Epithelium/enzymology , Female , Fibroblasts/enzymology , Humans , Immunoenzyme Techniques , Male , Middle Aged , Pterygium/enzymology , Recurrence
15.
Braz. j. med. biol. res ; 41(4): 283-288, Apr. 2008. graf
Article in English | LILACS | ID: lil-479675

ABSTRACT

GM1 gangliosidosis is an autosomal recessive disorder caused by the deficiency of lysosomal acid hydrolase ß-galactosidase (ß-Gal). It is one of the most frequent lysosomal storage disorders in Brazil, with an estimated frequency of 1:17,000. The enzyme is secreted and can be captured by deficient cells and targeted to the lysosomes. There is no effective treatment for GM1 gangliosidosis. To determine the efficiency of an expression vector for correcting the genetic defect of GM1 gangliosidosis, we tested transfer of the ß-Gal gene (Glb1) to fibroblasts in culture using liposomes. ß-Gal cDNA was cloned into the expression vectors pSCTOP and pREP9. Transfection was performed using 4 µL lipofectamine 2000 and 1.5-2.0 µg DNA. Cells (2 x 10(5)/well) were harvested 24 h, 48 h, and 7 days after transfection. Enzyme specific activity was measured in cell lysate and supernatant by fluorometric assay. Twenty-four hours after transfection, treated cells showed a higher enzyme specific activity (pREP9-ß-Gal: 621.5 ± 323.0, pSCTOP-ß-Gal: 714.5 ± 349.5, pREP9-ß-Gal + pSCTOP-ß-Gal: 1859.0 ± 182.4, and pREP9-ß-Gal + pTRACER: 979.5 ± 254.9 nmol·h-1·mg-1 protein) compared to untreated cells (18.0 ± 3.1 for cell and 32.2 ± 22.2 nmol·h-1·mg-1 protein for supernatant). However, cells maintained in culture for 7 days showed values similar to those of untreated patients. In the present study, we were able to transfect primary patients' skin fibroblasts in culture using a non-viral vector which overexpresses the ß-Gal gene for 24 h. This is the first attempt to correct fibroblasts from patients with GM1 gangliosidosis by gene therapy using a non-viral vector.


Subject(s)
Humans , Fibroblasts/enzymology , Genetic Vectors , Gangliosidosis, GM1/enzymology , Transfection/methods , beta-Galactosidase/metabolism , DNA, Complementary , Fluorometry , Gangliosidosis, GM1/therapy , Liposomes , Plasmids/genetics , beta-Galactosidase/genetics
16.
Article in English | WPRIM | ID: wpr-177643

ABSTRACT

During radiotherapy of cancer, neighboring normal cells may receive sub-lethal doses of radiation. To investigate whether such low levels of radiation modulate normal cell responses to death stimuli, primary cultured human fibroblasts were exposed to various doses of gamma-rays. Analysis of cell viability using an exclusion dye propidium iodide revealed that the irradiation up to 10 Gy killed the fibroblasts only to a minimal extent. In contrast, the cells efficiently lost their viability when exposed to 0.5-0.65 mM H2O2. This type of cell death was accompanied by JNK activation, and was reversed by the use of a JNK-specific inhibitor SP600125. Interestingly, H2O2 failed to kill the fibroblasts when these cells were pre-irradiated, 24 h before H2O2 treatment, with 0.25-0.5 Gy of gamma-rays. These cytoprotective doses of gamma-rays did not enhance cellular capacity to degrade H2O2, but elevated cellular levels of p21Cip/WAF1, a p53 target that can suppress H2O2-induced cell death by blocking JNK activation. Consistently, H2O2-induced JNK activation was dramatically suppressed in the pre-irradiated cells. The overall data suggests that ionizing radiation can impart normal fibroblasts with a survival advantage against oxidative stress by blocking the process leading to JNK activation.


Subject(s)
Antioxidants/pharmacology , Cell Death , Cells, Cultured , Enzyme Activation/radiation effects , Fibroblasts/enzymology , Gamma Rays , Heat-Shock Proteins/metabolism , Humans , JNK Mitogen-Activated Protein Kinases/antagonists & inhibitors , Oxidative Stress/radiation effects , Water/pharmacology
17.
Article in English | WPRIM | ID: wpr-226076

ABSTRACT

Hepatitis C Virus (HCV) is associated with a severe liver disease and increased frequency in the development of hepatocellular carcinoma. Overexpression of HCV core protein is known to transform fibroblast cells. Phospholipase D (PLD) activity is commonly elevated in response to mitogenic signals, and has also been overexpressed and hyperactivated in some human cancer cells. The aim of this study was to understand how PLD was regulated in the HCV core protein-transformed NIH3T3 mouse fibroblast cells. We observed that PLD activity was elevated in the NIH3T3 cells overexpressing HCV core protein over the vector alone-transfected control cells, however, expression levels of PLD protein and protein kinase C (PKC) in the HCV core protein-transformed cells was similar to the control cells. Phorbol 12-myristate 13-acetate (PMA), which is known to activate PKC, stimulated PLD activity significantly more in the core protein-transformed cells, in comparison with that of the control cells. PLD activity assay using PKC isozyme-specific inhibitor and PKC translocation experiment showed that PKC-delta was mainly involved in the PMA- induced PLD activation in the core-transformed cells. Moreover, in cells overexpressing HCV core protein, PMA also stimulated p38 kinase more potently than that of the control cells, and an inhibitor of p38 kinase abolished PMA-induced PLD activation in cells overexpressing HCV core protein. Taken together, these results suggest that PLD might be implicated in core protein-induced transformation.


Subject(s)
Animals , Cell Line, Transformed , Cell Transformation, Viral , Fibroblasts/enzymology , Hepacivirus/genetics , Mice , NIH 3T3 Cells , Phospholipase D/metabolism , Protein Kinase C/antagonists & inhibitors , Protein Transport/drug effects , Tetradecanoylphorbol Acetate/analogs & derivatives , Transfection , Up-Regulation , Viral Core Proteins/genetics , p38 Mitogen-Activated Protein Kinases/physiology
18.
J Biosci ; 2000 Mar; 25(1): 33-40
Article in English | IMSEAR | ID: sea-110721

ABSTRACT

PTP-S2 is a tyrosine specific protein phosphatase that binds to DNA and is localized to the nucleus in association with chromatin. It plays a role in the regulation of cell proliferation. Here we show that the subcellular distribution of this protein changes during cell division. While PTP-S2 was localized exclusively to the nucleus in interphase cells, during metaphase and anaphase it was distributed throughout the cytoplasm and excluded from condensed chromosomes. At telophase PTP-S2 began to associate with chromosomes and at cytokinesis it was associated with chromatin in the newly formed nucleus. It was hyperphosphorylated and showed retarded mobility in cells arrested in metaphase. In vitro experiments showed that it was phosphorylated by CK2 resulting in mobility shift. Using a deletion mutant we found that CK2 phosphorylated PTP-S2 in the C-terminal non-catalytic domain. A heparin sensitive kinase from mitotic cell extracts phosphorylated PTP-S2 resulting in mobility shift. These results are consistent with the suggestion that during metaphase PTP-S2 is phosphorylated (possibly by CK2 or a CK2-like enzyme), resulting in its dissociation from chromatin.


Subject(s)
Amino Acid Sequence , Animals , Casein Kinase II , Catalytic Domain , Cell Line , Cell Nucleus/enzymology , Chromatin/enzymology , Chromosomes/enzymology , Fibroblasts/enzymology , HeLa Cells , Humans , Isoenzymes/metabolism , Microscopy, Confocal , Mitosis , Molecular Sequence Data , Phosphorylation , Protein Tyrosine Phosphatases/metabolism , Protein Serine-Threonine Kinases/metabolism , Rats
19.
Rev. bras. clín. ter ; 25(2): 88-92, mar. 1999. tab
Article in English | LILACS | ID: lil-252908

ABSTRACT

A Síndrome de Sjögren-Larsson é uma doença autossômica recessiva rara com distribuiçäo universal. Consiste em ictiose, displegia espástica e retardo mental causado por um defeito enzimático na oxidaçäo do álcool-graxo. Nós relatamos dois casos e fazemos uma revisäo da literatura respectiva. As duas crianças tinham atividade deficiente da NAD oxidorredutase. Foram estudados os lípides de membrana das células plasmáticas e eritrócitos. Bons resultados foram obtidos em um dos pacientes quando submetido a dieta na infância precoce o que se correlacionou com diminuiçäo do álcool-graxo no plasma. Entretanto näo obtivemos melhora clínica no outro paciente cujo tratamento teve início tardio.Terapia com etretinato foi necessária para controlar os sintomas cutâneos neste segundo paciente.


Subject(s)
Humans , Infant , Child, Preschool , Infant, Newborn , Female , Male , Lipids/analysis , Alcohol Oxidoreductases/metabolism , Sjogren-Larsson Syndrome/diet therapy , Fatty Alcohols/metabolism , Fatty Acids, Essential , Fibroblasts/enzymology , Hospitalization , Ichthyosis/pathology , Intellectual Disability/pathology , Muscle Spasticity/pathology , Skin
20.
Rev. chil. pediatr ; 66(5): 265-9, sept.-oct. 1995. ilus
Article in Spanish | LILACS | ID: lil-164982

ABSTRACT

Se describen tres niños, uno varón, de 4, 6 y 2 años de edad, afectados de aciduria glutárica tipo I. Su desarrollo fue normal hasta la segunda mitad del primer año de vida, cuando sufrieron alteración de conciencia y convulsiones, seguidas de pérdida de las habilidades adquiridas, distonía y movimientos anormales. La tomografía axial y resonancia nuclear magnética de cerebro mostraron atrofia frontotemporal y de los núcleos caudados y putámenes. Habíagran cantidad de ácidos glutárico, 3-hidroxiglutárico y glutacónico en orina y actividad disminuida de la enzima glutaril CoA deshidrogenasa en cultivos de fibroblastos de los tres niños, confirmándose así el diagnóstico de esta afección metabólica


Subject(s)
Humans , Male , Female , Child, Preschool , Glutamic Acid/urine , Acidosis/urine , Amino Acid Metabolism, Inborn Errors/diagnosis , Oxidoreductases/deficiency , 3-Hydroxyacyl CoA Dehydrogenases/urine , Acidosis/diet therapy , Basal Ganglia Diseases/enzymology , Clinical Enzyme Tests , Dystonia/etiology , Encephalitis/etiology , Amino Acid Metabolism, Inborn Errors/diet therapy , Fibroblasts/enzymology , Psychomotor Disorders/etiology
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