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Article in English | WPRIM | ID: wpr-881035


Taurochenodeoxycholic acid (TCDCA) is one of the main effective components of bile acid, playing critical roles in apoptosis and immune responses through the TGR5 receptor. In this study, we reveal the interaction between TCDCA and TGR5 receptor in TGR5-knockdown H1299 cells and the regulation of inflammation via the cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA)-cAMP response element binding (CREB) signal pathway in NR8383 macrophages. In TGR5-knockdown H1299 cells, TCDCA significantly activated cAMP level via TGR5 receptor, indicating TCDCA can bind to TGR5; in NR8383 macrophages TCDCA increased cAMP content compared to treatment with the adenylate cyclase (AC) inhibitor SQ22536. Moreover, activated cAMP can significantly enhance gene expression and protein levels of its downstream proteins PKA and CREB compared with groups of inhibitors. Additionally, TCDCA decreased tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, IL-8 and IL-12 through nuclear factor kappa light chain enhancer of activated B cells (NF-κB) activity. PKA and CREB are primary regulators of anti-inflammatory and immune response. Our results thus demonstrate TCDCA plays an essential anti-inflammatory role via the signaling pathway of cAMP-PKA-CREB induced by TGR5 receptor.

Animals , Cell Line , Cyclic AMP/metabolism , Cyclic AMP Response Element-Binding Protein/metabolism , Cyclic AMP-Dependent Protein Kinases/metabolism , Cytokines/metabolism , Humans , Inflammation , Macrophages , Rats , Receptors, G-Protein-Coupled/metabolism , Signal Transduction/drug effects , Taurochenodeoxycholic Acid/pharmacology
Bol. latinoam. Caribe plantas med. aromát ; 19(6): 555-568, 2020. tab, ilus
Article in English | LILACS | ID: biblio-1284299


Despite the development of modern medicine, alternative medicine, which has not lost its timeliness, remains attractive for the treatment of various diseases. Glabridin, a major flavonoid of Glycyrrhiza glabra, is known for its antioxidant and anti-inflammatory activity. The aim of this study was: 1) to determine the possible protective role of glabridin against ischemia/reperfusion (I/R) injury of the intestine; 2) to evaluate the in vitrocontractile responses of ileum smooth muscles to acetylcholine after an intestinal I/R; and 3) to explain the underlying molecular mechanism of its effect. Rats were assigned to groups of six rats each; 1) I/R, 2) gla10, 3) gla20, 4) gla40, 5) N5-[imino(nitroamino)methyl]-L-ornithine, methyl ester monohydrochloride (L-NAME)+gla40, and 6) Sham group. The healing effect of glabridin was abolished by L-NAME. Glabridin did not cause contractility of the smooth muscles to acetylcholine-induced contractile responses in intestinal I/R. Yet, it increased to spontaneous basal activity.

A pesar del desarrollo de la medicina moderna, la medicina alternativa, sin perder su vigencia, sigue siendo atractiva para el tratamiento de varias enfermedades. Glabradina, el flavonoide mayoritario de Glycyrrhiza glabra, es conocido por su actividad antioxidante y antiinflamatoria. Los propósitos de este estudio fueron: 1) Determinar el posible rol protector de glabradina ante daños intestinales por isquemia/reperfusion (I/R) 2) Evaluar in vitrolas respuestas de contracción de los músculos lisos del ileum ante acetilcolina después de I/R intestinal; y 3) Explicar el mecanismo molecular subyacente de este efecto. Se asignaron grupos de seis ratas: 1) I/R, 2) gla10, 3) gla20, 4) gla40, 5) N5-[imino(nitroamino)metil]-L-ornithina, metil ester monohidrochloruro (L-NAME)+gla40, y 6) Grupo testigo. El efecto curativo de glabridina fue abolido por L-NAME. Glabridina no causó contracción en el músculo liso como respuesta acetilcolina-inducida I/R. Además, incrementa la actividad basal expontánea.

Animals , Rats , Phenols/administration & dosage , Reperfusion Injury/drug therapy , Cyclic AMP/metabolism , Glycyrrhiza , Isoflavones/administration & dosage , Phenols/pharmacology , Rats, Wistar , Cyclic AMP/analysis , Cyclic GMP/metabolism , Oxidative Stress/drug effects , NG-Nitroarginine Methyl Ester , Ileum/drug effects , Ileum/chemistry , Isoflavones/pharmacology , Malondialdehyde/analysis , Muscle, Smooth/drug effects
Braz. j. med. biol. res ; 49(7): e5285, 2016. tab, graf
Article in English | LILACS | ID: biblio-951689


Beta-adrenergic receptor (βAR)-dependent blood vessel relaxation is impaired in older animals and G protein activation has been suggested as the causative mechanism. Here, we investigated the role of βAR subtypes (β1AR, β2AR, and β3AR) and cAMP in maturation-dependent vasorelaxation impairment. Aortic rings from 15 Sprague-Dawley male rats (3 or 9 weeks old) were harvested and left intact or denuded of the endothelium. Vascular relaxation in aortic rings from younger and older groups was compared in the presence of βAR subtype agonists and antagonists along with cAMP and cGMP antagonists. Isolated aortic rings were used to evaluate relaxation responses, protein expression was evaluated by western blot or real time PCR, and metabolites were measured by ELISA. Expression of βAR subtypes and adenylyl cyclase was assessed, and cAMP activity was measured in vascular tissue from both groups. Isoproterenol- and BRL744-dependent relaxation in aortic rings with and without endothelium from 9-week-old rats was impaired compared with younger rats. The β1AR antagonist CGP20712A (10-7 M) did not affect isoproterenol or BRL744-dependent relaxation in arteries from either group. The β2AR antagonist ICI-118,551 (10-7 M) inhibited isoproterenol-dependent aortic relaxation in both groups. The β3AR antagonist SR59230A (10-7 M) inhibited isoproterenol- and BRL744-dependent aortic ring relaxation in younger but not in older rats. All βAR subtypes were expressed in both groups, although β3AR expression was lower in the older group. Adenylyl cyclase (SQ 22536) or protein kinase A (H89) inhibitors prevented isoproterenol-induced relaxation in younger but not in older rats. Production of cAMP was reduced in the older group. Adenylyl cyclase III and RyR3 protein expression was higher in the younger group. In conclusion, altered expression of β3AR and adenylyl cyclase III may be responsible for reduced cAMP production in the older group.

Animals , Male , Aorta, Thoracic/drug effects , Aorta, Thoracic/physiopathology , Vasodilation/drug effects , Vasodilation/physiology , Adrenergic beta-1 Receptor Antagonists/pharmacology , Adenylyl Cyclase Inhibitors/pharmacology , Aorta, Thoracic/physiology , Time Factors , Gene Expression , Adenylyl Cyclases/physiology , Blotting, Western , Age Factors , Cyclic AMP/analysis , Cyclic AMP/metabolism , Albuterol/pharmacology , Dobutamine/pharmacology
Clinics ; 70(3): 157-161, 03/2015. tab
Article in English | LILACS | ID: lil-747105


OBJECTIVE: To determine whether Tai Chi Chuan or ballroom dancing promotes better performance with respect to postural balance, gait, and postural transfer among elderly people. METHODS: We evaluated 76 elderly individuals who were divided into two groups: the Tai Chi Chuan Group and the Dance Group. The subjects were tested using the NeuroCom Balance Master¯ force platform system with the following protocols: static balance tests (the Modified Clinical Tests of Sensory Interaction on Balance and Unilateral Stance) and dynamic balance tests (the Walk Across Test and Sit-to-stand Transfer Test). RESULTS: In the Modified Clinical Test of Sensory Interaction on Balance, the Tai Chi Chuan Group presented a lower sway velocity on a firm surface with open and closed eyes, as well as on a foam surface with closed eyes. In the Modified Clinical Test of Sensory Interaction on Unilateral Stance, the Tai Chi Chuan Group presented a lower sway velocity with open eyes, whereas the Dance Group presented a lower sway velocity with closed eyes. In the Walk Across Test, the Tai Chi Chuan Group presented faster walking speeds than those of the Dance Group. In the Sit-to-stand Transfer Test, the Tai Chi Chuan Group presented shorter transfer times from the sitting to the standing position, with less sway in the final standing position. CONCLUSION: The elderly individuals who practiced Tai Chi Chuan had better bilateral balance with eyes open on both types of surfaces compared with the Dance Group. The Dance Group had better unilateral postural balance with eyes closed. The Tai Chi Chuan Group had faster walking speeds, shorter transfer times, and better postural balance in the final standing position during the Sit-to-stand Test. .

/metabolism , Cyclic AMP/metabolism , Dictyostelium/enzymology , Dictyostelium/genetics , GTP-Binding Protein alpha Subunits, Gi-Go/metabolism , Protozoan Proteins/metabolism , /genetics , Dictyostelium/growth & development , Dictyostelium/metabolism , Down-Regulation/drug effects , Extracellular Matrix Proteins/genetics , Extracellular Matrix Proteins/metabolism , Folic Acid/pharmacology , /deficiency , /genetics , /metabolism , GTP-Binding Protein alpha Subunits, Gi-Go/deficiency , GTP-Binding Protein alpha Subunits, Gi-Go/genetics , Mutation , Mitogen-Activated Protein Kinase 1/genetics , Mitogen-Activated Protein Kinase 1/metabolism , Protozoan Proteins/genetics , Signal Transduction , Spores, Protozoan/enzymology , Spores, Protozoan/genetics , Vitamin B Complex/pharmacology
Mem. Inst. Oswaldo Cruz ; 110(1): 56-64, 03/02/2015. tab, graf
Article in English | LILACS | ID: lil-741619


Histology is the gold standard for diagnosing acute rejection and hepatitis C recurrence after liver transplantation. However, differential diagnosis between the two can be difficult. We evaluated the role of C4d staining and quantification of hepatitis C virus (HCV) RNA levels in liver tissue. This was a retrospective study of 98 liver biopsy samples divided into four groups by histological diagnosis: acute rejection in patients undergoing liver transplant for hepatitis C (RejHCV+), HCV recurrence in patients undergoing liver transplant for hepatitis C (HCVTx+), acute rejection in patients undergoing liver transplant for reasons other than hepatitis C and chronic hepatitis C not transplanted (HCVTx-). All samples were submitted for immunohistochemical staining for C4d and HCV RNA quantification. Immunoexpression of C4d was observed in the portal vessels and was highest in the HCVTx- group. There was no difference in C4d expression between the RejHCV+ and HCVTx+ groups. However, tissue HCV RNA levels were higher in the HCVTx+ group samples than in the RejHCV+ group samples. Additionally, there was a significant correlation between tissue and serum levels of HCV RNA. The quantification of HCV RNA in liver tissue might prove to be an efficient diagnostic test for the recurrence of HCV infection.

Animals , Humans , Mice , Annexin A1/pharmacology , Macrophages/drug effects , Macrophages/immunology , Neutrophils/cytology , Neutrophils/immunology , Apoptosis , Actins/metabolism , Annexin A1/deficiency , Annexin A1/genetics , Annexin A1/immunology , Cyclic AMP-Dependent Protein Kinases/metabolism , Cyclic AMP/metabolism , Dexamethasone/pharmacology , In Vitro Techniques , /biosynthesis , Mice, Knockout , Macrophages/metabolism , Peptides , Phagocytosis/drug effects , Transforming Growth Factor beta/biosynthesis
Rev. bras. cir. cardiovasc ; 30(1): 77-83, Jan-Mar/2015. tab, graf
Article in English | LILACS | ID: lil-742899


Introduction: Previous studies have demonstrated that thymoquinone has protective effects against ischemia reperfusion injury to various organs like lungs, kidneys and liver in different experimental models. Objective: We aimed to determine whether thymoquinone has favorable effects on lung, renal, heart tissues and oxidative stress in abdominal aorta ischemia-reperfusion injury. Methods: Thirty rats were divided into three groups as sham (n=10), control (n=10) and thymoquinone (TQ) treatment group (n=10). Control and TQ-treatment groups underwent abdominal aorta ischemia for 45 minutes followed by a 120-min period of reperfusion. In the TQ-treatment group, thymoquinone was given 5 minutes. before reperfusion at a dose of 20 mg/kg via an intraperitoneal route. Total antioxidant capacity, total oxidative status (TOS), and oxidative stress index (OSI) in blood serum were measured and lung, kidney, and heart tissue histopathology were evaluated with light microscopy. Results: Total oxidative status and oxidative stress index activity in blood samples were statistically higher in the control group compared to the sham and TQ-treatment groups (P<0.001 for TOS and OSI). Control group injury scores were statistically higher compared to sham and TQ-treatment groups (P<0.001 for all comparisons). Conclusion: Thymoquinone administered intraperitoneally was effective in reducing oxidative stress and histopathologic injury in an acute abdominal aorta ischemia-reperfusion rat model. .

Introdução: Estudos prévios demonstraram que a timoquinona tem efeitos protetores contra a lesão de isquemia/reperfusão em vários órgãos como pulmão, rins e fígado em diferentes modelos experimentais. Objetivo: Determinar se timoquinona tem efeitos positivos em tecidos do pulmão, rim e coração e no estresse oxidativo em lesão de isquemia/perfusão da aorta abdominal. Métodos: Trinta ratos foram divididos em três grupos: sham (n=10), controle (n=10) e tratamento com timoquinona (TQ) (n=10). Os grupos controle e de tratamento com TQ foram submetidos à isquemia da aorta abdominal durante 45 minutos, seguido por um período de 120 minutos de reperfusão. No grupo de tratamento com TQ, a timoquinona foi administrada 5 minutos antes da reperfusão, dose de 20 mg/kg através da via intraperitoneal. A capacidade total antioxidante, estado oxidativo total (TOS) e o índice de estresse oxidativo (OSI) no soro do sangue foram medidos, e a histopatologia dos tecidos do pulmão, rim e coração foram avaliados com microscopia de luz. Resultados: Estado oxidativo e índice de estresse oxidativo total em amostras de sangue foram estatisticamente mais altos no grupo controle em relação aos grupos sham e tratamento com TQ (P<0,001 para TOS e OSI). Escores de lesões no grupo controle foram estatisticamente mais altos em relação aos grupos sham e tratamento com TQ (P<0,001 para todas as comparações). Conclusão: A timoquinona administrada por via intraperitoneal foi eficaz na redução do estresse oxidativo e lesão histopatológica em modelo de rato de isquemia/reperfusão aguda da aorta abdominal. .

Animals , Humans , Mice , Endothelial Cells/enzymology , Glucagon-Like Peptide 1/pharmacology , Microvessels/cytology , Nitric Oxide Synthase Type II/metabolism , Nitric Oxide/metabolism , Poly(ADP-ribose) Polymerases/metabolism , Recombinant Proteins/pharmacology , Signal Transduction/drug effects , Apoptosis/drug effects , Cell Line , Cell Proliferation/drug effects , Cyclic AMP/metabolism , Endothelial Cells/drug effects , Gene Expression/drug effects , Lipoproteins, LDL/pharmacology
Rev. bras. enferm ; 67(6): 898-904, Nov-Dec/2014. tab
Article in Portuguese | LILACS, BDENF | ID: lil-732821


Objetivou-se analisar as características demográficas e clínicas dos clientes diagnosticados com Síndrome de Stevens Johnson (SSJ) e Necrólise Epidérmica Tóxica (NET), bem como identificar as ações dos profissionais de saúde para o manejo das Reações Adversas a Medicamentos (RAM) em um hospital público do Distrito Federal. Pesquisa descritiva, retrospectiva, com abordagem quantitativa. Dados coletados em todos os prontuários de 22 clientes internados de janeiro de 2005 a setembro de 2012. Análise mediante estatística descritiva. Houve aumento gradativo de casos, com maior número nos anos de 2007 e 2012. Dos casos analisados, 9 foram diagnosticados com NET e 7 com SSJ; predominaram as mulheres (14) e a faixa etária de 21 aos 40 anos (10); 21 obtiveram cura. Os fármacos associados a RAM mais frequentes foram os antiepilépticos (10). Observou-se fragilidade nos registros clínicos nos prontuários e nas ações de monitoramento de RAM no serviço estudado.

This study aimed to analyze demographic and clinical aspects of patients diagnosed with Stevens Johnson Syndrome (SJS) and Toxic Epidermal Necrolysis (TEN), as well as identifying the actions of health professionals for the management of Adverse Drug Reactions (ADR) in a public hospital in Distrito Federal, Brazil. A descriptive and retrospective research was held, with quantitative approach. Data collected from all the records of 22 patients admitted with diagnosed with SJS and TEN, from January 2005 to September 2012. Data were analyzed using descriptive statistics. Of these cases, 9 were diagnosed with NET and 7, with SJS; there were more females (14); aged from 21 to 40 years (10); 21 were cured; the drugs more used were the antiepileptic ones (10). Fragility in clinical registers and in the actions to monitor the cases of ADR in this health service was observed.

Este estudio tuvo como objetivo analizar aspectos demográficos y clínicos de clientes con diagnóstico de Síndrome de Stevens Johnson (SSJ) y Necrólisis Epidérmica Tóxica (NET), así como la identificación de las acciones de los profesionales de la salud para el manejo de reacciones adversas a medicamentos (RAM) en un hospital público del Distrito Federal. Se realizó investigación descriptiva, retrospectiva con enfoque cuantitativo. Datos recogidos de prontuarios clínicos de los 22 clientes ingresados con diagnóstico de SJS y NET, de enero de 2005 a septiembre de 2012. Fueron analizados utilizando estadística descriptiva. De estos casos, 9 fueron diagnosticados con NET, 7 con SJS; había más mujeres (14); edad entre 21 y 40 años (10); 21 se curaron; predominaran los antiepilépticos (10). Fue observado que hay fragilidad en registros clínicos en los prontuarios y en las acciones de monitoreo de las RAM en este servicio de salud.

Animals , Male , Rats , Insulin , Somatostatin/pharmacology , Adenylyl Cyclases/metabolism , Calcium/metabolism , Cyclic AMP/metabolism , In Vitro Techniques , Islets of Langerhans/drug effects , Islets of Langerhans/metabolism , Islets of Langerhans , Rats, Inbred Strains
Medicina (B.Aires) ; 73(2): 155-162, abr. 2013. ilus
Article in English | LILACS | ID: lil-694758


Autosomal dominant polycystic kidney disease (ADPKD), a most common genetic cause of chronic renal failure, is characterized by the progressive development and enlargement of cysts in kidneys and other organs. The cystogenic process is highly complex and involves a high proliferative rate, increased apoptosis, altered protein sorting, changed secretory characteristics, and disorganization of the extracellular matrix. ADPKD is caused by mutations in the genes encoding polycystin-1 (PC-1) or polycystin-2 (PC-2). PC-1 undergoes multiple cleavages that intervene in several signaling pathways involved in cellular proliferation and differentiation mechanisms. One of these cleavages releases the cytoplasmic C-terminal tail of PC-1. In addition, the C-terminal cytoplasmic tails of PC-1 and PC-2 interact in vitro and in vivo. The purpose of this review is to summarize recent literature that suggests that PC-1 and PC-2 may function through a common signaling pathway necessary for normal tubulogenesis. We hope that a better understanding of PC-1 and PC-2 protein function will lead to progress in diagnosis and treatment for ADPKD.

La poliquistosis renal autosómica dominante (ADPKD por sus siglas en inglés) es una causa genética muy común de falla renal crónica que se caracteriza por el progresivo desarrollo y agrandamiento de quistes en los riñones y en otros órganos. El proceso de cistogénesis comprende incrementos en la proliferación y muerte celular por apoptosis, así como alteraciones en la distribución intracelular de proteínas, el movimiento transcelular de solutos y organización de la matriz extracelular. ADPKD es causada por mutaciones en los genes que codifican para policistina-1 (PC-1) o policistina-2 (PC-2). PC-1 puede sufrir múltiples clivajes y los fragmentos generados intervienen en diferentes cascadas de señalización involucradas en mecanismos de proliferación y diferenciación celular. Uno de estos clivajes libera el extremo C-terminal citoplasmático de la PC-1. Se ha demostrado que los extremos C-terminal citoplasmático de PC-1 y PC-2 pueden interactuar tanto in vitro como in vivo. El propósito de esta revisión es resumir la literatura más reciente que sugiere que PC-1 y PC-2 pueden funcionar a través de una cascada de señalización común necesaria para la tubulogénesis normal. Creemos que una mejor comprensión de los mecanismos moleculares de acción de PC-1 y PC-2 contribuirán al progreso en el diagnóstico y tratamiento de ADPKD.

Animals , Humans , Polycystic Kidney, Autosomal Dominant/metabolism , TRPP Cation Channels/metabolism , Apoptosis/physiology , Cell Proliferation , Calcium Channels/metabolism , Cell Nucleus/metabolism , Cyclic AMP/metabolism , Kidney Tubules/metabolism , Mutation , Polycystic Kidney, Autosomal Dominant/genetics
Yonsei Medical Journal ; : 772-777, 2013.
Article in English | WPRIM | ID: wpr-211908


PURPOSE: SNF2L belongs to Imitation Switch family and plays an essential role in neural tissues and gonads. In our previous studies, we have demonstrated that the basal transcription of human SNF2L gene is regulated by two cis-elements, cAMP response element (CRE)- and Sp1-binding sites. Recent studies suggested that cyclic adenosine monophosphate (cAMP) stimulation significantly up-regulated SNF2L expression in ovarian granulose cells. These data suggested that protein kinase-mediated signal pathways might also regulate SNF2L expression in neural cells. We therefore investigated the effects of agents that activate protein kinases A on SNF2L gene expression in neural cells. MATERIALS AND METHODS: To increase intracellular cAMP levels, all neural cells were treated with forskolin and dbcAMP, two cAMP response activators. We exmined the effects of cAMP on the promoter activity of human SNF2L gene by luciferase reporter gene assays, and further examined the effects of cAMP on endogenous SNF2L mRNA levels by qPCR. RESULTS: Transient expression of a luciferase fusion gene under the control of the SNF2L promoter was significantly increased by treatment of rat primary neurons with forskolin or dbcAMP, but not PC12, C6 and SH-SY5Y cells. Consistently, treatment with forskolin or dbcAMP could enhance endogenous SNF2L mRNA levels also only in rat primary neurons. CONCLUSION: These results suggest that the CRE consensus sequence in the SNF2L proximal promoter most likely confers constitutive activation and regulation by cAMP in neural cells.

Animals , Bucladesine/pharmacology , Cell Line , Colforsin/pharmacology , Cyclic AMP/metabolism , DNA-Binding Proteins/chemistry , Gene Expression Regulation , Humans , Luciferases/analysis , Neurons/metabolism , PC12 Cells , Promoter Regions, Genetic , RNA, Messenger/metabolism , Rats , Rats, Wistar , Recombinant Fusion Proteins/analysis , Response Elements , Transcription Factors/chemistry
Article in English | IMSEAR | ID: sea-138997


Background & objectives: Chapekar established a model of ovarian tumourigenesis in mice by splenic transplantation of ovaries, resulting in sustained luteinizing hormone (LH) levels because of absence of feedback inhibition. There is increasing evidence of the differential response to LH or hCG under various experimental conditions. The effect of sustained hormonal stimulation in long term cultures is sparsely investigated. The study is aimed to determine the role of hCG and LH stress on caprine ovarian granulosa cells and their downstream signaling in short and long term cultures. Methods: To study the response of hCG and LH stress and downstream signaling, short term cultures were set up by exposing goat ovarian granulosa cells in primary cultures to hCG and LH stress (levels beyond their physiological doses) for 5 days (P0). Cells were sub-cultured at sixth day and subjected to prolonged LH/ hCG stress for two weeks in passage 1(P1) (long term cultures). Downstream cell signaling molecules were assessed. Intracellular cAMP was estimated by ELISA. For PKA and PKC, activity assays were performed. pERK protein expressions in short term cultures were assessed by Western blot and flowcytometry; in long term cultures, pERK expression was analyzed by flowcytometry. Results: Differential effects on cell proliferation were observed in long term cultures, where the untreated and hCG exposed cells showed markedly reduced cell proliferation after second week of exposure while LH treated cells continued to proliferate. Different levels of cAMP, PKA, PKC and phosphorylated ERK1/2 were observed on short term and long term LH stimulation. On sustained hormonal stimulation, cAMP levels were significantly (P<0.05) higher in hCG treated cultures as compared to controls and LH treated cultures. LH led to maximal elevation of ERK in long term cultures. Interpretation & Conclusions: As pERK1/2 promotes cellular proliferation, activation of ERK1/2 in LH treated cultures may be responsible for sustained growth. Prolonged LH treatment promoted growth and proliferation in caprine ovarian granulosa cells whereas prolonged exposure to hCG led to elevated levels of cAMP and decreased the rate of proliferation. Defining the signals and second messengers that act as survival or apoptotic mediators may help in elucidation of the mechanisms controlling proliferation or programmed cell death in granulosa cells.

Animals , Cell Culture Techniques , Cell Proliferation/drug effects , Chorionic Gonadotropin/pharmacology , Cyclic AMP/metabolism , Cyclic AMP-Dependent Protein Kinases/metabolism , Female , Goats , Granulosa Cells/drug effects , Luteinizing Hormone/pharmacology , Mitogen-Activated Protein Kinase 3/metabolism , Primary Cell Culture , Protein Kinase C/metabolism , Signal Transduction/drug effects
Article in English | IMSEAR | ID: sea-135373


Campylobacter jejuni is a foodborne pathogen and a leading cause of diarrhoea worldwide. It is believed that a cholera toxin-like toxin (CTLT) produced by C. jejuni may mediate watery diarrhoea. However, the production of a CTLT by C. jejuni is controversial. A cholera toxin gene (ctx) homologue has not been identified in Campylobacter species. We investigated the identity of the CT cross-reactive antigen from Campylobacter species previously and the results are reviewed here. Filtrates of C. jejuni grown in four different liquid media, reported to promote CTLT production, were tested by Chinese hamster ovary (CHO) cell elongation assay for functional toxin and for reactivity with CT antibody using GM1 ganglioside ELISA (GM1 ELISA) and immunoblotting. Protein sequence of the CT antibody-reactive band was determined by matrix-assisted laser desorption ionization-time of flight (MALDI TOF-TOF). Non-jejuni species (C. coli, C. lari, C. foetus, C. hyointestinalis and C. upsaliensis) were investigated by CHO cell assay and immunoblotting. Filtrates from seven C. jejuni reference strains reported to produce CTLT and from 80 clinical strains were negative in the CHO cell assay. However, filtrates from three reference strains and 16 clinical strains were positive by GM1 ELISA. All strains irrespective of GM1 ELISA reactivity, possessed a 53-kDa protein which reacted with CT antibody by immunoblotting. This band was identified as the major outer membrane protein (PorA) of C. jejuni. CT antibody reacted with a C. jejuni recombinant PorA on immunoblotting. All non-C. jejuni strains were negative by CHO cell assay, but the common 53-kDa proteins reacted with CT antibody on immunoblots. The cross-reactivity of PorAs of Campylobacter species with CT may lead to the erroneous conclusion that Campylobacter species produce a functional CTLT.

Animals , Bacterial Proteins/metabolism , CHO Cells , Campylobacter/metabolism , Campylobacter/pathogenicity , Cholera Toxin/metabolism , Cricetinae , Cricetulus , Cyclic AMP/metabolism , Membrane Proteins/metabolism
Braz. j. med. biol. res ; 43(1): 36-42, Jan. 2010. tab, ilus
Article in English | LILACS | ID: lil-535640


Transforming growth factor-â1 (TGF-â1) plays an important role in the fibrogenic process in the liver. The aim of the present study was to explore the action of TGF-â1 on fibronectin expression in rat hepatic stem-like cells and the underlying mechanisms. The level of fibronectin expression was determined in hepatic stem-like cells (WB cells) before and after TGF-â1 stimulation by RT-PCR and Western blot methods. Using immunogold transmission electron microscopy and the Western blot method, we observed the result of the expression and the distribution of cAMP, phosphorylated Smad3 and Smad7 before and after TGF-â1 treatment. The levels of fibronectin expression in both mRNA and protein increased 4- to 5-fold after TGF-â1 stimulation, reaching an optimum level after 8 h and then gradually falling back. Similarly, TGF-â1 stimulation resulted in an increase of cAMP in WB cells, peaking at 8 h. After treatment with TGF-â1 for 24 h, the expression of cAMP gradually decreased. In addition, we found that TGF-â1 treatment also contributed to the increased expression and to changes in cellular distribution of phosphorylated Smad3 (translocation from the cytoplasm to the nucleus) and Smad7 (translocation from the nucleus to the cytoplasm) in WB cells. The present study demonstrates that TGF-â is involved in the fibrogenic process in hepatic stem cells through up-regulation of fibronectin expression, and the mechanisms underlying this process may be associated with the activation of cAMP and Smad pathways.

Animals , Rats , Fibronectins/metabolism , Hepatocytes/metabolism , Stem Cells/metabolism , Transforming Growth Factor beta/pharmacology , Blotting, Western , Cyclic AMP/metabolism , Fibronectins/genetics , Hepatocytes/pathology , Liver Cirrhosis/etiology , Microscopy, Electron, Transmission , Reverse Transcriptase Polymerase Chain Reaction , RNA, Messenger/metabolism , Smad Proteins/metabolism , Stem Cells/pathology
Article in English | WPRIM | ID: wpr-104279


Theophylline is commonly used to treat severe asthma and chronic obstructive pulmonary disease (COPD) characterized by non-eosinophilic inflammation. Acetyl salicylic acid (ASA) is one of the most widely used medications worldwide, but up to 20% of patients with asthma experience aggravated respiratory symptoms after taking ASA. Here we evaluated the adverse effect of ASA on the therapeutic effect of theophylline in mice with non-eosinophilic asthma. A non-eosinophilic asthma mouse model was induced by airway sensitization with lipopolysaccharide-containing allergen and then challenged with allergen alone. Therapeutic intervention was performed during allergen challenge. Theophylline inhibited lung inflammation partly induced by Th1 immune response. ASA attenuated the beneficial effects of theophylline. However, co-administration of the ASA metabolite salicylic acid (SA) showed no attenuating effect on theophylline treatment. The therapeutic effect of theophylline was associated with increase in cAMP levels, which was blocked by co-treatment of theophylline and ASA. ASA co-treatment also attenuated the anti-inflammatory effects of a specific phosphodiesterase 4 inhibitor. These results demonstrate that ASA reverses anti-inflammatory effects of theophylline, and that ASA exerts its adverse effects through the inhibition of cAMP production. Our data suggest that ASA reverses lung inflammation in patients taking theophylline, although clinical evidence will be needed.

Animals , Anti-Inflammatory Agents/therapeutic use , Aspirin/therapeutic use , Asthma/drug therapy , Blotting, Western , Bronchoalveolar Lavage Fluid , Cyclic AMP/metabolism , Enzyme-Linked Immunosorbent Assay , Immunoprecipitation , Mice , Mice, Inbred C57BL , Mice, Knockout , Pulmonary Eosinophilia/drug therapy , Theophylline/therapeutic use
Arch. argent. dermatol ; 59(6): 245-249, 2009. ilus, graf
Article in Spanish | LILACS | ID: lil-620543


La neurofibromina es una proteína citoplasmática codificada por el gen NF1, localizado en el cromosoma 17q11.2. Participa en la regulación de diversas vías de señalización molecular: MAPK, P13K, Caveolina y PKA. Cada vía cumple una función particular, con un objetivo final común, el control de genes involucrados en el crecimiento, apoptosis, diferenciación y migración celular. Realizamos una revisión sobre la neurofibromina, las vías de señalización en las que está implicada y su relación con las manifestaciones clínicas de la neurofibromatosis.

Humans , Neurofibromatosis 1/genetics , Neurofibromatosis 1/pathology , Neurofibromin 1/physiology , Neurofibromin 1/isolation & purification , Neurofibromin 1/metabolism , Cyclic AMP/metabolism , Neurofibroma/pathology , ras Proteins
Article in English | WPRIM | ID: wpr-107284


In this study, the essential oil from lotus flower extract, including petals and stamens, was assessed with regard to its effects on melanogenesis in human melanocytes. The lotus flower essential oil was shown to stimulate melanin synthesis and tyrosinase activity in a dose-dependent manner. The lotus flower essential oil induced the expression of tyrosinase, microphthalmia-associated transcription factor M (MITF-M), and tyrosinase-related proten-2 (TRP-2) proteins, but not tyrosinase mRNA. Moreover, it increased the phosphorylation of ERK and cAMP response element binding protein (CREB). In order to verify the effective components of the lotus flower oil, its lipid composition was assessed. It was found to be comprised of palmitic acid methyl ester (22.66%), linoleic acid methyl ester (11.16%), palmitoleic acid methyl ester (7.55%) and linolenic acid methyl ester (5.16%). Among these components, palmitic acid methyl ester clearly induced melanogenesis as the result of increased tyrosinase expression, thereby indicating that it may play a role in the regulation of melanin content. Thus, our results indicate that lotus flower oil may prove useful in the development of gray hair prevention agents or tanning reagents.

Blotting, Western , Cell Proliferation , Cyclic AMP/metabolism , Cyclic AMP Response Element-Binding Protein/genetics , Flowers/chemistry , Gas Chromatography-Mass Spectrometry , Humans , Intramolecular Oxidoreductases/genetics , Lotus/chemistry , Melanins/biosynthesis , Melanocytes/drug effects , Microphthalmia-Associated Transcription Factor/genetics , Monophenol Monooxygenase/genetics , Phosphorylation , Plant Oils/pharmacology , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction , Skin/cytology
Indian J Exp Biol ; 2008 Nov; 46(11): 764-9
Article in English | IMSEAR | ID: sea-61661


In the present in vitro study, the involvement of cAMP dependent-protein kinase A (PKA) and calcium-dependent protein kinase C (PKC) in the regulation of forebrain (telencephalon and hypothalamus) tyrosine hydroxylase (TH) activity was demonstrated during the reproductive seasons of the female catfish H. fossilis. In the concentration studies conducted in prespawning phase, cAMP (0.05 nM, 0.5 nM, 1 mM and 2.0 mM) or the phosphodiesterase inhibitor isobutylmethylxanthine (IBMX-0.5-2.0 mM) stimulated enzyme activity. Likewise, the incubation of the enzyme preparations with the cAMP dependent-protein kinase A inhibitor H-89 (1 and 10 microM) and PKC inhibitor calphostin C (cal C; 1 and 10 microM) inhibited enzyme activity in a concentration-dependent manner. In seasonal studies, the incubation of the enzyme preparations with cAMP (1 mM), IBMX (1 mM), H-89 (10 microM) and cal-C (10 microM) produced season-dependent effects on enzyme activity. The stimulatory effect of cAMP and IBMX and the inhibitory effect of H-89 and cal C were greater in the resting and spawning phases. The results suggest the involvement of both signal transduction pathways in TH activation vis-à-vis catecholaminergic activity with a more dominant role by the cAMP-PKA pathway.

1-Methyl-3-isobutylxanthine/pharmacology , Animals , Brain/enzymology , Calcium/metabolism , Catfishes , Cyclic AMP/metabolism , Cyclic AMP-Dependent Protein Kinases/metabolism , Female , Fossils , Isoquinolines/pharmacology , Naphthalenes/pharmacology , Protein Kinase C/metabolism , Seasons , Signal Transduction , Sulfonamides/pharmacology , Tyrosine 3-Monooxygenase/chemistry
J Biosci ; 2007 Dec; 32(7): 1281-9
Article in English | IMSEAR | ID: sea-110792


Cyclic AMP (cAMP)is a natural chemoattractant of the social amoeba Dictyostelium discoideum. It is detected by cell surface cAMP receptors. Besides a signalling cascade involving phosphatidylinositol 3,4,5-trisphosphate (PIP3), Ca2+ signalling has been shown to have a major role in chemotaxis. Previously, we have shown that arachidonic acid (AA) induces an increase in the cytosolic Ca2+ concentration by causing the release of Ca2+ from intracellular stores and activating influx of extracellular Ca2+. Here we report that AA is a chemoattractant for D. discoideum cells differentiated for 8-9 h. Motility towards a glass capillary filled with an AA solution was dose-dependent and qualitatively comparable to cAMP-induced chemotaxis. Ca2+ played an important role in AA chemotaxis of wild-type Ax2 as ethyleneglycol-bis(b-aminoethyl)-N,N,N',N'-tetraacetic acid (EGTA) added to the extracellular buffer strongly inhibited motility. In the HM1049 mutant whose iplA gene encoding a putative Ins(1,4,5)P3 -receptor had been knocked out, chemotaxis was only slightly affected by EGTA. Chemotaxis in the presence of extracellular Ca2+ was similar in both strains. Unlike cAMP, addition of AA to a cell suspension did not change cAMP or cGMP levels. A model for AA chemotaxis based on the findings in this and previous work is presented.

Animals , Arachidonic Acid/pharmacology , Calcium/metabolism , Chemotaxis/drug effects , Cyclic AMP/metabolism , Cyclic GMP/metabolism , Dictyostelium/cytology , Egtazic Acid/pharmacology , Mutation
J Biosci ; 2007 Sep; 32(6): 1185-94
Article in English | IMSEAR | ID: sea-110648


Intrafollicular factors play an important role in folliculogenesis. The follicle-stimulating hormone (FSH)-binding inhibitor (FSHBI), purified by our laboratory from human ovarian follicular fluid, has been shown to suppress ovulation and induce follicular atresia/apoptosis in mice as well as impair fertility in marmosets, the New World monkeys. The octapeptide, a peptide corresponding to the N-terminal region of human FSHBI (hFSHBI), has been synthesized and also shows FSHBI activity in vitro. In the present study, we have attempted to identify the mechanism of action of the peptide in granulosa cell cultures. Rat granulosa cell cultures were treated with varying concentrations of the octapeptide or partially purified hFSHBI (gel chromatography fraction hGF 2) in the presence or absence of human FSH (hFSH) and the amount of progesterone (P;4) secreted in the culture supernatants after 3 h/48 h was estimated. Both hGF2 and the octapeptide failed to alter basal levels as well as 8-bromo cAMP-induced P;4 production, while FSH-induced P 4 secretion was inhibited in a dose-dependent manner. These studies reveal that the octapeptide, a fragment of FSHBI, and the native protein have similar activity in vitro and both compounds alter FSH action at the receptor level upstream of cAMP formation.

Animals , Carrier Proteins/physiology , Cells, Cultured , Cyclic AMP/metabolism , Down-Regulation/physiology , Female , Follicle Stimulating Hormone/antagonists & inhibitors , Glycopeptides/physiology , Granulosa Cells/metabolism , Humans , Intercellular Signaling Peptides and Proteins/physiology , Peptide Fragments/physiology , Progesterone/antagonists & inhibitors , Rats , Rats, Sprague-Dawley , Receptors, FSH/antagonists & inhibitors
Arq. bras. endocrinol. metab ; 51(5): 654-671, jul. 2007. ilus, tab
Article in English | LILACS | ID: lil-463385


Thyroid cancers are the most frequent endocrine neoplasms and mutations in the thyrotropin receptor (TSHR) are unusually frequent. Here we present the state-of-the-art concerning the role of TSHR in thyroid cancer and discuss it in light of the cancer stem cell theory or the classical view. We briefly review the gene and protein structure updating the cancer related TSHR mutations database. Intriguingly, hyperfunctioning TSHR mutants characterise differentiated cancers in contrast to undifferentiated thyroid cancers which very often bear silenced TSHR. It remains unclear whether TSHR alterations in thyroid cancers play a role in the onset or they appear as a consequence of genetic instability during evolution, but the presence of functional TSHR is exploited in therapy. We outline the signalling network build up in the thyrocyte between TSHR/PKA and other proliferative pathways such as Wnt, PI3K and MAPK. This networks integrity surely plays a role in the onset/evolution of thyroid cancer and needs further research. Lastly, future investigation of epigenetic events occurring at the TSHR and other loci may give better clues for molecular based therapy of undifferentiated thyroid carcinomas. Targeted demethylating agents, histone deacetylase inhibitors combined with retinoids and specific RNAis may help treatment in the future.

Os cânceres de tiróide são as neoplasias endócrinas mais frequentes e as mutações no receptor de tirotrofina (TSHR) são incomumente frequentes. Nesta revisão nós apresentamos o "estado da arte" com relação ao papel do TSHR no câncer de tiróide e o discutimos à luz da teoria da célula matriz do câncer ou a visão clássica. Revisamos brevemente a estrutura do gene e da proteína, atualizando a base de dados das mutações do TSHR relacionadas ao câncer. Curiosamente, mutações do TSHR com hiperfunção caracterizam cânceres diferenciados, em contraste com os cânceres de tiróide indiferenciados, os quais muito comumente mostram TSHR silenciados. Permanece obscuro se as alterações do TSHR em cânceres de tiróide têm algum papel no surgimento ou se elas aparecem como conseqüência da instabilidade genética durante seu desenvolvimento, mas a presença de TSHR funcional é explorada na terapia. Nós delineamos a rede de sinalizacão desenvolvida no tirócito entre TSHR/PKA e outras vias proliferativas como a Wnt, PI3k e MAPK. A integridade desta rede certamente tem um papel no surgimento/evolução do câncer de tiróide e necessita de novas pesquisas. Finalmente, novas investigacões sobre os eventos epigenéticos que ocorrem no TSHR e outros locais poderão trazer novas informações para uma terapia de base molecular nos carcinomas indiferenciados de tiróide. Agentes demetilantes direcionados, inibidores da histona-deacetilase, combinados com retinóides e RNAs específicos poderão auxiliar no tratamento futuro.

Humans , Receptors, Thyrotropin/metabolism , Signal Transduction/physiology , Thyroid Neoplasms/genetics , Thyrotropin/metabolism , Cell Proliferation , Cyclic AMP Response Element-Binding Protein/metabolism , Cyclic AMP/metabolism , GTP-Binding Proteins/metabolism , Gene Expression Regulation/physiology , MAP Kinase Signaling System/physiology , Mutation/genetics , Neoplastic Stem Cells/metabolism , Receptors, Thyrotropin/genetics , Thyroid Neoplasms/metabolism , Thyrotropin/genetics , Wnt Proteins/metabolism
J Biosci ; 2007 Mar; 32(2): 329-38
Article in English | IMSEAR | ID: sea-111149


The three basic cell types in the migrating slug of Dictyostelium discoideum show differential chemotactic response to cyclic AMP (cAMP) and differential sensitivity to suppression of the chemotaxis by ammonia.The values of these parameters indicate a progressive maturation of chemotactic properties during the transdifferentiation of slug cell types.We present a model that explains the localization of the three cell types within the slug based on these chemotactic differences and on the maturation of their chemotactic properties.

Ammonia/toxicity , Animals , Cell Differentiation/physiology , Chemotactic Factors/metabolism , Chemotaxis/drug effects , Cyclic AMP/metabolism , Dictyostelium/cytology , Models, Biological