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1.
Biomed Pharmacother ; 163: 114860, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37196540

RESUMO

The antibiotic-induced intestinal injury (AIJ) is associated with diarrhoea and gastrointestinal discomfort. However, the pathological intestinal mechanisms and related side effects associated with antibiotic use/misuse may be counteracted by probiotics. This study aims to evaluate the effect and the protective mechanisms of a probiotic formulation containing Alkalihalobacillus clausii (formerly Bacillus clausii; BC) spores in an experimental model of AIJ. C57/Bl6J mice were orally challenged with a high dose of ceftriaxone for five days along with BC treatment which lasted up to the 15th day. Our results showed the beneficial effect of the probiotic in preserving colonic integrity and limiting tissue inflammation and immune cell infiltration in AIJ mice. BC increased tight junction expression and regulated the unbalanced production of colonic pro- and anti-inflammatory cytokines, converging toward the full resolution of the intestinal damage. These findings were supported by the histological evaluation of the intestinal mucosa, suggesting a potential restoration of mucus production. Notably, BC treatment increased gene transcription of the secretory products responsible for epithelium repair and mucus synthesis and normalized the expression of antimicrobial peptides involved in immune activation. Reconstruction of complex and diverse gut microbiota in antibiotic-induced dysbiosis was recorded upon BC supplementation. Specifically, the expansion of A. clausii, Prevotella rara and Eubacterium ruminatium drove intestinal microbiota rebalance by primarily impacting Bacteroidota members. Taken together, our data indicate that BC administration alleviates AIJ by multiple converging mechanisms leading to restoring gut integrity and homeostasis and reshaping microbiota composition.


Assuntos
Bacillus clausii , Microbioma Gastrointestinal , Enteropatias , Probióticos , Animais , Camundongos , Antibacterianos/uso terapêutico , Bacillus clausii/fisiologia , Esporos Bacterianos , Enteropatias/tratamento farmacológico , Mucosa Intestinal , Probióticos/farmacologia
2.
Int J Immunopathol Pharmacol ; 23(1): 227-34, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20378008

RESUMO

Pseudomonas fluorescens is a Gram-negative bacterium generally considered of scarce clinical significance. However, in the last few years, the isolation of P. fluorescens as the causative agent of nosocomial infections has rapidly increased. P. fluorescens is a psychrophile microorganism which grows at an optimal temperature of 25-30 degrees Celcius. In spite of this constraint, it has recently been reported that the human physiological temperature does not appear to be a barrier for this microorganism. In this study we examined the ability of P. fluorescens, grown at 28 degrees C or at 37 degrees C, to adhere to cultured human A549 pulmonary cells and to form biofilm. The ability of P. fluorescens to induce expression of proinflammatory cytokines, beta-defensin 2 and the intercellular adhesion molecule-1 was also investigated. Our results clearly indicate that inflammatory mediators are induced when the microorganism is grown at a lower temperature, while biofilm is formed only at 37 degrees C. The results presented are consistent with previous reports indicating P. fluorescens as an opportunistic pathogen and underscore the urgent need for further studies to better characterize the virulence of this microorganism.


Assuntos
Pseudomonas fluorescens/fisiologia , Aderência Bacteriana , Biofilmes , Linhagem Celular , Citocinas/biossíntese , Humanos , Molécula 1 de Adesão Intercelular/biossíntese , Pseudomonas fluorescens/patogenicidade , Temperatura , beta-Defensinas/biossíntese
3.
J Cell Physiol ; 214(3): 582-7, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-17786942

RESUMO

In this study, we have evaluated the effects on cell cycle regulation of VacA alone and in combination with other two Helicobacter pylori proteins, cytotoxin-associated protein (CagA) and HspB, using the human gastric epithelial cells (AGS). Our results indicate that VacA alone was able to inhibit the G1 to S progression of the cell cycle. The VacA capacity of inhibiting cell progression from G1 to S phase was also observed when cells were co-transfected with CagA or HspB. Moreover, VacA over-expression caused apoptosis in AGS cells through activation of caspase 8 and even more of caspase 9, thus indicating an involvement of both the receptor-mediated and the mitochondrial pathways of apoptosis. Indeed, the two pathways probably can co-operate to execute cell death with a prevalence of the mitochondrial pathways. Our data taken together provide additional information to further enhance our understanding of the molecular mechanism by which H. pylori proteins alter the growth status of human gastric epithelial cells.


Assuntos
Apoptose , Proteínas de Bactérias/metabolismo , Ciclo Celular , Células Epiteliais/citologia , Helicobacter pylori/metabolismo , Estômago/citologia , Antígenos de Bactérias/metabolismo , Caspases/metabolismo , Linhagem Celular , Ativação Enzimática , Células Epiteliais/enzimologia , Citometria de Fluxo , Proteínas de Choque Térmico/metabolismo , Humanos , Immunoblotting , Proteína do Retinoblastoma/metabolismo , Estômago/enzimologia , Transfecção
4.
Biochim Biophys Acta ; 1496(2-3): 285-95, 2000 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-10771097

RESUMO

The effect of nontoxic, low concentrations (10(-8) M) of retinoic acid (RA) for a relatively long time (28 days) on a Kirsten ras-virus transformed cell line (Ki-SVC1), derived from the rat seminal vesicle epithelium, was investigated. In these experimental conditions, the cell treatment with RA induced a decrease of the proliferation rate, apoptosis and a marked reduction of both anchorage-independent growth and tumorigenicity. These biological responses were either preceded or associated with important changes in adenylate cyclase/protein kinase C signaling pathways, the activation of important apoptosis-linked genes and a marked decrease of the v-Ki-ras p21 protein. The significance of these findings is discussed.


Assuntos
Antineoplásicos/farmacologia , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Tretinoína/farmacologia , Adenilil Ciclases/metabolismo , Animais , Apoptose/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Linhagem Celular Transformada , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , AMP Cíclico/análise , Citosol/efeitos dos fármacos , Citosol/metabolismo , Relação Dose-Resposta a Droga , Regulação para Baixo , Hemangiossarcoma/patologia , Transplante de Neoplasias , Proteína Quinase C/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/análise , Proteínas Proto-Oncogênicas p21(ras)/genética , RNA Mensageiro/análise , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacos
5.
Cell Prolif ; 37(6): 413-26, 2004 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15548174

RESUMO

3-O-Methylfunicone (OMF) is a secondary metabolite produced by the soil fungus Penicillium pinophilum which has cytostatic properties. The aim of this study was to investigate the mechanisms by which such properties are exerted, with special reference to any anti-proliferative and apoptotic potential, on HeLa cells. OMF treatment caused about 44% inhibition of cell growth after 24 h, and modifications in the tubulin fibre organization. In addition, a significant increase in p21 mRNA expression and a decrease in cyclin D1 and Cdk4 mRNA expression resulted at the same time. Apoptosis induction was demonstrated by the annexin V assay, cytofluorimetric analysis of the DNA content of the sub-G1 fraction and DNA laddering. Taken together, our data showed that the compound inhibits proliferation of HeLa cells by several mechanisms, such as disruption of tubulin fibres, cell cycle arrest and apoptosis induction. The capacity of the compound to affect the cell cycle and to modulate apoptosis is indicative of a potential for the development of a new agent for cancer chemotherapy.


Assuntos
Citotoxinas/toxicidade , Inibidores do Crescimento/toxicidade , Penicillium/metabolismo , Pironas/toxicidade , Anexina A5/metabolismo , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/fisiologia , Proteínas de Ciclo Celular/genética , Divisão Celular/efeitos dos fármacos , Divisão Celular/fisiologia , Ciclina D1/genética , Quinase 4 Dependente de Ciclina , Inibidor de Quinase Dependente de Ciclina p21 , Quinases Ciclina-Dependentes/genética , Fragmentação do DNA/efeitos dos fármacos , Fragmentação do DNA/fisiologia , Ensaios de Seleção de Medicamentos Antitumorais , Células HeLa , Humanos , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Estrutura Molecular , Proteínas Proto-Oncogênicas/genética , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Tubulina (Proteína)/efeitos dos fármacos , Tubulina (Proteína)/metabolismo
6.
J Mol Endocrinol ; 24(1): 83-94, 2000 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10657000

RESUMO

Retinoic acid (RA) and sodium butyrate (NaB) are regulators of cell growth and differentiation. We studied their effect on normal (SVC1) or v-Ki-ras-transformed (Ki-SVC1) rat seminal vesicle (SV) epithelial cell lines. The treatment of these cells with 10(-((7( M RA did not produce significant changes in the morphological and biochemical parameters analyzed. When RA was used in combination with 2 mM NaB, the treatment induced substantial morphological changes, apoptosis-independent growth arrest, up-regulation of tissue transglutaminase (tTGase), and down-regulation of beta and gamma RA receptor (RAR) mRNA expression. The same cells did not express RAR alpha either before or after NaB/RA treatment. A similar treatment did not change the amount of mRNA coding for the protein SV-IV (a typical differentiation marker of the SV epithelium) in normal or ras-transformed cells nor the level of v-Ki-ras mRNA in Ki-SVC1 cells. These findings suggest that a defective RA/RARs signaling pathway is probably the biochemical condition that underlies the unresponsiveness to RA of our in vitro culture system, and indirectly points to the possibility that the NaB/RA-induced effects were brought about by a cooperation at the transcription level between the histone deacetylase inhibitory activity of NaB and the ability of RA/RAR to modulate the expression of various genes involved in the control of cell growth and differentiation.


Assuntos
Butiratos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Glândulas Seminais/efeitos dos fármacos , Tretinoína/farmacologia , Animais , Apoptose , Sequência de Bases , Linhagem Celular Transformada , Primers do DNA , Regulação para Baixo , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Genes ras , Masculino , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Receptores do Ácido Retinoico/genética , Receptores do Ácido Retinoico/metabolismo , Glândulas Seminais/citologia , Glândulas Seminais/metabolismo , Transglutaminases/metabolismo
7.
Res Microbiol ; 150(1): 13-9, 1999.
Artigo em Inglês | MEDLINE | ID: mdl-10096130

RESUMO

This project focused on the effects of aflatoxin B1 (AFB1), a food-contaminating mycotoxin produced by fungi, genus Aspergillus, on the release and genetic expression of some important cytokines, i.e., (interleukin-1 alpha (IL-1 alpha), IL-6, tumor necrosis factor-alpha (TNF alpha)) by human monocytes. Monocytes, preincubated for different time periods with concentrations of AFB1 ranging from 0.01 to 1.0 pg/mL, were then activated with bacterial lipopolysaccharide. Cytokine levels were measured by immunoassay and mRNA by cDNA amplification. Pretreatment of monocytes with AFB1 resulted in a decrease in IL-1, IL-6 and TNF alpha release already at a concentration of 0.05 pg/mL. The gene expression of the cytokines considered was drastically affected by treatment with AFB1. In fact, AFB1 completely blocked the transcription of IL-1 alpha, IL-6 and TNF alpha mRNAs, while it did not affect beta-actin mRNA at the concentrations used. It therefore appears that AFB1 exerts its effect on cytokine release through selective inhibition of specific mRNA, without affecting general protein synthesis.


Assuntos
Aflatoxina B1/toxicidade , Aspergillus/metabolismo , Carcinógenos/toxicidade , Citocinas/efeitos dos fármacos , Leucócitos Mononucleares/efeitos dos fármacos , Citocinas/biossíntese , Relação Dose-Resposta a Droga , Humanos , Leucócitos Mononucleares/imunologia , Fatores de Tempo
8.
Arch Dermatol Res ; 293(8): 407-13, 2001 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-11686516

RESUMO

We showed the ability of human dermal fibroblasts to take up Malassezia furfur and the effect of ketoconazole and cytoskeleton inhibitors, including cytochalasin D and colchicine, on invasivity. Engulfment was evaluated by May Grunwald Giemsa stain and confirmed by acridine orange staining and electron microscopy. Both revealed the different steps of engulfment, including a fusion event between lysosomes and phagosomes containing M. furfur. Subinhibitory concentrations of ketoconazole (5 microg/ml) reduced the invasive capacity compared to controls (52.0+/-6.3 vs 10.0+/-1.2). M. furfur induced changes in the cytoskeleton of human dermal fibroblasts, with signs of disaggregation of actin fibres. We also studied the effect of the cytoskeleton inhibitors, cytochalasin D (1 microg/ml) and colchicine (1 microg/ml), on engulfment. Cytochalasin D, an inhibitor of actin polymers, inhibited the uptake of M. furfur by human dermal fibroblasts. Colchicine, a microtubule inhibitor, reduced the uptake of M. furfur less markedly. This suggests that the process of engulfment is F-actin-dependent, but the integrity of microtubules is also important in "non-professional" phagocytic cells such as dermal fibroblasts.


Assuntos
Colchicina/farmacologia , Fibroblastos/microbiologia , Fibroblastos/fisiologia , Malassezia/fisiologia , Micoses/microbiologia , Fenômenos Fisiológicos da Pele , Pele/microbiologia , Antifúngicos/farmacologia , Células Cultivadas , Citocalasina D/farmacologia , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/ultraestrutura , Humanos , Cetoconazol/farmacologia , Microscopia Eletrônica , Pele/citologia
9.
Arch Dermatol Res ; 293(8): 414-9, 2001 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-11686517

RESUMO

The lipophilic yeast Malassezia furfur is a member of the cutaneous microbiota, also associated with several chronic diseases such as pityriasis versicolor, folliculitis, seborrhoeic dermatitis, and some forms of atopic dermatitis, psoriasis and confluent and reticulate papillomatosis. In this study we determined the immunomodulatory and invasive capacity of M. furfur in a human keratinocyte cell culture, HaCat. At a yeast cell to HaCat ratio of 30:1, M. furfur penetration was only 30% with poor phagolysosome fusion and with cytoskeleton modification. Transglutaminase I gene expression was also inhibited, supporting the hypothesis that M. furfur causes an initial break in the barrier function of the epidermis. Moreover, we demonstrated that M. furfur modulates proinflammatory and immunomodulatory cytokine synthesis by downregulating IL-1alpha and by inhibiting IL-6 and TNF-alpha and by upregulating IL-10 and TGF-beta1. The suppressed inflammatory response induced by M. furfur may play a role in chronic disease.


Assuntos
Citocinas/metabolismo , Citoesqueleto/ultraestrutura , Molécula 1 de Adesão Intercelular/metabolismo , Queratinócitos/metabolismo , Queratinócitos/microbiologia , Malassezia/fisiologia , Divisão Celular , Linhagem Celular , Citoesqueleto/fisiologia , Expressão Gênica/fisiologia , Humanos , Queratinócitos/citologia , Transglutaminases/genética
10.
Arch Dermatol Res ; 304(3): 237-40, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22037626

RESUMO

Pemphigus is an autoimmune blistering disease characterized by severe and chronic course, histopathologically characterized by infiltration of a large quantity of eosinophils, neutrophils, and activated Th1 and Th2 cells around the blister. Polarization of Th cells to Th1 or Th2 phenotypes, a critical aspect of cell-mediated immunity, is influenced by production of early cytokines, including osteopontin. To determine the involvement of osteopontin in pemphigus vulgaris patients in active stage of the disease, auto-antibodies to desmoglein-1 and desmoglein-3 and plasmatic osteopontin levels were examined by ELISA tests. In this work, significant plasmatic level of osteopontin in PV patients with active stage of disease were found particularly in those patients with both skin and oral pemphigus. OPN might drive the immune responses playing an important role in pemphigus onset.


Assuntos
Autoanticorpos/sangue , Osteopontina/sangue , Pênfigo/sangue , Adulto , Idoso , Desmogleína 1/imunologia , Desmogleína 3/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pênfigo/imunologia
11.
Arch Dermatol Res ; 304(6): 481-5, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22684781

RESUMO

Psoriasis is a chronic skin inflammatory disease in which a pleiotropic cytokine, tumor necrosis factor alpha (TNF-α), plays a central role, as demonstrated by the clinical success of anti-TNF-α therapy. Among the multiple effects of TNF-α on keratinocytes, the induction of matrix metalloproteinase-9 (MMP-9), a collagenase implicated in joint inflammation, might be one of the key mechanisms in psoriasis pathogenesis. Interestingly, MMP-9 expression can be enhanced also by osteopontin (OPN), a glycosylated protein whose levels are increased in skin and peripheral blood mononuclear cells (PBMC) of psoriasis patients. The aim of the current study is to investigate the relationship between OPN, MMP-9 and TNF-α in psoriasis. Our survey identified high levels of both OPN and MMP-9 in PBMC as well as skin of psoriatic patients with respect to healthy controls. Significant reduction of OPN and MMP-9 levels in PBMC, plasma and lesional skin of psoriasis patients was observed after 24 weeks of anti-TNF-α therapy. Moreover, OPN and MMP-9 were enhanced by TNF-α and down-regulated by anti-TNF-α treatment in healthy PBMC. These findings may suggest that OPN and MMP-9 may be regulated by TNF-α, indicating a possible role in the pathogenesis of psoriasis.


Assuntos
Metaloproteinase 9 da Matriz/sangue , Osteopontina/sangue , Psoríase/sangue , Psoríase/tratamento farmacológico , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Humanos , Leucócitos Mononucleares/química , Metaloproteinase 9 da Matriz/fisiologia , Osteopontina/fisiologia , Psoríase/etiologia
12.
Cell Prolif ; 44(5): 401-9, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-21951283

RESUMO

OBJECTIVES: Cancer stem cells make up a subpopulation of cells within tumours that drive tumour initiation, growth and recurrence. They are resistant to many current types of cancer treatment, causing failure of such therapeutic approaches, including chemotherapy and radiotherapy. In the study described here, anti-proliferative effects of 3-O-methylfunicone (OMF), a metabolite from Penicillium pinophilum, were investigated on human breast cancer MCF-7 cells and cancer stem cells selected as mammospheres derived from MCF-7s. MATERIALS AND METHODS: Stemness markers were analysed on isolated mammospheres showing positive expression of CD24, CD29, CD44, CD133, CD184 and CD338. Cell proliferation and apoptosis were analysed by flow cytometry and RT-PCR. Cell colony formation assays were performed to evaluate colony formation of mammospheres. RESULTS AND CONCLUSION: OMF treatment affected both MCF-7 and mammosphere growth, inducing apoptosis. In addition, OMF strongly reduced stemness markers and survivin, hTERT and Nanog-1 gene expression. Growth of colonies in soft-agar was significantly affected by OMF treatment, too. Lastly, we tested ability of MCF-7 cells to form mammospheres after treatment with OMF or cisplatin, demonstrating that OMF treatment resulted in drastic reduction in number of mammospheres. These results introduce OMF as an effective molecule in suppressing breast cancer stem cells.


Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Células-Tronco Neoplásicas/efeitos dos fármacos , Pironas/farmacologia , Antineoplásicos/isolamento & purificação , Apoptose/efeitos dos fármacos , Sequência de Bases , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Primers do DNA/genética , Feminino , Expressão Gênica/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Humanos , Proteínas Inibidoras de Apoptose/genética , Proteína Homeobox Nanog , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Penicillium/química , Reação em Cadeia da Polimerase , Pironas/isolamento & purificação , Esferoides Celulares/efeitos dos fármacos , Esferoides Celulares/patologia , Survivina , Telomerase/genética , Ensaio Tumoral de Célula-Tronco
13.
Cell Prolif ; 43(2): 114-23, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20447056

RESUMO

OBJECTIVES: 3-O-methylfunicone (OMF), a secondary metabolite produced by Penicillium pinophilum, affects cell proliferation and motility in a variety of human solid tumours. The aim of this study was to demonstrate whether OMF has the ability to arrest cell division and motility, in a human mesothelioma cell line. Malignant mesothelioma is an aggressive cancer that does not respond to standard therapies the cells of which are considered to be highly resistant to apoptosis. MATERIAL AND METHODS: Cell motility and invasion were measured using a modified Boyden chamber. Gene expression was examined by RT-PCR, while ERK1/2 was investigated by Western blot analysis. All experiments were also performed on primary cultures of mesothelial cells. RESULTS: The present study shows that OMF inhibited motility of the NCI mesothelioma cell line by modulating ERK signalling activity, and affected alphaVbeta5 integrin and MMP-2 expression, inducing marked downregulation at both mRNA and protein levels. Substantial downregulation of VEGF gene expression was also demonstrated. These effects were not observed in normal mesothelial cell cultures. CONCLUSION: OMF may have potential as a naturally derived anti-tumour drug for treatment of mesothelioma.


Assuntos
Movimento Celular/efeitos dos fármacos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Pironas/farmacologia , Linhagem Celular Tumoral , Regulação para Baixo , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Mesotelioma/genética , Mesotelioma/metabolismo , Penicillium/metabolismo , Receptores de Vitronectina/metabolismo , Transdução de Sinais/genética
14.
Cell Prolif ; 42(4): 541-53, 2009 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-19486013

RESUMO

OBJECTIVES: Melanoma cells take advantage of impaired ability to undergo programmed cell death in response to different external stimuli and chemotherapeutic drugs; this makes prevention of tumour progression very difficult. The aim of this study was to demonstrate whether 3-O-methylfunicone (OMF), a metabolite of Penicillium pinophilum, has the ability to arrest cell population growth and to induce apoptosis in A375P (parental) and A375M (metastasis derivatived) melanoma cell lines. MATERIALS AND METHODS: Cell proliferation and apoptosis were analysed by flow cytometry, DNA fragmentation, caspase-3 and caspase-9 activation, and PARP-1 cleavage. RESULTS: We demonstrated that OMF affected cell proliferation in a time- and dose-dependent manner, reaching the best effect at concentration of 80 microg/ml for 24 h. Flow cytometry revealed that OMF caused significant G(2) phase arrest, which was associated with marked decrease in cyclin B1/p34(cdc2) complex and p21 induction. OMF also induced marked decrease of survivin expression. Reduced levels of apoptosis were evident after silencing p21 expression in both cell lines. Finally, the effect exercised by OMF on hTERT and TEP-1 gene expression confirmed the ability of this molecule to interfere with replicative ability of cells. CONCLUSIONS: The results reported here seem to suggest that OMF as a promising molecule to include in strategies for treatment of melanoma.


Assuntos
Proliferação de Células/efeitos dos fármacos , Melanoma/tratamento farmacológico , Penicillium/metabolismo , Pironas/farmacologia , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 9/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Poli(ADP-Ribose) Polimerase-1 , Poli(ADP-Ribose) Polimerases/metabolismo , Pironas/isolamento & purificação , RNA Interferente Pequeno/genética , Proteínas rho de Ligação ao GTP/genética
15.
Clin Diagn Lab Immunol ; 8(1): 206-8, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11139223

RESUMO

Treatment of human spermatozoa with porins or lipopolysaccharide (LPS) increases spontaneous apoptosis in these cells. Porins and LPS were extracted from Salmonella enterica serovar Typhimurium and Pasteurella multocida and were mixed with human spermatozoa for detection of levels of apoptosis.


Assuntos
Apoptose , Lipopolissacarídeos/metabolismo , Porinas/metabolismo , Espermatozoides/patologia , Sobrevivência Celular/efeitos dos fármacos , Fragmentação do DNA , Humanos , Lipopolissacarídeos/farmacologia , Masculino , Pasteurella multocida , Polimixina B/farmacologia , Porinas/farmacologia , Salmonella typhimurium , Espermatozoides/efeitos dos fármacos
16.
Br J Dermatol ; 150(6): 1070-80, 2004 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15214891

RESUMO

BACKGROUND: Captopril is an angiotensin-converting enzyme inhibitor with sulphydryl groups in its chemical structure. It is commonly used as an antihypertensive drug. The occurrence of pemphigus vulgaris has repeatedly been reported in patients receiving captopril. The capacity of captopril and pemphigus serum to induce acantholysis, in vivo or in vitro, has been demonstrated experimentally. OBJECTIVES: To show that captopril and pemphigus serum, acting by a biochemical and immunological mechanism, respectively, trigger apoptosis. METHODS: Human keratinocyte cells were treated with 15 mmol L-1 captopril or with pemphigus serum. DNA was extracted and the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labelling method was used to detect apoptosis. RESULTS: DNA fragmentation occurred after 72 h of treatment. Increased expression of p53, c-myc and inducible nitric oxide (NO) synthase (iNOS) mRNA were observed by polymerase chain reaction (PCR) in the treated cells compared with the untreated ones. The increase in iNOS gene expression was associated with overproduction of NO. Moreover, the addition of 1 mmol L-1N-monomethyl-L-arginine, a structural analogue of arginine, reduced nitrite levels by about 70% in cells treated with captopril or pemphigus serum. Western blot analysis revealed an overexpression of heat shock protein 70 (hsp70) in cells treated with captopril or pemphigus serum. Finally, total inhibition of the keratinocyte transglutaminase gene was shown by PCR analysis in the same samples, compared with control cells. CONCLUSIONS: These data demonstrate the involvement of apoptosis in keratinocytes treated with captopril or pemphigus serum, with induction of the iNOS gene and hsp70 in the cascade of events leading to programmed cell death.


Assuntos
Inibidores da Enzima Conversora de Angiotensina/efeitos adversos , Apoptose , Captopril/efeitos adversos , Proteínas de Choque Térmico HSP70/análise , Queratinócitos/efeitos dos fármacos , Óxido Nítrico Sintase/análise , Pênfigo/induzido quimicamente , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Autoanticorpos/farmacologia , Western Blotting/métodos , Captopril/farmacologia , Células Cultivadas , Meios de Cultivo Condicionados , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/ultraestrutura , Fragmentação do DNA , Relação Dose-Resposta a Droga , Enalapril/efeitos adversos , Enalapril/farmacologia , Ativação Enzimática , Humanos , Queratinócitos/metabolismo , Queratinócitos/patologia , Microscopia de Fluorescência , Óxido Nítrico Sintase Tipo II , Nitritos/análise , Pênfigo/imunologia , Pênfigo/patologia , Proteínas Proto-Oncogênicas c-myc/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transglutaminases/análise , Proteína Supressora de Tumor p53/análise
17.
Br J Dermatol ; 148(3): 424-33, 2003 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-12653733

RESUMO

BACKGROUND: Recent evidence assigns the vitronectin receptors (VnRs) an important role in regulating tumour cell invasion and dissemination. In vivo and in vitro studies document that all trans-retinoid acids (ATRAs) inhibit growth-inducing apoptosis in melanomas. OBJECTIVES: We have analysed the effects of ATRA treatment on melanoma cell adhesion and motility. METHODS: Human M14 melanoma cells were treated with 10 micromol L-1 ATRA for different times and stained with rhodamine-phalloidin to analyse the effect of treatment on cytoskeleton organization. Cell adhesion and cell migration assays were performed to analyse the role of VnRs in the ATRA-induced early stages of apoptosis. VnR expression was evaluated by Western blot, immunoprecipitation and immunocytochemistry assays. RESULTS: First, using an annexin V assay, we found that apoptosis was triggered by 48 h with 10 micromol L-1 ATRA exposure. At this time point, decrease in the F-actin polymerization as well as inhibition of cell adhesive ability to vitronectin (Vn) was exerted by ATRA treatment. In the presence of serum, exposure to 10 micromol L-1 ATRA for 48 h produced a dramatic inhibition of the cell adhesion ability that was comparable with that exerted by untreated cells preincubated with anti-alpha(v)beta(3) or anti-alpha(v)beta(5) VnR monoclonal antibodies. Functionally, the treatment of melanoma cells with 10 micromol L-1 ATRA for 48 h causes an inhibition of directional cell migration towards Vn-coated filters. Therefore, we analysed the effect of ATRA on the VnR expression. Both alpha(v)beta(3) and alpha(v)beta(5) VnR levels were reduced upon exposure to 10 micromol L-1 ATRA for 48 h as shown by Western blot, immunoprecipitation and immunocytochemistry assays. CONCLUSIONS: Altogether, our data indicate that treatment of M14 melanoma cells with ATRA downregulates VnR expression and that this reduction is closely correlated with the ATRA-dependent inhibition of actin-fibre organization, cell adhesion and migration. Although the mechanism by which ATRA regulates the expression of VnR in M14 melanoma cells needs further elucidation, this system may represent a model for understanding the molecular basis of ATRA therapy in melanoma.


Assuntos
Antineoplásicos/farmacologia , Apoptose , Integrina alfaVbeta3/metabolismo , Melanoma Experimental/metabolismo , Proteínas de Neoplasias/metabolismo , Tretinoína/farmacologia , Western Blotting/métodos , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Citoesqueleto/efeitos dos fármacos , DNA de Neoplasias/análise , Regulação para Baixo/efeitos dos fármacos , Humanos , Testes de Precipitina , Células Tumorais Cultivadas/efeitos dos fármacos
18.
Br J Dermatol ; 141(6): 1033-9, 1999 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-10606848

RESUMO

Pemphigus is an autoimmune disease where both endogenous (genetic) and exogenous (environmental) factors play a part. Viral infections, in particular herpesvirus infections, have been identified as a possible triggering factor for pemphigus. In this study, using the polymerase chain reaction, we studied peripheral blood mononuclear cells (PBMC) and skin biopsies from patients with pemphigus, and in some of these were able to demonstrate the presence of DNA sequences of herpes simplex virus 1/2 (50% in PBMC and 71% in skin biopsies), Epstein-Barr virus (15% in PBMC and 5% in skin biopsies) and human herpesvirus 6 (20% in PBMC only). However, the inability to detect herpesvirus DNA consistently in these cases suggests that viral infection may only be an occasional factor triggering the outbreak or exacerbation of the disease. The possible role of interferons and interleukins in the pathogenesis of virus-induced pemphigus is discussed.


Assuntos
DNA Viral/análise , Herpesviridae/isolamento & purificação , Leucócitos Mononucleares/virologia , Pênfigo/virologia , Pele/virologia , Adulto , Idoso , Anticorpos Antivirais/sangue , Criança , Feminino , Herpesviridae/imunologia , Herpesvirus Humano 4/isolamento & purificação , Herpesvirus Humano 6/isolamento & purificação , Teste de Histocompatibilidade , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Simplexvirus/isolamento & purificação
19.
J Clin Immunol ; 21(3): 210-7, 2001 May.
Artigo em Inglês | MEDLINE | ID: mdl-11403228

RESUMO

Our objective was to investigate the phenotype of helper T cells in the peripheral blood of patients with systemic sclerosis (SSc). PBMC from 15 patients with SSc and 15 sex- and age-matched controls were investigated for lymphocyte subpopulations (CD3, CD4, CD8, CD19, CD16/CD56, CD3-DR); IL-2, IL-4, and IFN-gamma mRNAs; and the relative cytokines in their cytoplasm. The last assay was carried out both in unstimulated and in PMA-activated PBMC. SSc patients presented a higher percentage of activated T cells, CD3+ DR+ (19.7 +/- 9.9 vs 5.1 +/- 2.5%; P < 0.0001); 12 of them presented IFN-gamma mRNA-positive cells; and none IL-2 or IL-4 mRNAs. Under basal conditions, PBMC from six SSc patients contained IL-2, IL-4, and IFN-gamma (i.e., they showed both Th1 and Th2 activation), and 1 IFN-gamma only. PMA-stimulated PBMC of patients differed from those of controls only in the increased percentage of IFN-gamma positive cells (52 +/- 12 vs 37 +/- 11%; P < 0.01). Our study demonstrates that Thl activation occurs in the peripheral blood of SSc patients. This evidence must be faced with from both a pathogenetic and a therapeutical point of view.


Assuntos
Escleroderma Sistêmico/imunologia , Células Th1/imunologia , Células Th2/imunologia , Adulto , Estudos de Casos e Controles , Feminino , Humanos , Técnicas In Vitro , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-2/genética , Interleucina-2/metabolismo , Interleucina-4/genética , Interleucina-4/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Escleroderma Sistêmico/genética , Acetato de Tetradecanoilforbol/farmacologia , Células Th1/efeitos dos fármacos , Células Th2/efeitos dos fármacos
20.
Infect Immun ; 67(9): 4794-800, 1999 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-10456933

RESUMO

Micromolar concentrations of porin, purified from the outer membranes of Pseudomonas aeruginosa, induced in vitro the classic morphological and biochemical signs of apoptosis in an epithelial cell line (SVC1) derived from the rat seminal vesicle secretory epithelium. The programmed cell death (PCD) was p53 independent and associated with significant decrease of bcl-2 expression, a marked increase of c-myc transcriptional activity, and an absence of the mRNA coding for tissue transglutaminase. The Ca(2+) influx, caused by the porin treatment of SVC1 cells, appears to play an important role in the triggering of apoptosis in our biological model. The possibility that the porin property of inducing PCD plays a role in the infertility of individuals chronically infected by gram-negative bacteria is discussed.


Assuntos
Apoptose , Células Epiteliais/patologia , Porinas/metabolismo , Pseudomonas aeruginosa , Glândulas Seminais/patologia , Animais , Cálcio/metabolismo , Linhagem Celular , Membrana Celular/fisiologia , Humanos , Masculino , Porinas/farmacologia , Proteínas Proto-Oncogênicas c-myc/metabolismo , Ratos , Proteína Supressora de Tumor p53/metabolismo
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