RESUMO
Schistosomiasis is a chronic parasitic disease caused by trematodes of the genus Schistosoma, endemic in tropical and subtropical regions. The hepatic pathology of this parasitic disease could develop complications, such as fibrosis and cirrhosis, which can be fatal. The Venezuelan endemic area is considered as one of low transmission, which complicates the detection of infected individuals and signals the importance of improving the sensitivity of immunodiagnostic methods. Using ELISA, an evaluation was conducted of IgM and IgG responses to soluble antigens of eggs and female worms (SEA and SFWA) and excretion-secretion products of eggs and female worms (ESPE and ESPAW) in infected Balb/c mice with different parasitic burden and infection times. A high positivity rate by IgM detection was observed for all antigen preparations in 7-week infections (100% by SEA, SFWA, ESPE, and ESPWA in high parasitic burden) as well as a reduction of this immunoglobulin in chronic infection. Positivity rate for IgG detection was higher in 20-week infections (100% by ESPE in low burden, 100% by SEA and ESPE in medium burden, and 100% by ESPE and ESPAW in high burden conditions). The potential use of combined or unique antigenic preparations associated with IgM or IgG for detection of active infection, regardless the parasitic burden, was demonstrated. Differences between immunoglobulin responses show its application for phase-specific diagnosis.
Assuntos
Anticorpos Anti-Helmínticos/biossíntese , Imunoglobulina G/biossíntese , Imunoglobulina M/biossíntese , Schistosoma mansoni/imunologia , Esquistossomose mansoni/imunologia , Animais , Anticorpos Anti-Helmínticos/sangue , Cricetinae , Ensaio de Imunoadsorção Enzimática , Fezes/parasitologia , Feminino , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Mesocricetus , Camundongos , Camundongos Endogâmicos BALB C , Testes de Precipitina , Esquistossomose mansoni/diagnóstico , Esquistossomose mansoni/parasitologiaRESUMO
The increase in the intracellular Ca(2+) concentration ([Ca(2+)]i) is the key variable for many different processes, ranging from regulation of cell proliferation to apoptosis. In this work we demonstrated that the sphingolipid sphingosine (Sph) increases the [Ca(2+)]i by inhibiting the sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA), in a similar manner to thapsigargin (Tg), a specific inhibitor of this Ca(2+) pump. The results showed that addition of sphingosine produced a release of Ca(2+) from the endoplasmic reticulum followed by a Ca(2+) entrance from the outside mileu. The results presented in this work support that this sphingolipid could control the activity of the SERCA, and hence sphingosine may participate in the regulation of [Ca(2+)]I in mammalian cells.
Assuntos
Cálcio/metabolismo , Neoplasias/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Esfingosina/metabolismo , Linhagem Celular Tumoral , Retículo Endoplasmático/metabolismo , Ativação Enzimática , HumanosRESUMO
PURPOSE: In search for new drugs derived from natural products for the possible treatment of cancer, we studied the action of agelasine B, a compound purified from a marine sponge Agelas clathrodes. METHODS: Agelasine B was purified from a marine sponge Agelas clathrodes and assayed for cytotoxicity by MTT on two human breast cancer cells (MCF-7 and SKBr3), on a prostate cancer cells (PC-3) and on human fibroblasts. Changes in the intracellular Ca(2+) concentrations were assessed with FURA 2 and by confocal microscopy. Determination of Ca(2+)-ATPase activity was followed by Pi measurements. Changes in the mitochondria electrochemical potential was followed with Rhodamine 123. Apoptosis and DNA fragmentation were determined by TUNEL experiments. RESULTS: Upon agelasine B treatment, cell viability of both human breast cancer cell lines was one order of magnitude lower as compared with fibroblasts (IC(50) for MCF-7 = 2.99 µM; SKBr3: IC(50) = 3.22 µM vs. fibroblasts: IC(50) = 32.91 µM), while the IC(50) for PC-3 IC(50) = 6.86 µM. Agelasine B induced a large increase in the intracellular Ca(2+) concentration in MCF-7, SKBr3, and PC-3 cells. By the use of confocal microscopy coupled to a perfusion system, we could observe that this toxin releases Ca(2+) from the endoplasmic reticulum (ER). We also demonstrated that agelasine B produces a potent inhibition of the ER Ca(2+)-ATPase (SERCA), and that this compound induced the fragmentation of DNA. Accordingly, agelasine B reduced the expression of the anti-apoptotic protein Bcl-2 and was able to activate caspase 8, without affecting the activity of caspase 7. CONCLUSIONS: Agelasine B in MCF-7 cells induce the activation of apoptosis in response to a sustained increase in the [Ca(2+)]( i ) after blocking the SERCA activity. The reproduction of the effects of agelasine B on cell viability and on the [Ca(2+)]( I ) obtained on SKBr3 and PC-3 cancer cells strongly suggests the generality of the mechanism of action of this toxin.