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1.
Vet Parasitol ; 223: 127-32, 2016 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-27198789

RESUMO

Toll like receptors (TLRs) are involved in the modulation of diverse host genes expression through a complex network of signalling events that allow for an appropriate response to a microbial pathogen. In the present work we used TLR6KO mice in order to study the role of TLR6 in the immune discrimination of lipids from two Babesia bovis strains, attenuated R1A (LA) and virulent S2P (LV), and the consequent macrophage activation. We demonstrated that TLR6 is required for lipid body induction in murine peritoneal macrophages by both LA and LV. Interestingly, as regards IL-10 and COX-2/PGE2 pathway induction by LA and LV, we observed differences in the biological effects produced by these lipid extracts. Our results indicate a role of TLR6 in the down-modulation of these immunoregulators only in the case of LA, whereas this receptor was not implicated in pro-inflammatory TNFα, IL-6 and KC release induced by LA. Remarkably, LV did not exert the down-modulatory effect observed for LA, supporting the notion that LA and LV possess different lipid composition that could correlate with the polar pathogenic effect of both B. bovis strains.


Assuntos
Babesia bovis/metabolismo , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/metabolismo , Interleucina-10/metabolismo , Macrófagos Peritoneais/efeitos dos fármacos , Receptor 6 Toll-Like/metabolismo , Animais , Babesia bovis/patogenicidade , Ciclo-Oxigenase 2/genética , Dinoprostona/genética , Interleucina-10/genética , Gotículas Lipídicas/fisiologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Macrófagos Peritoneais/metabolismo , Camundongos , Camundongos Knockout , Receptor 6 Toll-Like/genética , Virulência
2.
Parasitology ; 140(4): 530-40, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23286221

RESUMO

The intra-erythrocytic protozoan Babesia bovis is an economically important pathogen that causes an acute and often fatal infection in adult cattle. Babesiosis limitation depends on the early activation of macrophages, essential cells of the host innate immunity, which can generate an inflammatory response mediated by cytokines and nitric oxide (NO). Herein, we demonstrate in bovine macrophages that lipids from B. bovis attenuated R1A strain (LA) produced a stronger NO release, an early TNFα mRNA induction and 2-fold higher IL-12p35 mRNA levels compared to the lipids of virulent S2P strain (LV). Neither LA nor LV induced anti-inflammatory IL-10. Regarding signalling pathways, we here report that LA induced a significant phosphorylation of p38 and extracellular signal-regulated kinases 1 and 2 (ERK1/2) whereas LV only induced a reduced activation of ERK1/2. Besides, NF-κB was activated by LA and LV, but LA produced an early degradation of the inhibitor IκB. Interestingly, LV and the majority of its lipid fractions, exerted a significant inhibition of concanavalin A-induced peripheral blood mononuclear cell proliferation with respect to LA and its corresponding lipid fractions. In addition, we determined that animals infected with R1A developed a higher increase in IgM anti-phosphatidylcholine than those inoculated with S2P. Collectively, S2P lipids generated a decreased inflammatory response contributing to the evasion of innate immunity. Moreover, since R1A lipids induced a pro-inflammatory profile, we propose these molecules as good candidates for immunoprophylactic strategies against babesiosis.


Assuntos
Babesia bovis/imunologia , Babesiose/veterinária , Interações Hospedeiro-Parasita/imunologia , Lipídeos/imunologia , Macrófagos/imunologia , Transdução de Sinais , Animais , Anti-Inflamatórios/farmacologia , Anticorpos Antifosfolipídeos/sangue , Babesia bovis/química , Babesia bovis/patogenicidade , Babesiose/imunologia , Babesiose/parasitologia , Bovinos , Doenças dos Bovinos/imunologia , Doenças dos Bovinos/parasitologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Leucócitos Mononucleares/citologia , Lipídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/parasitologia
3.
Mol Immunol ; 47(4): 747-55, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19910051

RESUMO

Babesia bovis is an intraerythrocytic apicomplexan protozoa of cattle that causes an acute infection with parasite persistence. Babesiosis limitation depends on macrophages, essential effector cells of the host innate defense, which generate inflammatory cytokines and nitric oxide. Herein, we report quantitative differences in the lipid composition of merozoites from two B. bovis strains with polar behaviour: attenuated R1A and virulent S2P. Accordingly, we observed a distinct inflammatory response induced by the total lipids of R1A (L(A)) and S2P (L(V)) in murine peritoneal macrophages. L(A) and particularly its fractions phosphatidic acid and phosphatidylserine+phosphatidylinositol (PS+PI), produced a strong activation of these cells with lipid body formation, cyclooxygenase-2 expression and pro-inflammatory TNFalpha, IL-6 and KC secretion. Although L(V) did not activate these cells, the corresponding PS+PI fraction induced TNFalpha, IL-6 and KC release. Therefore, these facts might be suggesting the presence of an inhibitor in L(V). Furthermore, the employment of wild type and toll like receptor 2 knockout (TLR2KO) mice allowed us to demonstrate that macrophage activation by the stimulating lipid fractions was mediated through TLR2. Interestingly, only L(A) activated the extracellular signal-regulated kinases 1 and 2 (ERK1/2). Inhibitory studies employing UO126, indicated that the ERK pathway was required for TNFalpha, IL-6 and KC release. In conclusion, the absence of inflammatory response observed with the lipids of S2P virulent strain could constitute an evasion mechanism of the innate immune response enabling parasite establishment in the host.


Assuntos
Babesia bovis/imunologia , Babesia bovis/patogenicidade , Lipídeos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Receptor 2 Toll-Like/imunologia , Animais , Babesia bovis/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Dinoprostona/biossíntese , Indução Enzimática/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/enzimologia , Macrófagos/metabolismo , Macrófagos/parasitologia , Merozoítos/efeitos dos fármacos , Merozoítos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Virulência/efeitos dos fármacos
4.
Parasitology ; 134(Pt 4): 491-502, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17121684

RESUMO

Here we have studied phospholipase A1 (Plase A1) from Trypanosoma cruzi infective stages and it's possible role regarding the interaction with mammalian host cells. Plase A1 was mainly detected as a membrane-bound activity in the infective amastigote and trypomastigote stages, being remarkably higher with respect to the non-infective epimastigotes. It is noteworthy that only the infective stages secreted Plase A1. Moreover, along the differentiation process from epimastigotes into metacyclic trypomastigotes, the secreted enzyme activity increased simultaneously with the appearance of metacyclic forms, as expected. Since this enzyme is predominantly membrane-associated and secreted by the infective stages, Vero cell lipid profile modifications were analysed after interaction with either intact infective parasites or purified T. cruzi Plase A1. Significant changes in Vero cell lipid composition were observed, with the appearance of free fatty acids, diacylglycerol and lysophosphatidylcholine. Concomitantly with the generation of second lipid messengers, host cell protein kinase C activation was demonstrated. These results indicate that T. cruzi Plase A1 could play a critical role in the early events of parasite-host cell interaction that precede invasion.


Assuntos
Metabolismo dos Lipídeos , Fosfolipases A/metabolismo , Proteína Quinase C/metabolismo , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/fisiologia , Animais , Chlorocebus aethiops , Ativação Enzimática , Fosfolipases A1 , Células Vero
5.
Parasitology ; 132(Pt 3): 345-55, 2006 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16238824

RESUMO

With the aim to study proteinases released to the culture medium during Trypanosoma cruzi metacyclogenesis, the presence of cysteine proteinases (CPs) was analysed in culture supernatants obtained throughout the differentiation induced by stimulation of epimastigotes with Triatoma infestans hindgut homogenate. In SDS-gelatin containing gels, an important endopeptidase activity with apparent molecular weight range between 97 and 116 kDa was encountered at pH 6, which was abolished by the specific cysteine proteinase inhibitor E-64 and TLCK, but not by pepstatin, 1,10 phenantroline or PMSF. This novel CP, named TcCPmet, showed affinity to cystatin-Sepharose, denoting its thiol-proteinase character as well as to ConA-Sepharose, indicating it contains N-linked oligosaccharides. However, it presented a different elution pattern on ConA-Sepharose than cruzipain and, in addition, it was not recognized by anti-cruzipain serum, facts that strongly suggest the different nature of both CPs. Moroever, evidence is presented indicating that TcCPmet was able to hydrolyse the same chromogenic peptides as cruzipain at optimal alkaline pH values, although with a different order of effectiveness. Our results indicate the presence of a novel CP secreted by metacyclic trypomastigotes and reinforces the important role of these enzymes in metacyclogenesis.


Assuntos
Cisteína Endopeptidases/isolamento & purificação , Estágios do Ciclo de Vida/fisiologia , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/fisiologia , Animais , Western Blotting/veterinária , Cromatografia de Afinidade/veterinária , Reações Cruzadas , Meios de Cultura/química , Técnicas de Cultura/veterinária , Cisteína Endopeptidases/química , Cisteína Endopeptidases/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Inibidores de Proteases/farmacologia , Triatoma/química , Trypanosoma cruzi/crescimento & desenvolvimento
6.
Biochem Biophys Res Commun ; 291(3): 516-21, 2002 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-11855818

RESUMO

During invasion of nonphagocytic cells by Trypanosoma cruzi (T. cruzi), host cell lysosomes are recruited to the plasma membrane attachment site followed by lysosomal enzyme secretion. The membrane trafficking events involved in invasion have not been delineated. We demonstrate here that T. cruzi invasion of nonphagocytic cells was completely abolished by overexpression of a dominant negative mutant of dynamin. Likewise, overexpression of a dominant negative mutant of Rab5, the rate-limiting GTPase for endocytosis, resulted in reduced infection rates compared with cells expressing Rab5 wild-type. Moreover, cells expressing the activated mutant of Rab5 experienced higher infection rates. A similar pattern was also observed when Rab7-transfected cells were examined. Confocal microscopy experiments showed that parasites colocalized with green fluorescent protein-Rab5-positive early endosomes after 5 min of invasion. These data clearly indicate that newly forming T. cruzi phagosomes first interact with an early endosomal compartment and subsequently with other late component markers before lysosomal interaction occurs.


Assuntos
Endossomos/enzimologia , GTP Fosfo-Hidrolases/fisiologia , Trypanosoma cruzi/patogenicidade , Proteínas rab de Ligação ao GTP/fisiologia , Proteínas rab5 de Ligação ao GTP/fisiologia , Animais , Células CHO , Cricetinae , Dinaminas , Endocitose , Células HeLa , Interações Hospedeiro-Parasita , Humanos , Lisossomos/química , Microscopia Confocal , Modelos Biológicos , Fagócitos/fisiologia , Vacúolos/enzimologia , Vacúolos/parasitologia , proteínas de unión al GTP Rab7
7.
Exp Cell Res ; 264(2): 211-8, 2001 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-11262178

RESUMO

Multiple signal transduction events are triggered in the host cell during invasion by the protozoan parasite Trypanosoma cruzi. Here, we report the regulation of host cell phosphatydilinositol 3-kinase (PI3K) and protein kinase B (PKB/Akt) activities by T. cruzi during parasite-host cell interaction. Treatment of nonphagocytic cells (Vero, L(6)E(9), and NIH 3T3) and phagocytic cells (human and J774 murine macrophages) with the selective PI3K inhibitors Wortmannin and LY294002 significantly impaired parasite invasion in a dose-dependent fashion. A strong activation of PI3K and PKB/Akt activities in Vero cells was detected when these cells were incubated with trypomastigotes or their isolated membranes. Consistently, we were unable to detect activation of PI3K or PKB/Akt activities in host cells during epimastigote (noninfective) membrane-host cell interaction. Infection of transiently transfected cells containing an inactive mutant PKB showed a significant inhibition of invasion compared with the active mutant-transfected cells. T. cruzi PI3K-like activity was also required in host cell invasion since treatment of trypomastigotes with PI3K inhibitors prior to infection reduced parasite entry. Taken together, these results indicate that PI3K and PKB/Akt activation in parasites, as in host cells induced by T. cruzi, is an early invasion signal required for successful trypomastigote internalization.


Assuntos
Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Serina-Treonina Quinases , Proteínas Proto-Oncogênicas/metabolismo , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/patogenicidade , Células 3T3 , Androstadienos/farmacologia , Animais , Chlorocebus aethiops , Cromonas/farmacologia , Ativação Enzimática , Inibidores Enzimáticos/farmacologia , Humanos , Camundongos , Morfolinas/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt , Trypanosoma cruzi/efeitos dos fármacos , Células Vero , Wortmanina
8.
J Eukaryot Microbiol ; 44(5): 497-502, 1997.
Artigo em Inglês | MEDLINE | ID: mdl-9304819

RESUMO

Trypanosoma cruzi trypomastigotes survive inside macrophages by promoting fusion between the parasitophorous vacuole and mature host lysosomes upon internalization. Since trypomastigotes can evade the lytic pathway, the earliest steps of endocytosis, such as early endosome fusion, may be affected. To test this hypothesis, we used an in vitro early endosome fusion assay. Our results show that trypomastigote-infected macrophage cytosols cannot promote fusion between early endosomes, compared to mock-infected cytosols (heat-killed trypomastigotes were used in the parasite-macrophage interaction assay). GTP gamma S addition potentiates the fusogenic activity driven by trypomastigote-infected macrophage cytosol-mediated assays, unlike the biphasic fusogenic effect obtained with GTP gamma S treatment of macrophage cytosol controls. Calcium-stimulated early endosome fusogenic processes are not affected in the assays mediated by infected macrophage cytosol. We conclude that GTP-regulated factors, and not calcium-regulated elements, are involved in the inhibition of the early endosome fusogenic process by the trypomastigote-infected macrophage cytosol. This primary impediment to the progress of a normal endocytosis may be a relevant step required for the lysosomal recruitment-fusion of the host lysosomes upon trypomastigote infection and further survival of the parasite within its host.


Assuntos
Citosol/fisiologia , Endossomos/fisiologia , Macrófagos/parasitologia , Fusão de Membrana/fisiologia , Trypanosoma cruzi/fisiologia , Animais , Cálcio/fisiologia , Citosol/parasitologia , Guanosina 5'-O-(3-Tiotrifosfato)/farmacologia , Guanosina Trifosfato/fisiologia , Macrófagos/ultraestrutura , Camundongos
9.
Exp Parasitol ; 83(2): 240-9, 1996 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-8682192

RESUMO

The possible role of intracellular Ca2+ level on Trypanosoma cruzi differentiation was explored. The addition to epimastigotes of a Triatoma infestans intestinal homogenate, which that triggers off the differentiation to the infective metacyclic form, induced a sudden rise in [Ca2+]i from the basal value, 94 +/- 28 to 584 +/- 43 nmole/liter. This increase was not affected by the presence of EGTA in the medium. Trypsin-treated intestinal homogenate did not alter the [Ca2+]i of epimastigotes. Calmodulin inhibitors (Calmidazolium, Trifluoperazine, and Chlorpromazine) blocked differentiation. Although the calcium ionophore ionomycin increased [Ca2+]i to 342 +/- 29 nmole/liter, it was unable to induce differentiation by itself. BAY K8644 and Methoxyverapamil (agonist and antagonist of Ca2+ channels, respectively) were unable to affect [Ca2+]i by themselves, or when added to stimulated parasites, and did not exert a stimulatory or inhibitory effect on morphogenesis. BAPTA/AM, a Ca2+ chelator, partially blocked the rise in [Ca2+]i and morphogenesis; this effect was reversed by ionomycin. The requirement of intracellular Ca2+ on epimastigote multiplication was also evaluated. The addition of EGTA to the culture medium led to a decrease in epimastigote multiplication till it practically ceased in the sixth passage. When such parasites were transferred to LIT they partially recovered the growth rate. Parasites from passages III, IV, and V in the Ca(2+)-depleted medium maintained their basal [Ca2+]i, but when treated with the intestinal homogenate, the rise in [Ca2+]i was abrogated. Accordingly, the differentiation percentages of such parasites dropped significantly compared with controls.


Assuntos
Cálcio/metabolismo , Trypanosoma cruzi/crescimento & desenvolvimento , Éster Metílico do Ácido 3-Piridinacarboxílico, 1,4-Di-Hidro-2,6-Dimetil-5-Nitro-4-(2-(Trifluormetil)fenil)/farmacologia , Animais , Calcimicina/farmacologia , Agonistas dos Canais de Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Quelantes/farmacologia , Clorpromazina/farmacologia , Antagonistas de Dopamina/farmacologia , Ácido Egtázico/análogos & derivados , Ácido Egtázico/farmacologia , Inibidores Enzimáticos/farmacologia , Ácido Gálico/análogos & derivados , Ácido Gálico/farmacologia , Galopamil/farmacologia , Imidazóis/farmacologia , Ionomicina/farmacologia , Ionóforos/farmacologia , Morfogênese/efeitos dos fármacos , Triatoma , Trifluoperazina/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/metabolismo
10.
Biochem Biophys Res Commun ; 222(2): 386-9, 1996 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-8670214

RESUMO

The interaction between Trypanosoma cruzi, the protozoan causative of Chagas's disease, and its host cell is a complex process in which multiple signals including those of Ca2+ are involved. Macrophage cytosolic Ca2+ levels were studied during the interaction of these cells with metacyclic trypomastigotes of T. cruzi, since this event is an initial step in the natural infection. In this model we detected an increase in the macrophage cytosolic Ca2+ concentration after infection, or incubation with a metacyclic lysate or with isolated membranes, suggesting that these increments could be necessary for parasite invasion. This fact was confirmed by treating macrophages with a Ca2+ chelator or a Ca2+ channel antagonist which decreased the infection percentages while parasitization levels increased after treatment with Ca2+ channel agonist.


Assuntos
Quelantes/farmacologia , Interações Hospedeiro-Parasita , Macrófagos/parasitologia , Trypanosoma cruzi/fisiologia , Éster Metílico do Ácido 3-Piridinacarboxílico, 1,4-Di-Hidro-2,6-Dimetil-5-Nitro-4-(2-(Trifluormetil)fenil)/farmacologia , Animais , Cálcio/metabolismo , Células Cultivadas , Citosol/metabolismo , Ácido Egtázico/análogos & derivados , Ácido Egtázico/farmacologia , Corantes Fluorescentes , Fura-2/análogos & derivados , Galopamil/farmacologia , Ionomicina/farmacologia , Cinética , Macrófagos/efeitos dos fármacos , Macrófagos/fisiologia , Espectrometria de Fluorescência , Fatores de Tempo , Trypanosoma cruzi/patogenicidade
11.
Cell Mol Biol (Noisy-le-grand) ; 41(6): 861-6, 1995 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-8535180

RESUMO

Activity of H(+)-ATPase of Trypanosoma cruzi (RA strain) submitochondrial particles is increased in parasites which have a low spermine content caused by growing in a polyamine free medium or in a medium containing inhibitors of polyamine synthesis. Under these conditions, the proliferation rate is markedly decreased. Kinetics of the enzyme inhibition by spermine indicates a non-competitive inhibition. Spermine, through its action on the H(+)-ATPase hydrophobic environment, affects the enzyme activity. Together with the ATPase protein inhibitor, this could be a mechanism regulating ATP levels needed for the parasite proliferation.


Assuntos
Mitocôndrias/enzimologia , ATPases Translocadoras de Prótons/metabolismo , Espermina/farmacologia , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/crescimento & desenvolvimento , Animais , Cicloeximida/farmacologia , Eflornitina/farmacologia , Cinética , ATPases Translocadoras de Prótons/efeitos dos fármacos , Putrescina/metabolismo , Espermidina/metabolismo , Espermina/metabolismo , Partículas Submitocôndricas/enzimologia , Trypanosoma cruzi/efeitos dos fármacos
12.
Proc Natl Acad Sci U S A ; 90(21): 10140-4, 1993 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-8234267

RESUMO

A peptide from hindguts of the Triatoma hematophagous Chagas insect vector activates adenylyl cyclase activity in Trypanosoma cruzi epimastigote membranes and stimulates the in vitro differentiation of epimastigotes to metacyclic trypomastigotes. Hindguts were obtained from insects fed 2 days earlier with chicken blood. Purification was performed by gel filtration and HPLC on C18 and C4 columns. SDS/PAGE of the purified peptide showed a single band of about 10 kDa. The following sequence was determined for the 20 amino-terminal residues of this peptide: H2N-Met-Leu-Thr-Ala-Glu-Asp-Lys-Lys-Leu-Ile-Gln- Gln-Ala-Trp-Glu-Lys-Ala-Ala-Ser-His. This sequence is identical to the amino terminus of chicken alpha D-globin. On a Western blot, the peptide immunoreacted with a polyclonal antibody against chicken globin D. A synthetic peptide corresponding to residues 1-40 of the alpha D-globin amino terminus also stimulated adenylyl cyclase activity and promoted differentiation. This 125I-labeled synthetic peptide bound specifically to T. cruzi epimastigote cells. Activation of epimastigote adenylyl cyclase by the hemoglobin-derived peptide may play an important role in T. cruzi differentiation and consequently in the transmission of Chagas disease.


Assuntos
Adenilil Ciclases/metabolismo , Globinas/farmacologia , Fragmentos de Peptídeos/farmacologia , Triatoma/metabolismo , Trypanosoma cruzi/enzimologia , Sequência de Aminoácidos , Animais , Galinhas , Cromatografia Líquida de Alta Pressão , Fenômenos Fisiológicos do Sistema Digestório , Eletroforese em Gel de Poliacrilamida , Globinas/química , Guanosina Trifosfato/farmacologia , Guanilil Imidodifosfato/farmacologia , Hemoglobinas/farmacologia , Humanos , Camundongos , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/isolamento & purificação , Peptídeos/síntese química , Peptídeos/farmacologia , Homologia de Sequência de Aminoácidos
13.
Biochem Mol Biol Int ; 30(3): 547-58, 1993 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-8401312

RESUMO

Growth and polyamine content of epimastigotes of Trypanosoma cruzi were studied in the presence of precursors or inhibitors of putrescine synthesis. Arginine and agmatine turned out to be better precursors than ornithine. alpha-D-difluoromethylarginine, an inhibitor of arginine decarboxylase, inhibited cellular proliferation and decreased putrescine, spermidine and spermine contents while that of cadaverine remained unchanged. These effects were reversed both by agmatine and putrescine. alpha-D-difluoromethylornithine, an inhibitor of ornithine decarboxylase did not inhibit growth of parasites grown in a polyamine free medium. These results suggest that epimastigotes need polyamines to grow, and that the parasite are able to synthesize them mainly through the arginine decarboxylase pathway.


Assuntos
Poliaminas/metabolismo , Trypanosoma cruzi/metabolismo , Agmatina/metabolismo , Animais , Arginina/análogos & derivados , Arginina/metabolismo , Arginina/farmacologia , Carboxiliases/antagonistas & inibidores , Eflornitina/farmacologia , Ornitina/metabolismo , Inibidores da Ornitina Descarboxilase , Proteínas de Protozoários/antagonistas & inibidores , Putrescina/biossíntese , Trypanosoma cruzi/crescimento & desenvolvimento
14.
Biol Res ; 26(1-2): 279-83, 1993.
Artigo em Inglês | MEDLINE | ID: mdl-7670540

RESUMO

A peptide from hindguts of the Triatoma infestans, the hematophagous Chagas' insect vector, activates adenylyl cyclase activity in Trypanosoma cruzi epimastigote membranes and stimulates the in vitro differentiation of epimastigotes (proliferative and non-infectious forms) to metacyclic trypomastigotes (non-proliferative and infectious forms). The peptide was purified from hindguts of insects fed two days before with chicken blood. After purification, the peptide showed upon SDS-PAGE a single band of about 10 kDa. The sequence for 20 residues of the amino terminus of this peptide was: H2N-Met-Leu-Thr-Ala-Glu-Asp-Lys-Lys-Leu-Ile-Gln-Gln-Ala-Trp-Glu-Lys-Ala- Ala-Ser-His. This sequence corresponds to the amino terminus of chicken alpha D-globin. A synthetic peptide with the sequence of the 40 amino acids corresponding to the amino terminus of alpha D-globin, also stimulated T. cruzi adenylyl cyclase activity and promoted metacyclogenesis.


Assuntos
Adenilil Ciclases/metabolismo , Globinas/fisiologia , Fragmentos de Peptídeos/fisiologia , Triatoma/metabolismo , Trypanosoma cruzi/enzimologia , Sequência de Aminoácidos , Animais , Galinhas , Fenômenos Fisiológicos do Sistema Digestório , Camundongos , Dados de Sequência Molecular , Fragmentos de Peptídeos/isolamento & purificação
15.
Biochem Biophys Res Commun ; 187(1): 108-13, 1992 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-1325784

RESUMO

Metacyclogenesis of Trypanosoma cruzi epimastigotes was evaluated in a medium supplemented with Triatoma infestans intestinal homogenate in the presence of sugars and derivates as are mannose, galactose, fucose, N-acetylglucosamine, mannose 6-P, and fructose 1,6-P at a concentration of 25 mM. Only mannose significantly inhibited metacyclogenesis. Sodium metaperiodate and trypsin treatment of the intestinal homogenate also inhibited differentiation. In our opinion there exists a proteinic factor in the intestine of the vector that promotes metacyclogenesis and is incorporated by the parasite. Treatment of the intestinal homogenate with alkaline phosphatase had no effect. Instead, high ionic strength in the medium (0.4 M NaCl) strongly inhibited metacyclogenesis indicating that, in these conditions, the possible binding of the differentiation factor to the parasite surface was inhibited.


Assuntos
Manose/farmacologia , Trypanosoma cruzi/crescimento & desenvolvimento , Animais , Carboidratos/farmacologia , Morfogênese/efeitos dos fármacos , Concentração Osmolar , Ácido Periódico/farmacologia , Cloreto de Sódio/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Tripsina/farmacologia
16.
Rev Argent Microbiol ; 24(2): 81-5, 1992.
Artigo em Inglês | MEDLINE | ID: mdl-1298017

RESUMO

During the morphogenic process from epimastigote to metacyclic trypomastigote stage changes at molecular levels are produced. In the present work the incorporation of macromolecule precursors L [3H]-leucine, [3H]-thymidine and [3H]-uridine was used to monitor the differences between parasites growing with and without specific stimulation for morphogenesis (SEpi and NEpi, respectively). The peak of maximum [3H]-uridine incorporation was earlier in the SEpi than in NEpi (3 vs 4 days, respectively) reaching 129% higher values. Even at day 2, SEpi already showed values of [3H]-uridine incorporation slightly higher (29%) than the maximum reached by NEpi. One the other hand the peak of L-[3H]-leucine incorporation was 48 h delayed comparing SEpi vs NEpi with quantitatively similar patterns, while no differences were registered for [3H]-thymidine incorporation. Therefore the [3H]-uridine incorporation may constitute an earlier marker of morphogenesis since the process was optically detected starting day at 6.


Assuntos
Leucina/metabolismo , Timidina/metabolismo , Trypanosoma cruzi/metabolismo , Uridina/metabolismo , Animais , Intestinos/química , Morfogênese/efeitos dos fármacos , Extratos de Tecidos/farmacologia , Triatoma/química , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/crescimento & desenvolvimento
17.
Rev. argent. microbiol ; 24(2): 81-5, 1992 Apr-Jun.
Artigo em Espanhol | LILACS-Express | LILACS, BINACIS | ID: biblio-1171563

RESUMO

During the morphogenic process from epimastigote to metacyclic trypomastigote stage changes at molecular levels are produced. In the present work the incorporation of macromolecule precursors L [3H]-leucine, [3H]-thymidine and [3H]-uridine was used to monitor the differences between parasites growing with and without specific stimulation for morphogenesis (SEpi and NEpi, respectively). The peak of maximum [3H]-uridine incorporation was earlier in the SEpi than in NEpi (3 vs 4 days, respectively) reaching 129


higher values. Even at day 2, SEpi already showed values of [3H]-uridine incorporation slightly higher (29


) than the maximum reached by NEpi. One the other hand the peak of L-[3H]-leucine incorporation was 48 h delayed comparing SEpi vs NEpi with quantitatively similar patterns, while no differences were registered for [3H]-thymidine incorporation. Therefore the [3H]-uridine incorporation may constitute an earlier marker of morphogenesis since the process was optically detected starting day at 6.

18.
Rev. argent. microbiol ; 24(2): 81-5, 1992 Apr-Jun.
Artigo em Espanhol | LILACS-Express | LILACS, BINACIS | ID: biblio-1171571

RESUMO

During the morphogenic process from epimastigote to metacyclic trypomastigote stage changes at molecular levels are produced. In the present work the incorporation of macromolecule precursors L [3H]-leucine, [3H]-thymidine and [3H]-uridine was used to monitor the differences between parasites growing with and without specific stimulation for morphogenesis (SEpi and NEpi, respectively). The peak of maximum [3H]-uridine incorporation was earlier in the SEpi than in NEpi (3 vs 4 days, respectively) reaching 129


higher values. Even at day 2, SEpi already showed values of [3H]-uridine incorporation slightly higher (29


) than the maximum reached by NEpi. One the other hand the peak of L-[3H]-leucine incorporation was 48 h delayed comparing SEpi vs NEpi with quantitatively similar patterns, while no differences were registered for [3H]-thymidine incorporation. Therefore the [3H]-uridine incorporation may constitute an earlier marker of morphogenesis since the process was optically detected starting day at 6.

19.
Rev. argent. microbiol ; 21(1): 9-14, ene.-mar. 1989. tab
Artigo em Inglês | LILACS | ID: lil-78197

RESUMO

En el presente trabajo se estudió el comportamieto de varias cepas y clones de Epi de T. cruzi en medios estimulantes de la diferenciación: M 16, TAUP, TAUS, LIT-hemina y G-IH. Los resultados mostraron que en nuestras condiciones experimentales el medio G-IH fue el único adecuado para producir morfogénesis Epi-Mtc. La temperatura óptima del proceso tanto de multiplicación como de diferenciación en ese medio fue 28-C. el empleo G-IH empobrecido en nutrientes por dilución no indujo diferenciación. Los parásitos que se obtuvieron de medio bifásico entre 24 a 48 h de cultivados fueron los óptimos para obtener morfogénesis. El bloqueo de la oferta de Ca++ externo afectó la capacidad de diferenciarse de las cepas Tul y RA en grado variable


Assuntos
Animais , Trypanosoma cruzi/crescimento & desenvolvimento , Meios de Cultura , Morfogênese
20.
Rev Argent Microbiol ; 21(1): 9-14, 1989.
Artigo em Inglês | MEDLINE | ID: mdl-2664877

RESUMO

This work is a study on the behaviour of Epi from various T. cruzi strains and clones grown in media stimulating differentiation such as M 16, TAUP, TAUS, LIT-hemin and G-IH. Results showed that in our experimental conditions the G-IH medium was the only suitable one to induce Epi-Mtc morphogenesis. Optimal temperature for both multiplication and differentiation in this medium was 28 degrees C. The use of G-IH medium after nutrient depletion by dilution failed to induce differentiation. Parasites obtained from biphasic medium at 24-48 h culture proved the best to achieve morphogenesis. External Ca++ supply blockage affected the differentiation capacity of Tul and RA strains to a variable degree.


Assuntos
Trypanosoma cruzi/crescimento & desenvolvimento , Animais , Meios de Cultura , Morfogênese , Temperatura
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