RESUMO
The palladacycle complex DPPE 1.2 was previously shown to inhibit Leishmania (Leishmania) amazonensis infection in vitro and in vivo. The present study aimed to evaluate the effect of DPPE 1.2 associated with a recombinant cysteine proteinase, rLdccys1, and the adjuvant Propionibacterium acnes on L. (L.) amazonensis infection in two mouse strains, BALB/c, and C57BL/6. Treatment with this association potentiated the leishmanicidal effect of DPPE 1.2 resulting in a reduction of parasite load in both strains of mice which was higher compared to that found in groups treated with either DPPE 1.2 alone or associated with P. acnes or rLdccys1. The reduction of parasite load in both mice strains was followed by immunomodulation mediated by an increase of memory CD4+ and CD8+ T lymphocytes, IFN-γ levels and reduction of active TGF-ß in treated animals. No infection relapse was observed 1 month after the end of treatment in mice which received DPPE 1.2 associated with rLdccys1 or rLdccys1 plus P. acnes in comparison to that exhibited by animals treated with DPPE 1.2 alone. Evaluation of serum levels of AST, ALT, urea, and creatinine showed no alterations among treated groups, indicating that this treatment schedule did not induce hepato or nephrotoxicity. These data indicate the potential use of this association as a therapeutic alternative for cutaneous leishmaniasis caused by L. (L) amazonensis.
Assuntos
Antiprotozoários/uso terapêutico , Cisteína Endopeptidases/uso terapêutico , Imunoterapia/métodos , Leishmaniose Cutânea/tratamento farmacológico , Propionibacterium acnes , Proteínas de Protozoários/uso terapêutico , Animais , Antiprotozoários/administração & dosagem , Antiprotozoários/toxicidade , Terapia Combinada , Cisteína Endopeptidases/administração & dosagem , Cisteína Endopeptidases/imunologia , Cisteína Endopeptidases/toxicidade , Avaliação Pré-Clínica de Medicamentos , Feminino , Memória Imunológica , Interferon gama/metabolismo , Leishmania mexicana , Leishmaniose Cutânea/imunologia , Linfonodos/imunologia , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas de Protozoários/administração & dosagem , Proteínas de Protozoários/imunologia , Proteínas de Protozoários/toxicidade , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/uso terapêutico , Proteínas Recombinantes/toxicidade , Subpopulações de Linfócitos T/imunologia , Fator de Crescimento Transformador beta/metabolismoRESUMO
Therapeutic vaccine research and development are especially important in Chagas disease considering the characteristics of the chronic infection and the number of people in the Americas living with a parasite infection for decades. We have previously reported the efficacy of attenuated Salmonella enterica (S) carrying plasmid encoding cruzipain (SCz) to protect against Trypanosoma cruzi infection. In the present work we investigated whether Cz DNA vaccine immunotherapy could be effective in controlling an ongoing T. cruzi infection in mice. We here report the intramuscular administration of naked Cz DNA or the oral administration of Salmonella as Cz DNA delivery system as therapeutic vaccines in mice during acute or chronic infection. The coadministration of a plasmid encoding GM-CSF improved vaccine performance, indicating that the stimulation of innate immune cells is needed in the event of an ongoing infection. These therapeutic vaccines were able to address the response to a protective and sustained Th1 biased profile not only against Cz but also against a variety of parasite antigens. The combined therapeutic vaccine during the chronic phase of infection prevents tissue pathology as shown by a reduced level of enzyme activity characteristic of tissue damage and a tissue status compatible with normal tissue. The obtained results suggest that immunotherapy with Cz and GM-CSF DNAs, either alone or in combination with other drug treatments, may represent a promising alternative for Chagas disease therapy.
Assuntos
Anticorpos Antiprotozoários/sangue , Doença de Chagas/tratamento farmacológico , Cisteína Endopeptidases/uso terapêutico , DNA/uso terapêutico , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Imunoterapia/métodos , Vacinas Protozoárias/imunologia , Trypanosoma cruzi/imunologia , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Doença de Chagas/parasitologia , Combinação de Medicamentos , Feminino , Imunidade Inata/genética , Imunidade Inata/imunologia , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Camundongos , Camundongos Endogâmicos C3H , Parasitemia/tratamento farmacológico , Parasitemia/prevenção & controle , Plasmídeos/genética , Plasmídeos/uso terapêutico , Proteínas de Protozoários , Salmonella/genética , Células Th1/imunologia , Trypanosoma cruzi/efeitos dos fármacos , Vacinas de DNA/genética , Vacinas de DNA/imunologiaRESUMO
Previous studies demonstrated that proteinases from latex of C. candamarcensis act as mitogens on fibroblast and epithelial cells and a subsequent report showed their protective, angiogenic and wound healing effects on gastric ulcers. In this study, we present evidence of skin healing activity by the group of proteinases known as P1G10. By using a hairless mouse model, we compared the healing effect following topical application of various concentrations of P1G10. The data confirm that healing actions take place between 0.1 and 1%, without adverse local irritation or systemic toxicological action after a prolonged period of use. The wound healing effect is unaltered when P1G10 is previously inhibited with iodoacetamide. The low permeation of the hydrosoluble formulation Polawax(®) supports the maintenance of the drug at the site of application. These results extend the healing properties of these groups of enzymes in situations of dermatological trauma and open the way to future clinical applications.
Assuntos
Cisteína Endopeptidases/farmacologia , Glicoproteínas/farmacologia , Látex/química , Fitoterapia , Pele/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Cicatrização/fisiologia , Animais , Peso Corporal/efeitos dos fármacos , Carica/enzimologia , Cisteína Endopeptidases/uso terapêutico , Dextranos , Eletroforese em Gel de Poliacrilamida , Feminino , Géis , Glicoproteínas/uso terapêutico , Masculino , Camundongos , Camundongos PeladosRESUMO
Cysteine proteinases are involved in virtually every aspect of plant physiology and development. They play a role in development, senescence, programmed cell death, storage and mobilization of germinal proteins, and in response to various types of environmental stress. In this review, we focus on a group of plant defensive enzymes occurring in germinal tissue of Caricaceae. These enzymes elicit a protective response in the unripe fruit after physical stress. We propose that these enzymes follow a strategy similar to mammalian serine proteinases involved in blood clotting and wound healing. We show evidence for the pharmacological role of plant cysteine proteinases in mammalian wound healing, immunomodulation, digestive conditions, and neoplastic alterations.
Assuntos
Anti-Helmínticos/uso terapêutico , Caricaceae/enzimologia , Cisteína Endopeptidases/metabolismo , Cisteína Endopeptidases/uso terapêutico , Proteínas de Plantas/metabolismo , Animais , Humanos , Modelos Moleculares , Papaína/química , Papaína/metabolismo , Pisum sativum/enzimologia , Proteínas de Plantas/uso terapêutico , Raízes de Plantas/enzimologia , Conformação Proteica , Serina Endopeptidases/metabolismo , Serina Endopeptidases/uso terapêutico , Cicatrização/efeitos dos fármacosRESUMO
Latex from Caricaceae contains proteolytic enzymes localized in the fruit, which are used ethnopharmacologically to treat digestive disorders. Some of these proteins display proliferative properties when probed with mammalian cells, suggesting a role in the reconstruction of wounded tissue. We tested the efficacy of a proteolytic fraction derived from Carica candamarcensis, designated as P1G10 in experimental rodent models, to protect and heal chemically induced gastric ulcers. The protective effect of oral administration of P1G10 fraction was analyzed in indomethacin-treated Wistar animals. The healing effect of P1G10 was studied following sub-serous injection of acetic acid in a Wistar rat model. The results show that P1G10 between 0.1 and 10 mg/kg protect indomethacin but not ethanol-induced gastric ulcers. The maximal protection attained was 67% with 10 mg/kg of P1G10. The healing rate by 10 mg/kg of P1G10 using the acetic acid ulcerogenic model is similar to that of omeprazole (10 mg/kg) or ranitidine (100 mg/kg). The effect of P1G10 at 10 mg/kg seems to be mediated by an increase in the mucus content by 25% and stimulation of angiogenesis by 64% in a manner similar to growth factors. These results confirm the protective and healing role of proteinases from C. candamarcensis.
Assuntos
Carica/enzimologia , Cisteína Endopeptidases/uso terapêutico , Látex/química , Fitoterapia , Úlcera Gástrica/tratamento farmacológico , Animais , Antiulcerosos , Carica/química , Cisteína Endopeptidases/isolamento & purificação , Cisteína Endopeptidases/farmacologia , Frutas/química , Frutas/enzimologia , Neovascularização Fisiológica/efeitos dos fármacos , Ratos , Ratos Wistar , Úlcera Gástrica/induzido quimicamente , Úlcera Gástrica/prevenção & controleRESUMO
In the present work, the antitumor effect of fastuosain, a cysteine proteinase from Bromelia fastuosa, was investigated. In the intravenous model of lung colonization in C57Bl/6 mice, fastuosain and bromelain injected intraperitoneally were protective, and very few nodules of B16F10-Nex2 melanoma cells were detected. Tumor cells treated with fastuosain showed reduced expression of CD44 and decreased invasion through Matrigel, lost their cytoplasmic extensions and substrate adherence, and became round and detached, forming strongly bound cell clusters in suspension. Peritoneal cells recruited and activated by fastuosain treatment (mainly monocytic cells and lymphocytes) migrated to the lung, where pulmonary melanoma metastases grew. Adoptive transference of peritoneal cells recruited by fastuosain had no protective effect against lung metastases in recipient mice. Treatment of green fluorescent protein-chimeric animals with fastuosain did not change the number of cells that migrated to the lung, compared to PBS-injected control mice, but the number of positive major histocompatibility complex class II cells increased with fastuosain treatment. Murine antibodies against fastuosain, bromelain, and cathepsins B and L cross-reacted in ELISA and recognized surface and cytoplasmic components expressed on B16F10-Nex2 cells. Anti-fastuosain antibodies were cytotoxic/lytic to B16F10-Nex2 cells. Antitumor effects of fastuosain involve mainly the direct effect of the enzyme and elicitation of protective antibodies.
Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Cisteína Endopeptidases/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/secundário , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/secundário , Transferência Adotiva , Animais , Formação de Anticorpos , Antígenos de Neoplasias/biossíntese , Antígenos de Neoplasias/imunologia , Antineoplásicos Fitogênicos/imunologia , Antineoplásicos Fitogênicos/farmacologia , Bromelaínas/imunologia , Bromelaínas/farmacologia , Bromelaínas/uso terapêutico , Linhagem Celular Tumoral/efeitos dos fármacos , Quimiotaxia de Leucócito/efeitos dos fármacos , Cisteína Endopeptidases/imunologia , Cisteína Endopeptidases/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Neoplasias Pulmonares/imunologia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/transplante , Masculino , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Papaína/imunologia , Papaína/farmacologia , Papaína/uso terapêutico , Quimera por RadiaçãoRESUMO
Endemic in most American countries, Chagas' disease causes high morbidity and mortality. Recent experimental and clinical evidence shows the importance of chemotherapy in both the acute and chronic phases of this disease. However, treatment is yet limited by the toxicity associated to available drugs. This review describes the design, evolution, and selection of dipeptides that interrupt the intracellular cycle of T. cruzi and cure acute experimental infections in laboratory animals. Peptido-mimetic inhibitors specifically bind cruzain, a T. cruzi cystein protease. The inhibitors cause alterations in the Golgi complex and ER, accumulation of unprocessed enzyme within Golgi cisternae, and decrease of mature cruzain within lysosomes. The most effective compound, N-Pip-F-hF-VS phi, cured an acute lethal infection in experimental animals. Myocardial lesions, lymphocyte infiltration and intracellular amastigote clusers were absent in treated animals. Preliminary toxicology and pharmacokinetic analyses suggest the lack of toxicity associated to high doses and prolonged treatment regimes. Protease inhibitors may soon become good chemotherapeutic alternatives for acute and chronic Chagas' disease.