RESUMEN
Human T-lymphotropic virus type 1 (HTLV-1) induces a strong activation of the immune system, especially in individuals with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Physalin F is a secosteroid with potent anti-inflammatory and immunomodulatory activities. The present study aimed to investigate the effects of physalin F on peripheral blood mononuclear cells (PBMC) of HAM/TSP subjects. A concentration-dependent inhibition of spontaneous proliferation of PBMC from HAM/TSP subjects was observed in the presence of physalin F, as evaluated by (3)H-thymidine uptake. The IC50 for physalin F was 0.97 ± 0.11 µM. Flow cytometry analysis using Cytometric Bead Array (CBA) showed that physalin F (10 µM) significantly reduced the levels of IL-2, IL-6, IL-10, TNF-α and IFN-γ, but not IL-17A, in supernatants of PBMC cultures. Next, apoptosis induction was addressed by using flow cytometry to evaluate annexin V expression. Treatment with physalin F (10 µM) increased the apoptotic population of PBMC in HAM/TSP subjects. Transmission electron microscopy analysis of PBMC showed that physalin F induced ultrastructural changes, such as pyknotic nuclei, damaged mitochondria, enhanced autophagic vacuole formation, and the presence of myelin-like figures. In conclusion, physalin F induces apoptosis of PBMC, decreasing the spontaneous proliferation and cytokine production caused by HTLV-1 infection.
Asunto(s)
Inmunosupresores/uso terapéutico , Leucocitos Mononucleares/patología , Paraparesia Espástica Tropical/sangre , Paraparesia Espástica Tropical/tratamiento farmacológico , Physalis/química , Secoesteroides/uso terapéutico , Proliferación Celular/efectos de los fármacos , Citocinas/metabolismo , Humanos , Inmunosupresores/farmacología , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/ultraestructura , Paraparesia Espástica Tropical/patología , Fosfatidilserinas/metabolismo , Secoesteroides/química , Secoesteroides/farmacologíaRESUMEN
We previously observed that physalins have immunomodulatory properties, as well as antileishmanial and antiplasmodial activities. Here, we investigated the anti-Trypanosoma cruzi activity of physalins B, D, F and G. We found that physalins B and F were the most potent compounds against trypomastigote and epimastigote forms of T. cruzi. Electron microscopy of trypomastigotes incubated with physalin B showed disruption of kinetoplast, alterations in Golgi apparatus and endoplasmic reticulum, followed by the formation of myelin-like figures, which were stained with MDC to confirm their autophagic vacuole identity. Physalin B-mediated alteration in Golgi apparatus was likely due to T. cruzi protease perturbation; however physalins did not inhibit activity of the trypanosomal protease cruzain. Flow cytometry examination showed that cell death is mainly caused by necrosis. Treatment with physalins reduced the invasion process, as well as intracellular parasite development in macrophage cell culture, with a potency similar to benznidazole. We observed that a combination of physalins and benznidazole has a greater anti-T. cruzi activity than when compounds were used alone. These results indicate that physalins, specifically B and F, are potent and selective trypanocidal agents. They cause structural alterations and induce autophagy, which ultimately lead to parasite cell death by a necrotic process.
Asunto(s)
Physalis/química , Secoesteroides/farmacología , Tripanocidas/farmacología , Trypanosoma cruzi/efectos de los fármacos , Animales , Femenino , Ratones , Ratones Endogámicos BALB C , Estructura Molecular , Secoesteroides/química , Tripanocidas/química , Trypanosoma cruzi/fisiología , Trypanosoma cruzi/ultraestructuraRESUMEN
Physalin B is a natural secosteroidal, extracted from the Solanaceae plant, Physalis angulata, and it presents immune-modulator effects on the bloodsucking bug, Rhodnius prolixus. In this work, R. prolixus was treated with physalin B at a concentration of 1 mg/ml of blood meal (oral application), or 20 ng/insect (applied topically) or 57 ng/cm(2) of filter paper (contact treatment), and infected with Trypanosoma cruzi Dm28c clone (2×10(6) epimastigotes/insect). The three types of applications significantly decreased the number of T. cruzi Dm28c in the gut comparing with the non-treated infected insects (controls). All groups of infected insects treated with physalin B had higher numbers of bacterial microbiota in the gut than the non-treated controls infected with T. cruzi. We observed that the infected physalin B insects with topical and contact treatments had a lower antibacterial activity in the gut when compared with control infected insects. Furthermore, infected insects with the physalin B oral treatment produced higher levels of nitrite and nitrate in the gut than control infected insects. These results demonstrate that physalin B decreases the T. cruzi transmission by inhibiting the parasite development in the insect vector R. prolixus. Herein the importance of physalin B modulation on the immune system and microbiota population in terms of parasite development and transmission are discussed.
Asunto(s)
Interacciones Huésped-Parásitos/efectos de los fármacos , Rhodnius/efectos de los fármacos , Secoesteroides/farmacología , Trypanosoma cruzi/efectos de los fármacos , Animales , Metagenoma/efectos de los fármacos , Muda/efectos de los fármacos , Nitratos/metabolismo , Nitritos/metabolismo , Especies de Nitrógeno Reactivo/metabolismo , Rhodnius/inmunología , Rhodnius/metabolismo , Rhodnius/microbiología , Rhodnius/parasitologíaRESUMEN
The antimalarial activities of physalins B, D, F, and G (1-4), isolated from Physalis angulata, were investigated. In silico analysis using the similarity ensemble approach (SEA) database predicted the antimalarial activity of each of these compounds, which were shown using an in vitro assay against Plasmodium falciparum. However, treatment of P. berghei-infected mice with 3 increased parasitemia levels and mortality, whereas treatment with 2 was protective, causing a parasitemia reduction and a delay in mortality in P. berghei-infected mice. The exacerbation of in vivo infection by treatment with 3 is probably due to its potent immunosuppressive activity, which is not evident for 2.
Asunto(s)
Antimaláricos/farmacología , Inmunosupresores/farmacología , Physalis/química , Plasmodium berghei/efectos de los fármacos , Plasmodium falciparum/efectos de los fármacos , Secoesteroides/farmacología , Animales , Antimaláricos/química , Antimaláricos/aislamiento & purificación , Inmunosupresores/química , Inmunosupresores/aislamiento & purificación , Malaria/tratamiento farmacológico , Ratones , Ratones Endogámicos BALB C , Parasitemia/tratamiento farmacológico , Secoesteroides/química , Secoesteroides/aislamiento & purificaciónRESUMEN
Physalis angulata L., Solanaceae, is an annual herb commonly used in popular medicine in many tropical and subtropical countries. P. angulata extracts contain a variety of substances, but little is known about their pharmacological activities. In this work we investigated the in vitro antileishmanial activity of seco-steroids (physalins) purified from P. angulata. Addition of physalins B, F, and G caused a concentration-dependent inhibition in the growth of L. amazonensis promastigotes, being the IC50 values were 6.8, 1.4, and 9.2 μM, respectively. Physalin D was less active and had an IC50 value of 30.5 μM. Physalins were also active in cultures of other Leishmania species (L. major, L. braziliensis, and L. chagasi). Our results demonstrate the potent antileishmanial activity of physalins in cultures of Leishmania species of the New and Old Worlds and suggest the therapeutic potential of these seco-steroids in leishmaniasis.
Physalis angulata L., Solanaceae, é uma erva anual utilizada na medicina popular em muitos países tropicais e subtropicais. Apesar dos extratos da P. angulata apresentarem uma grande variedade de substâncias, pouco é conhecido sobre a sua atividade farmacológica. Neste trabalho foi investigado a atividade antileishmania in vitro de seco-esteroides (fisalinas) purificados da P. angulata. O tratamento com as fisalinas B, F e G causou uma inibição concentração-dependente do crescimento de promastigotas de Leishmania amazonensis em cultura axênica, com valores de IC50 de 6,8, 1,4, e 9,2 μM respectivamente. A fisalina D foi menos ativa, com valores de IC50 de 30,5 μM. Foi também observada uma atividade leishmanicida em culturas de outras espécies de Leishmania (L. major, L. braziliensis e L. chagasi). Nossos resultados demonstram que as fisalinas inibem o crescimento dos promastigotas com o tratamento de espécies de Leishmania do Velho e do Novo Mundos e sugerem o potencial terapêutico destas moléculas na leishmaniose.
RESUMEN
OBJECTIVES: We have previously demonstrated the immunomodulatory effects of physalins, secosteroids purified from Physalis angulata. Here we investigate the antileishmanial activity of physalins in vitro and in vivo in a model of cutaneous leishmaniasis. METHODS: The antileishmanial activity of physalins B, D and F was tested in Leishmania-infected macrophage cultures. For the in vivo studies, BALB/c mice were infected with Leishmania amazonensis subcutaneously in the ear pinna and treated with physalin F by topical administration. RESULTS: Physalins B and F were able to reduce the percentage of Leishmania-infected macrophages and the intracellular parasite number in vitro at concentrations non-cytotoxic to macrophages. More importantly, topical treatment with physalin F significantly reduced the lesion size, the parasite load and histopathological alterations in BALB/c mice infected with L. amazonensis. CONCLUSIONS: Our results demonstrate the potent antileishmanial activity of physalins, especially physalin F, and suggest these molecules as the basis for the development of new therapeutic options for cutaneous leishmaniasis.
Asunto(s)
Antiprotozoarios/farmacología , Antiprotozoarios/uso terapéutico , Leishmania mexicana/efectos de los fármacos , Leishmaniasis Cutánea/tratamiento farmacológico , Physalis/química , Secoesteroides/farmacología , Secoesteroides/uso terapéutico , Animales , Antiprotozoarios/aislamiento & purificación , Femenino , Humanos , Macrófagos/parasitología , Ratones , Ratones Endogámicos BALB C , Secoesteroides/aislamiento & purificación , Piel/parasitología , Piel/patologíaRESUMEN
Physalins are seco-steroids obtained from plants of the family Solanaceae. Herein, we tested Physalis angulata L purified physalin B as an immunomodulatory compound in 5th-instar larvae of Rhodnius prolixus, which were systemically infected with the H14 Trypanosoma rangeli strain protozoan. In uninfected insects, the effective concentration of physalin B, which inhibited 50% of the blood ingested (ED(50)) volume, was 15.2+/-1.6 microg/ml of the meal. Ecdysis processes and mortality in uninfected larvae, treated orally with physalin B in concentrations ranging from 1 to 10 microg/ml, was similar to that observed in insects not treated with physalin B. However, R. prolixus larvae previously fed on blood containing 1.0, 0.1, and 0.01 microg of physalin B/ml exhibited mortality rates of 78.1, 54.3, and 12.7%, respectively, 6 days after inoculation of T. rangeli (1 x 10(3) parasites/insect), whereas only 7.2% mortality was observed in the control group, injected with sterile culture medium. The insects treated with physalin B (0.1 microg/ml) and inoculated with T. rangeli did not modify the phenoloxidase (PO) activity and total hemocyte count in the hemolymph. However, physalin B treatment caused a reduction in hemocyte micro-aggregation and nitric oxide production and enhanced the parasitemia in the hemolymph. These results demonstrate that physalin B from P. angulata is a potent immunomodulatory substance for the bloodsucking insect, R. prolixus.
Asunto(s)
Lactonas/farmacología , Rhodnius/inmunología , Esteroides/farmacología , Trypanosoma/inmunología , Animales , Catecol Oxidasa/metabolismo , Agregación Celular/efectos de los fármacos , Recuento de Células , Activación Enzimática/efectos de los fármacos , Precursores Enzimáticos/metabolismo , Hemocitos/citología , Hemocitos/efectos de los fármacos , Lactonas/química , Larva/efectos de los fármacos , Larva/inmunología , Larva/parasitología , Nitratos/metabolismo , Nitritos/metabolismo , Rhodnius/efectos de los fármacos , Rhodnius/parasitología , Secoesteroides , Esteroides/químicaRESUMEN
Reperfusion of an ischaemic tissue is associated with an intense inflammatory response and inflammation-mediated tissue injury. Physalins, a group of substances with secosteroidal chemical structure, are found in Physalis angulata stems and leaves. Here, we assessed the effects of physalins on the local, remote and systemic injuries following intestinal ischaemia and reperfusion (I/R) in mice and compared with the effects of dexamethasone. Following I/R injury, dexamethasone (10 mg kg(-1)) or physalin B or F markedly prevented neutrophil influx, the increase in vascular permeability in the intestine and the lungs. Maximal inhibition occurred at 20 mg kg(-1). Moreover, there was prevention of haemorrhage in the intestine of reperfused animals. Dexamethasone or physalins effectively suppressed the increase in tissue (intestine and lungs) and serum concentrations of TNF-alpha. Interestingly, treatment with the compounds was associated with enhancement of IL-10. The anti-inflammatory effects of dexamethasone or physalins were reversed by pretreatment with the corticoid receptor antagonist RU486 (25 mg kg(-1)). The drug compounds suppressed steady-state concentrations of corticosterone, but did not alter the reperfusion-associated increase in levels of corticosterone. The IL-10-enhancing effects of the drugs were not altered by RU486. In conclusion, the in vivo anti-inflammatory actions of physalins, natural steroidal compounds, appear to be mostly due to the activation of glucocorticoid receptors. Compounds derived from these natural secosteroids may represent novel therapeutic options for the treatment of inflammatory diseases.
Asunto(s)
Antiinflamatorios no Esteroideos/uso terapéutico , Enfermedades Intestinales/prevención & control , Lactonas/uso terapéutico , Daño por Reperfusión/prevención & control , Esteroides/uso terapéutico , Animales , Antiinflamatorios/farmacología , Permeabilidad Capilar/efectos de los fármacos , Quimiocinas/biosíntesis , Citocinas/biosíntesis , Dexametasona/farmacología , Relación Dosis-Respuesta a Droga , Hemorragia Gastrointestinal/prevención & control , Glucocorticoides/farmacología , Hemoglobinas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Peroxidasa/metabolismo , SecoesteroidesRESUMEN
BACKGROUND AND PURPOSE: Prior reports have described increased signal intensity (SI) of CSF on fluid-attenuated inversion recovery (FLAIR) images of anesthetized patients receiving 100% O(2). This appearance can simulate that of diseases. We evaluated the relationship between the concentration of inhaled O(2) and the development of increased SI of CSF on FLAIR images. METHODS: FLAIR was performed in 25 healthy volunteers breathing room air and 100% O(2) through a face mask for 5, 10, and 15 minutes. MR imaging, including FLAIR imaging, was performed in 52 patients with no potential meningeal abnormalities under general anesthesia: 21 received an equal mixture of N(2)O and O(2), and 31 received 100% O(2). The SI of CSF in volunteers and patients was graded in several locations by using a three-point scale. RESULTS: SI of CSF significantly increased (P <.05) in various locations, in both volunteers and patients breathing 100% O(2), when compared with SI in the same volunteers breathing room air. Hyperintensity of CSF was not significantly different in volunteers receiving 100% O(2) through a face mask compared with anesthetized patients receiving 100% O(2) through a laryngeal airway or an endotracheal tube. No significant increase in SI occurred in patients receiving 50% O(2), when compared with the SI of volunteers breathing room air. CONCLUSION: Supplemental oxygen at 100% is a main cause of artifactual CSF hyperintensity on FLAIR images, regardless of the anesthetic drug used. This artifact does not develop when 50% O(2) is administered.
Asunto(s)
Encéfalo/anatomía & histología , Líquido Cefalorraquídeo , Imagen por Resonancia Magnética , Terapia por Inhalación de Oxígeno , Adolescente , Adulto , Anciano , Anestesia General , Anestésicos , Artefactos , Ángulo Pontocerebeloso/anatomía & histología , Niño , Preescolar , Femenino , Humanos , Lactante , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Espacio Subaracnoideo/anatomía & histologíaRESUMEN
Physalis angulata L. is an annual herb widely used in popular medicine for the treatment of a variety of pathologies. Here, we tested immunomodulatory activities of physalins, seco-steroids purified from P. angulata extracts. Addition of physalins B, F or G, but not D, caused a reduction in nitric oxide production by macrophages stimulated with lipopolysaccaride and interferon-gamma. In the presence of physalin B, macrophages stimulated with lipopolysaccaride, alone or in combination with interferon-gamma, produced lower levels of tumour necrosis factor (TNF)-alpha, interleukin-6 and interleukin-12. The inhibitory activity of physalin B, unlike that of dexamethasone, was not reversed by RU486 [(4-dimethylamino) phenyl-17beta-hydroxy-17-(1-propynyl)estra-4,9-dien-3-one], an antiglucocorticoid. Physalin B-treated mice had lower levels of serum TNF-alpha than control mice after lipopolysaccaride challenge. More importantly, mice injected with physalins B, F or G survived after a lethal lipopolysaccaride challenge. These results demonstrate that seco-steroids from P. angulata are potent immunomodulatory substances and act through a mechanism distinct from that of dexamethasone.