RESUMEN
BACKGROUND: The challenge in anti-malarial chemotherapy is based on the emergence of resistance to drugs and the search for medicines against all stages of the life cycle of Plasmodium spp. as a therapeutic target. Nowadays, many molecules with anti-malarial activity are reported. However, few studies about the cellular and molecular mechanisms to understand their mode of action have been explored. Recently, new primaquine-based hybrids as new molecules with potential multi-acting anti-malarial activity were reported and two hybrids of primaquine linked to quinoxaline 1,4-di-N-oxide (PQ-QdNO) were identified as the most active against erythrocytic, exoerythrocytic and sporogonic stages. METHODS: To further understand the anti-malarial mode of action (MA) of these hybrids, hepg2-CD81 were infected with Plasmodium yoelii 17XNL and treated with PQ-QdNO hybrids during 48 h. After were evaluated the production of ROS, the mitochondrial depolarization, the total glutathione content, the DNA damage and proteins related to oxidative stress and death cell. RESULTS: In a preliminary analysis as tissue schizonticidals, these hybrids showed a mode of action dependent on peroxides production, but independent of the activation of transcription factor p53, mitochondrial depolarization and arrest cell cycle. CONCLUSIONS: Primaquine-quinoxaline 1,4-di-N-oxide hybrids exert their antiplasmodial activity in the exoerythrocytic phase by generating high levels of oxidative stress which promotes the increase of total glutathione levels, through oxidation stress sensor protein DJ-1. In addition, the role of HIF1a in the mode of action of quinoxaline 1,4-di-N-oxide is independent of biological activity.
Asunto(s)
Antimaláricos/farmacología , Plasmodium yoelii/efectos de los fármacos , Primaquina/farmacología , Quinoxalinas/farmacología , Combinación de Medicamentos , Eritrocitos/parasitología , Células Hep G2 , Humanos , Esporozoítos/efectos de los fármacosRESUMEN
Compound 1-methyl-7-nitro-4-(5-(piperidin-1-yl)pentyl)-3,4-dihydroquinoxalin-2(1H)-one (VAM2-6) was evaluated against a blood-induced infection with chloroquine-sensitive Plasmodium yoelii yoelii lethal strain in CD1 mice in a 4-day test scheme. LD50 of the compound was 56.51 mg/kg and LD10 was 20.58 mg/kg (taken as the highest dose). Animals were treated by oral gavage of 20, 10, and 5 mg/kg. Mice in the untreated control group showed a progressively increasing parasitemia leading to mouse death on 6 days post-infection; in this group, all mice showed parasites in the blood on the fifth day of sampling; the mean parasitemia on that day was 19.4%. A 4-day dosage of 20 mg/kg of VAM2-6 showed a 97% chemosuppression of total parasitemia on the fifth day, a 28 days survival time, and 20% of cured animals. A 4-day dosage of 10 and 5 mg/kg showed 85 and 37%, respectively, chemosuppression of total parasitemia on the fifth day; but all mice died from days 6 to 9 post-infection with increasing parasitemia. Mice treated with chloroquine at 5 mg/kg survived during the experiment. The results obtained in this study showed that the infection outcome of P. yoelii yoelii-infected mice is affected by VAM2-6 compound by slowing down the parasite replication, retarding the patency time, and increasing their survival time. Although compound VAM2-6 was active at higher doses than chloroquine, these results leaves a door open to the study of its structure in order to improve its antimalarial activity.
Asunto(s)
Antimaláricos/uso terapéutico , Malaria/tratamiento farmacológico , Plasmodium yoelii/efectos de los fármacos , Quinoxalinas/uso terapéutico , Administración Oral , Animales , Bioensayo , Modelos Animales de Enfermedad , Masculino , Ratones , Análisis de Supervivencia , Resultado del TratamientoRESUMEN
Malaria is still a major health problem in developing countries. It is caused by the protist parasite Plasmodium, in which proteases are activated during the cell cycle. Ca(2+) is a ubiquitous signalling ion that appears to regulate protease activity through changes in its intracellular concentration. Proteases are crucial to Plasmodium development, but the role of Ca(2+) in their activity is not fully understood. Here we investigated the role of Ca(2+) in protease modulation among rodent Plasmodium spp. Using fluorescence resonance energy transfer (FRET) peptides, we verified protease activity elicited by Ca(2+) from the endoplasmatic reticulum (ER) after stimulation with thapsigargin (a sarco/endoplasmatic reticulum Ca(2+)-ATPase (SERCA) inhibitor) and from acidic compartments by stimulation with nigericin (a K(+)/H(+) exchanger) or monensin (a Na(+)/H(+) exchanger). Intracellular (BAPTA/AM) and extracellular (EGTA) Ca(2+) chelators were used to investigate the role played by Ca(2+) in protease activation. In Plasmodium berghei both EGTA and BAPTA blocked protease activation, whilst in Plasmodium yoelii these compounds caused protease activation. The effects of protease inhibitors on thapsigargin-induced proteolysis also differed between the species. Pepstatin A and phenylmethylsulphonyl fluoride (PMSF) increased thapsigargin-induced proteolysis in P. berghei but decreased it in P. yoelii. Conversely, E64 reduced proteolysis in P. berghei but stimulated it in P. yoelii. The data point out key differences in proteolytic responses to Ca(2+) between species of Plasmodium.
Asunto(s)
Activación Enzimática/efectos de los fármacos , Eritrocitos/parasitología , Péptido Hidrolasas/metabolismo , Péptidos/farmacología , Plasmodium berghei/enzimología , Plasmodium yoelii/enzimología , Animales , Calcio/metabolismo , Calcio/farmacología , Señalización del Calcio , Transferencia Resonante de Energía de Fluorescencia , Monensina/farmacología , Nigericina/farmacología , Péptidos/química , Plasmodium berghei/efectos de los fármacos , Plasmodium yoelii/efectos de los fármacos , Tapsigargina/farmacologíaRESUMEN
Neurological impairments are frequently detected in children surviving cerebral malaria (CM), the most severe neurological complication of infection with Plasmodium falciparum. The pathophysiology and therapy of long lasting cognitive deficits in malaria patients after treatment of the parasitic disease is a critical area of investigation. In the present study we used several models of experimental malaria with differential features to investigate persistent cognitive damage after rescue treatment. Infection of C57BL/6 and Swiss (SW) mice with Plasmodium berghei ANKA (PbA) or a lethal strain of Plasmodium yoelii XL (PyXL), respectively, resulted in documented CM and sustained persistent cognitive damage detected by a battery of behavioral tests after cure of the acute parasitic disease with chloroquine therapy. Strikingly, cognitive impairment was still present 30 days after the initial infection. In contrast, BALB/c mice infected with PbA, C57BL6 infected with Plasmodium chabaudi chabaudi and SW infected with non lethal Plasmodium yoelii NXL (PyNXL) did not develop signs of CM, were cured of the acute parasitic infection by chloroquine, and showed no persistent cognitive impairment. Reactive oxygen species have been reported to mediate neurological injury in CM. Increased production of malondialdehyde (MDA) and conjugated dienes was detected in the brains of PbA-infected C57BL/6 mice with CM, indicating high oxidative stress. Treatment of PbA-infected C57BL/6 mice with additive antioxidants together with chloroquine at the first signs of CM prevented the development of persistent cognitive damage. These studies provide new insights into the natural history of cognitive dysfunction after rescue therapy for CM that may have clinical relevance, and may also be relevant to cerebral sequelae of sepsis and other disorders.
Asunto(s)
Trastornos del Conocimiento/patología , Trastornos del Conocimiento/prevención & control , Malaria Cerebral/complicaciones , Malaria Cerebral/parasitología , Plasmodium berghei/efectos de los fármacos , Plasmodium yoelii/efectos de los fármacos , Animales , Antimaláricos/uso terapéutico , Antioxidantes/uso terapéutico , Conducta Animal , Combinación de Medicamentos , Eritrocitos/efectos de los fármacos , Eritrocitos/parasitología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Estrés Oxidativo/efectos de los fármacos , Parasitemia/patología , Parasitemia/prevención & control , Especies Reactivas de Oxígeno/metabolismo , Tasa de SupervivenciaRESUMEN
Pyrimethamine is an antimalarial drug that has also been used successfully to treat autoimmune diseases such as lymphoproliferative syndrome. In this work, the effect of pyrimethamine (PYR) on the production of free radicals in malaria-infected mice was studied to better understand the drug's immunomodulatory properties. BALB/c and CBA/Ca mice were infected with Plasmodium yoelii 17XL. Seven days after infection, mice were treated with PYR or vehicle and sacrificed 24h later. Treatment with PYR increased superoxide dismutase and glutathione peroxidase activities in erythrocytes and the liver, augmented the levels of nitric oxide in the serum, and upregulated mRNA levels of superoxide dismutase, glutathione peroxidase, catalase, and iNOS in the spleen. In addition, PYR increased lipoperoxidation and protein carbonylation in infected mice. Our results indicate that P. yoelii 17XL reduces oxidative stress in infected cells, while PYR induces it, which is associated with increased parasite elimination. Thus, it is possible that oxidative stress generated by pyrimethamine is also involved in its immunomodulatory mechanism of action.
Asunto(s)
Antimaláricos/farmacología , Malaria/tratamiento farmacológico , Estrés Oxidativo/efectos de los fármacos , Plasmodium yoelii/efectos de los fármacos , Pirimetamina/farmacología , Animales , Antimaláricos/uso terapéutico , Catalasa/biosíntesis , Catalasa/efectos de los fármacos , Catalasa/genética , Eritrocitos/efectos de los fármacos , Eritrocitos/metabolismo , Femenino , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Glutatión Peroxidasa/biosíntesis , Glutatión Peroxidasa/efectos de los fármacos , Glutatión Peroxidasa/genética , Peroxidación de Lípido/efectos de los fármacos , Hígado/efectos de los fármacos , Hígado/metabolismo , Malaria/inmunología , Malaria/metabolismo , Masculino , Malondialdehído/análisis , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos CBA , Óxido Nítrico/sangre , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Óxido Nítrico Sintasa de Tipo II/efectos de los fármacos , Óxido Nítrico Sintasa de Tipo II/genética , Estrés Oxidativo/inmunología , Plasmodium yoelii/genética , Plasmodium yoelii/inmunología , Carbonilación Proteica/efectos de los fármacos , Pirimetamina/uso terapéutico , Organismos Libres de Patógenos Específicos , Bazo/efectos de los fármacos , Bazo/metabolismo , Superóxido Dismutasa/biosíntesis , Superóxido Dismutasa/efectos de los fármacos , Superóxido Dismutasa/genética , Factores de TiempoRESUMEN
The molecular mechanisms by which the malarial parasite has managed to develop resistance to many antimalarial drugs remain to be completely elucidated. Mutations in the pfmdr1 gene of Plasmodium falciparum, as well as an increase in pfmdr1 copy number, have been associated with resistance to the quinoline-containing antimalarial drugs. We investigated the mechanisms of drug resistance in Plasmodium using a collection of P. yoelii lines with different drug resistance profiles. The mdr1 gene of P. yoelii (pymdr1) was identified and characterized. A 2- to 3-fold increase in the pymdr1 gene copy number was observed in the P. yoelii ART line (artemisinin resistant) when compared with the NS parental line. The pymdr1 gene was mapped to a chromosome of 2.1 Mb in all lines analyzed. Reverse transcriptase-polymerase chain reaction and Western blot experiments confirmed the expression of the gene at the RNA and protein levels.
Asunto(s)
Antimaláricos/farmacología , Artemisininas/farmacología , Genes MDR/genética , Plasmodium yoelii/genética , Secuencia de Aminoácidos , Animales , Western Blotting , Mapeo Cromosómico , Resistencia a Múltiples Medicamentos/genética , Femenino , Dosificación de Gen , Expresión Génica/genética , Genes MDR/fisiología , Malaria/tratamiento farmacológico , Malaria/parasitología , Ratones , Sistemas de Lectura Abierta/genética , Plasmodium yoelii/efectos de los fármacos , Proteínas Protozoarias/química , Proteínas Protozoarias/genética , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Chloroquine resistance of Plasmodium falciparum first and of P. vivax more recently, stimulated the search for new antimalarics. Chinese investigators have introduced new compounds obtained from extracts of Artemisia annua which possess an antimalaric active principle different from those of the drugs in use. In Mexico eight species of Artemisia have been described and among them just A. ludoviciana has been empirically used in the treatment of intermittent fever. To know whether mexican Artemisia had antimalaric activity several in vivo experiments were performed. Different type of extracts from two Artemisia species were prepared and assayed in five different doses on mice infected by Plasmodium yoelii yoelii, in a four-day test scheme. Here, only the results of the assays on ethanolic extract of A. ludoviciana are presented. The results of the in vivo experiments showed that the parasite reproduction was inhibited up to 98.6% at the fifth day, as compared with the controls; the ED50 was of 29.2 mg/kg and the SM50 of 28.7. We looked after the presence of artemisinin in the ethanolic extract, without success.