ABSTRACT
First synthesis of novel coumarin-trioxane hybrids is reported. The synthesis was achieved via condensation of ß-hydroxyhydroperoxides with coumarinic-aldehydes in presence of p-toluenesulfonic acid in good yields and the novel hybrids were evaluated for their antimalarial activity both in vitro and in vivo.
Subject(s)
Antimalarials/chemical synthesis , Coumarins/chemistry , Heterocyclic Compounds/chemistry , Malaria, Falciparum/drug therapy , Plasmodium falciparum/drug effects , Plasmodium yoelii/drug effects , Aldehydes/chemistry , Animals , Antimalarials/pharmacology , Benzenesulfonates/chemistry , Coumarins/pharmacology , Erythrocytes/drug effects , Erythrocytes/parasitology , Heterocyclic Compounds/pharmacology , Humans , Inhibitory Concentration 50 , Malaria, Falciparum/parasitology , Mice , Peroxides/chemistry , Plasmodium falciparum/parasitology , Plasmodium yoelii/parasitology , Structure-Activity RelationshipABSTRACT
Methods facilitating research in malaria are of pivotal relevance. Flow cytometry offers the possibility of rapid enumeration of parasitemia. It relies on staining the parasite DNA to distinguish between infected and non-infected red blood cell (RBC) populations. Unfortunately, in rodents abundant reticulocyte RNA interferes with the application of the method. This results in time-consuming sample preparation protocols that offer no clear advantage over microscopic counting. We re-evaluated the use of the DNA/RNA discriminating vital fluorochrome acridine orange (AO) for rapid flow cytometric enumeration of parasitemia in rodents. Whole blood from rodents infected with Plasmodium berghei and Plasmodium yoelii was stained with AO and analyzed by flow cytometer. A newly developed two-channel (FL1/FL3) detection method was compared with conventional one-channel (FL1) detection and microscopic counting. The new AO two-channel detection method clearly discriminated between infected and non-infected RBC populations. It showed to be linear above parasitemias of 0.3%. Sample processing time amounted to approximately 5 min. It is shown that AO can be used for rapid, precise, and accurate enumeration of parasitemia in rodents. Due to its ease of handling the method might find widespread application in malaria research.
Subject(s)
Acridine Orange , Erythrocytes/parasitology , Flow Cytometry/methods , Fluorescent Dyes , Malaria/parasitology , Plasmodium berghei/isolation & purification , Plasmodium yoelii/isolation & purification , Animals , Cricetinae , Malaria/diagnosis , Mesocricetus , Mice , Mice, Inbred ICR , Parasitemia/parasitology , Plasmodium berghei/parasitology , Plasmodium yoelii/parasitology , Staining and LabelingABSTRACT
This study explores the utility of recombinant adeno-associated virus (rAAV) as a genetic vaccine delivery system using muscle as a target tissue. A single injection of rAAV encoding the malarial antigens MSP4 (Plasmodium falciparum) or MSP4/5 (Plasmodium yoelii) stimulated long-term antigen-specific antibody responses. Anti-MSP4/5 immunity stimulated by AAV was not protective against P. yoelii infection and efforts taken to augment antibody responses against MSP4/5, either by priming with plasmid DNA or AAV and boosting with rAAV were unsuccessful. Alternative strategies such as inclusion of genetic adjuvants into the AAV vector will be necessary to stimulate an adequate level of anti-malarial protective immunity in this model.
Subject(s)
Antigens, Protozoan/immunology , Dependovirus/genetics , Malaria Vaccines/administration & dosage , Malaria/immunology , Membrane Proteins/immunology , Plasmodium falciparum/immunology , Plasmodium yoelii/immunology , Protozoan Proteins/immunology , Adjuvants, Immunologic/pharmacology , Animals , Antibody Formation/immunology , Antigens, Protozoan/genetics , Dependovirus/immunology , Epitopes , Female , Genetic Vectors/genetics , Genetic Vectors/immunology , HeLa Cells , Humans , Injections, Intramuscular , Malaria/parasitology , Malaria/prevention & control , Malaria Vaccines/genetics , Malaria Vaccines/immunology , Membrane Proteins/genetics , Mice , Mice, Inbred BALB C , Plasmodium falciparum/parasitology , Plasmodium yoelii/parasitology , Protozoan Proteins/geneticsABSTRACT
The youngest (rings and young trophozoites) erythrocytic stages of Plasmodium yoelii nigeriensis and P. yoelii killicki, two rodent malaria subspecies developing very asynchronously in the blood, were separated from the other stages using a discontinuous Percoll-glucose gradient. They were inoculated into mice and the subsequent infection remained synchronous for two generations. The duration of the asexual cycle was found to be 18 h.
