Cellular responses to modified Plasmodium falciparum MSP119 antigens in individuals previously exposed to natural malaria infection

<p>Abstract</p> <p>Background</p> <p>MSP1 processing-inhibitory antibodies bind to epitopes on the 19 kDa C-terminal region of the <it>Plasmodium falciparum </it>merozoite surface protein 1 (MSP1₁₉), inhibiting erythrocyte invasion. Blocking antibodies also bind to this antigen but prevent inhibitory antibodies binding, allowing invasion to proceed. Recombinant MSP1₁₉had been modified previously to allow inhibitory but not blocking antibodies to continue to bind. Immunization with these modified proteins, therefore, has the potential to induce more effective protective antibodies. However, it was unclear whether the modification of MSP1₁₉would affect critical T-cell responses to epitopes in this antigen.</p> <p>Methods</p> <p>The cellular responses to wild-type MSP1₁₉and a panel of modified MSP1₁₉antigens were measured using an <it>in-vitro </it>assay for two groups of individuals: the first were malaria-naïve and the second had been naturally exposed to <it>Plasmodium falciparum </it>infection. The cellular responses to the modified proteins were examined using cells from malaria-exposed infants and adults.</p> <p>Results</p> <p>Interestingly, stimulation indices (SI) for responses induced by some of the modified proteins were at least two-fold higher than those elicited by the wild-type MSP1₁₉. A protein with four amino acid substitutions (Glu27→Tyr, Leu31→Arg, Tyr34→Ser and Glu43→Leu) had the highest stimulation index (SI up to 360) and induced large responses in 64% of the samples that had significant cellular responses to the modified proteins.</p> <p>Conclusion</p> <p>This study suggests that specific MSP1₁₉variants that have been engineered to improve their antigenicity for inhibitory antibodies, retain T-cell epitopes and the ability to induce cellular responses. These proteins are candidates for the development of MSP1-based malaria vaccines.</p

Clustering-based approaches to SAGE data mining

Serial analysis of gene expression (SAGE) is one of the most powerful tools for global gene expression profiling. It has led to several biological discoveries and biomedical applications, such as the prediction of new gene functions and the identification of biomarkers in human cancer research. Clustering techniques have become fundamental approaches in these applications. This paper reviews relevant clustering techniques specifically designed for this type of data. It places an emphasis on current limitations and opportunities in this area for supporting biologically-meaningful data mining and visualisation

Monocyte Gene Expression Signature of Patients with Early Onset Coronary Artery Disease

The burden of cardiovascular disease (CVD) cannot be fully addressed by therapy targeting known pathophysiological pathways. Even with stringent control of all risk factors CVD events are only diminished by half. A number of additional pathways probably play a role in the development of CVD and might serve as novel therapeutic targets. Genome wide expression studies represent a powerful tool to identify such novel pathways. We compared the expression profiles in monocytes from twenty two young male patients with premature familial CAD with those from controls matched for age, sex and smoking status, without a family history of CVD. Since all patients were on statins and aspirin treatment, potentially affecting the expression of genes in monocytes, twelve controls were subsequently treated with simvastatin and aspirin for 6 and 2 weeks, respectively

Gene Expression Signature in Peripheral Blood Detects Thoracic Aortic Aneurysm

BACKGROUND: Thoracic aortic aneurysm (TAA) is usually asymptomatic and associated with high mortality. Adverse clinical outcome of TAA is preventable by elective surgical repair; however, identifying at-risk individuals is difficult. We hypothesized that gene expression patterns in peripheral blood cells may correlate with TAA disease status. Our goal was to identify a distinct gene expression signature in peripheral blood that may identify individuals at risk for TAA. METHODS AND FINDINGS: Whole genome gene expression profiles from 94 peripheral blood samples (collected from 58 individuals with TAA and 36 controls) were analyzed. Significance Analysis of Microarray (SAM) identified potential signature genes characterizing TAA vs. normal, ascending vs. descending TAA, and sporadic vs. familial TAA. Using a training set containing 36 TAA patients and 25 controls, a 41-gene classification model was constructed for detecting TAA status and an overall accuracy of 78+/-6% was achieved. Testing this classifier on an independent validation set containing 22 TAA samples and 11 controls yielded an overall classification accuracy of 78%. These 41 classifier genes were further validated by TaqMan real-time PCR assays. Classification based on the TaqMan data replicated the microarray results and achieved 80% classification accuracy on the testing set. CONCLUSIONS: This study identified informative gene expression signatures in peripheral blood cells that can characterize TAA status and subtypes of TAA. Moreover, a 41-gene classifier based on expression signature can identify TAA patients with high accuracy. The transcriptional programs in peripheral blood leading to the identification of these markers also provide insights into the mechanism of development of aortic aneurysms and highlight potential targets for therapeutic intervention. The classifier genes identified in this study, and validated by TaqMan real-time PCR, define a set of promising potential diagnostic markers, setting the stage for a blood-based gene expression test to facilitate early detection of TAA

Tobacco use induces anti-apoptotic, proliferative patterns of gene expression in circulating leukocytes of Caucasian males

Abstract Background Strong epidemiologic evidence correlates tobacco use with a variety of serious adverse health effects, but the biological mechanisms that produce these effects remain elusive. Results We analyzed gene transcription data to identify expression spectra related to tobacco use in circulating leukocytes of 67 Caucasian male subjects. Levels of cotinine, a nicotine metabolite, were used as a surrogate marker for tobacco exposure. Significance Analysis of Microarray and Gene Set Analysis identified 109 genes in 16 gene sets whose transcription levels were differentially regulated by nicotine exposure. We subsequently analyzed this gene set by hyperclustering, a technique that allows the data to be clustered by both expression ratio and gene annotation (e.g. Gene Ontologies). Conclusion Our results demonstrate that tobacco use affects transcription of groups of genes that are involved in proliferation and apoptosis in circulating leukocytes. These transcriptional effects include a repertoire of transcriptional changes likely to increase the incidence of neoplasia through an altered expression of genes associated with transcription and signaling, interferon responses and repression of apoptotic pathways

Formation of the Food Vacuole in Plasmodium falciparum: A Potential Role for the 19 kDa Fragment of Merozoite Surface Protein 1 (MSP119)

Plasmodium falciparum Merozoite Surface Protein 1 (MSP1) is synthesized during schizogony as a 195-kDa precursor that is processed into four fragments on the parasite surface. Following a second proteolytic cleavage during merozoite invasion of the red blood cell, most of the protein is shed from the surface except for the C-terminal 19-kDa fragment (MSP119), which is still attached to the merozoite via its GPI-anchor. We have examined the fate of MSP119 during the parasite's subsequent intracellular development using immunochemical analysis of metabolically labeled MSP119, fluorescence imaging, and immuno-electronmicroscopy. Our data show that MSP119 remains intact and persists to the end of the intracellular cycle. This protein is the first marker for the biogenesis of the food vacuole; it is rapidly endocytosed into small vacuoles in the ring stage, which coalesce to form the single food vacuole containing hemozoin, and persists into the discarded residual body. The food vacuole is marked by the presence of both MSP119 and the chloroquine resistance transporter (CRT) as components of the vacuolar membrane. Newly synthesized MSP1 is excluded from the vacuole. This behavior indicates that MSP119 does not simply follow a classical lysosome-like clearance pathway, instead, it may play a significant role in the biogenesis and function of the food vacuole throughout the intra-erythrocytic phase

A principal target of human immunity to malaria identified by molecular population genetic and immunological analyses

New strategies are required to identify the most important targets of protective immunity in complex eukaryotic pathogens. Natural selection maintains allelic variation in some antigens of the malaria parasite Plasmodium falciparum (1–3). Analysis of allele frequency distributions could identify the loci under most intense selection (4–7). The merozoite surface protein 1 (Msp1) is the most-abundant surface component on the erythrocyte-invading stage of P. falciparum (8–10). Immunization with whole Msp1 has protected monkeys completely against homologous (11) and partially against non-homologous (12) parasite strains. The singlecopy msp1 gene, of about 5 kilobases, has highly divergent alleles (13) with stable frequencies in endemic populations (14,15). To identify the region of msp1 under strongest selection to maintain alleles within populations, we studied multiple intragenic sequence loci in populations in different regions of Africa and Southeast Asia. On both continents, the locus with the lowest inter-population variance in allele frequencies was block 2, indicating selection in this part of the gene. To test the hypothesis of immune selection, we undertook a large prospective longitudinal cohort study. This demonstrated that serum IgG antibodies against each of the two most frequent allelic types of block 2 of the protein were strongly associated with protection from P. falciparum malaria

New insights for native production of MSP1(19), the disulfide-rich C-terminal fragment from Plasmodium falciparum merozoite surface protein 1.

Malaria represents a major public health problem and an important cause of mortality and morbidity. The malaria parasites are becoming resistant to drugs used to treat the disease and still no efficient vaccine has been developed. One promising vaccine candidate is the merozoite surface protein 1 (MSP1), which has been extensively investigated as a vaccine target. The surface protein MSP1 plays an essential role in the erythrocyte invasion process and is an accessible target for the immune system. Antibodies to the carboxy-terminal region of the protein, named MSP119, can inhibit erythrocyte invasion and parasite growth. In order to develop an effective MSP119- based vaccine against malaria, production of an antigen that is recognized by protective antibodies is mandatory. To this aim, we propose a method to produce the disulfide-rich MSP119 in its native conformation based on its in vitro oxidative refolding. The native conformation of the renatured MSP119 is carefully established by immunochemical reactivity experiments, circular dichroism and NMR. MSP119 can successfully be refolded in vitro as an isolated protein or as a fusion with the maltose binding protein. The possibility to properly fold MSP119in vitro paves the way to new approaches for high titer production of native MSP119 using Escherichia coli as a host