Goar Gevorkian

Two Epitopes Shared by Taenia crassiceps and Taenia solium Confer Protection against Murine T. crassiceps Cysticercosis along with a Prominent T1 Response

ABSTRACT Taenia crassiceps recombinant antigens KETc1 and KETc12 have been shown to induce high level of protection against experimental murine T. crassiceps cysticercosis, an experimental model successfully used to test candidate antigens for use in vaccination against porcine Taenia solium cysticercosis. Based on the deduced amino acid sequence, KETc1 and KETc12 were chemically synthesized in linear form. Immunization with KETc1 induced 66.7 to 100% protection against murine cysticercosis, and immunization with KETc12 induced 52.7 to 88.1% protection. The elicited immune response indicated that both peptides contain at least one B-cell epitope (as demonstrated by their ability to induce specific antibodies) and one T-cell epitope that strongly stimulated the proliferation of T cells primed with either the free peptide or total cysticercal T. crassiceps antigens. The high percentage of spleen cells expressing inflammatory cytokines points to the likelihood of a T1 response being involved in protection. The protective capacity of the peptides and their presence in all developmental stages of T. solium point to these two epitopes as strong candidates for inclusion in a polyepitopic synthetic vaccine against T. solium pig cysticercosis.

Protection against murine cysticercosis using cDNA expression library immunization.

Genetic or DNA-based immunization, including genomic immunization, has shown to be a viable alternative approach to induce protective immunity against a number of pathogens in several disease models. Here we describe a new method, cDNA expression library immunization (cDELI), based on the use of a large number of cDNA clones. This immunization strategy was tested in experimental murine Taenia crassiceps cysticercosis model. A partial cDNA expression library of 2 x 10(4) members was constructed in eukaryotic expression vector pcDNA3 and used to immunize BALB/c female mice subcutaneously (s.c.) and intramuscularly (i.m.). In both cases significant reduction of parasite load (up to 65%) was obtained. We were unable to directly measure T. crassiceps-specific humoral immune response in any of the immunized mice, although the expression of pathogen proteins in vitro in macrophages transfected with cDNA expressing plasmids was demonstrated. Also, in three out of five randomly selected immunized mice detectable levels of interferon-gamma (IFN-gamma) were obtained. cDELI has additional advantages compared with recently developed single gene or genomic immunization approaches because a cDNA population represents only those genes that are being expressed in the pathogen cells and the selection of stage-specific antigens is possible. The use of cDELI could be particularly attractive for the pathogens with complicated life cycles and large genomes.

Amyloid-β Peptide Binds to Cytochrome C Oxidase Subunit 1

Extracellular and intraneuronal accumulation of amyloid-beta aggregates has been demonstrated to be involved in the pathogenesis of Alzheimers disease (AD). However, the precise mechanism of amyloid-beta neurotoxicity is not completely understood. Previous studies suggest that binding of amyloid-beta to a number of macromolecules has deleterious effects on cellular functions. Mitochondria were found to be the target for amyloid-beta, and mitochondrial dysfunction is well documented in AD. In the present study we have shown for the first time that Aβ 1–42 bound to a peptide comprising the amino-terminal region of cytochrome c oxidase subunit 1. Phage clone, selected after screening of a human brain cDNA library expressed on M13 phage and bearing a 61 amino acid fragment of cytochrome c oxidase subunit 1, bound to Aβ 1–42 in ELISA as well as to Aβ aggregates present in AD brain. Aβ 1–42 and cytochrome c oxidase subunit 1 co-immunoprecipitated from mitochondrial fraction of differentiated human neuroblastoma cells. Likewise, molecular dynamics simulation of the cytochrome c oxidase subunit 1 and the Aβ 1–42 peptide complex resulted in a reliable helix-helix interaction, supporting the experimental results. The interaction between Aβ 1–42 and cytochrome c oxidase subunit 1 may explain, in part, the diminished enzymatic activity of respiratory chain complex IV and subsequent neuronal metabolic dysfunction observed in AD.

Immunodominant synthetic peptides of Taenia crassiceps in murine and human cysticercosis

The screening of a cDNA library of Taenia crassiceps revealed a clone designated KETc7 that induced high levels of protection against murine cysticercosis in previous experiments. The molecular structure of the deduced 100-amino acid sequence of the corresponding proline-rich polypeptide was studied to detect potentially immunologically active epitopes. Several candidate epitopes were identified, three of which were synthesized by solid-phase peptide synthesis and used as antigens in enzyme-linked immunosorbent assay (ELISA) for detection of specific antibodies in a selected panel of sera from mice infected with Taenia crassiceps and pigs infected with Taenia solium, as well as in the serum and cerebrospinal fluid of human patients with neurocysticercosis. The three peptides detected antibodies in serum from all infected mice. Seven of nine sera from patients with neurocysticercosis reacted strongly with peptide GK-3, and four of them with peptides GK-1 and GK-2. A lower reactivity was observed in sera from experimentally infected pigs. Peptide GK-3 reacted also with 45 out of 77 cerebrospinal fluids (CSF) from patients with confirmed neurocysticercosis and with 14 out of 68 CSF from control patients with other neurological disorders. This is the first report on synthetic peptides that are prominent in the humoral response of murine, porcine and human cysticercosis. Their identification implies finer molecular tools in the exploration of this form of host-parasite relationship, as well as hints to their application in immunodiagnosis and in vaccine design.

LPS-induced Murine Neuroinflammation Model: Main Features and Suitability for Pre-clinical Assessment of Nutraceuticals

Neuroinflammation is an important feature in the pathogenesis and progression of neurodegenerative diseases such as Alzheimer´s disease (AD), Parkinson´s disease (PD), frontotemporal dementia and amyotrophic lateral sclerosis. Based on current knowledge in the field, suggesting that targeting peripheral inflammation could be a promising additional treatment/prevention approach for neurodegenerative diseases, drugs and natural products with anti-inflammatory properties have been evaluated in animal models of neuroinflammation and neurodegeneration. In this review, we provide an extensive analysis of one of the most important and widely-used animal models of peripherally induced neuroinflammation and neurodegeneration - lipopolysaccharide (LPS)-treated mice, and address the data reproducibility in published research. We also summarize briefly basic features of various natural products, nutraceuticals, with known anti-inflammatory effects and present an overview of data on their therapeutic potential for reducing neuroinflammation in LPS-treated mice.

Peptide mimotopes of Mycobacterium tuberculosis carbohydrate immunodeterminants.

Cell-surface saccharides of Mycobacterium tuberculosis appear to be crucial factors in tuberculosis pathogenicity and could be useful antigens in tuberculosis immunodiagnosis. In the present study, we report the successful antigenic and immunogenic mimicry of mannose-containing cell-wall compounds of M. tuberculosis by dodecamer peptides identified by phage-display technology. Using a rabbit antiserum raised against M. tuberculosis cell-surface saccharides as a target for biopanning, peptides with three different consensus sequences were identified. Phage-displayed and chemically synthesized peptides bound to the anticarbohydrate antiserum. Rabbit antibodies elicited against the peptide QEPLMGTVPIRAGGGS recognize the mannosylated M. tuberculosis cell-wall antigens arabinomannan and lipoarabinomannan, and the glycosylated recombinant protein alanine/proline-rich antigen. Furthermore, antibodies were also able to react with mannan from Saccharomyces cerevisiae, but not with phosphatidylinositol dimannosides or arabinogalactan from mycobacteria. These results suggest that the immunogenic peptide mimics oligomannosidic epitopes. Interestingly, this report provides evidence that, in contrast with previously known carbohydrate mimotopes, no aromatic residues are necessary in a peptide sequence for mimicking unusual glycoconjugates synthesized by mycobacteria. The possible usefulness of the identified peptide mimotopes as surrogate reagents for immunodiagnosis and for the study of functional roles of the native non-peptide epitopes is discussed.

New approaches to improve a peptide vaccine against porcine Taenia solium cysticercosis.

Cysticercosis caused by Taenia solium frequently affects human health and rustic porciculture. Cysticerci may localize in the central nervous system of humans causing neurocysticercosis, a major health problem in undeveloped countries. Prevalence and intensity of this disease in pigs and humans are related to social factors (poor personal hygiene, low sanitary conditions, rustic rearing of pigs, open fecalism) and possibly to biological factors such as immunity, genetic background, and gender. The indispensable role of pigs as an obligatory intermediate host in the life cycle offers the possibility of interfering with transmission through vaccination of pigs. An effective vaccine based on three synthetic peptides against pig cysticercosis has been successfully developed and proved effective in experimental and field conditions. The well-defined peptides that constitute the cysticercosis vaccine offer the possibility to explore alternative forms of antigen production and delivery systems that may improve the cost/benefit of this and other vaccines. Encouraging results were obtained in attempts to produce large amounts of these peptides and increased its immunogenicity by expression in recombinant filamentous phage (M13), in transgenic plants (carrots and papaya), and associated to bacterial immunogenic carrier proteins.

Cysticercosis: towards the design of a diagnostic kit based on synthetic peptides

Cysticercosis caused by Taenia solium is a very common disease in developing countries that seriously affects human health. Diagnosis can only be confirmed with the aid of computerized tomography or nuclear magnetic resonance (NMR) creating obvious difficulties for epidemiological studies. Reliable immunoassays employing cerebrospinal fluid (CSF) have been developed, based on the use of cysticercal antigens. However, the reliance on parasite material is restrictive. Herein, we report the advances in the design of a diagnostic kit based on immunodominant synthetic peptides, targeting four candidate epitopes KETc1, KETc12, 410 and 413 which were identified from three different clones (KETc1, 12 and 4) selected from a cDNA library of Taenia crassiceps. CSF antibodies against T. solium cysticercal antigens (TCA) as well as the four peptides were determined by enzyme-linked immunoabsorbent assays (ELISA) using two panels of CSF from patients with confirmed neurocysticercosis and other neurological diseases. In the first CSF panel which included patients with high level of antibodies against TCA, KETc12 exhibited almost the same sensitivity (87.5%) as TCA (93.7%) and 100% specificity. In the second panel of 110 CSF collected at random, two peptides (KETc1 and KETc12) exhibited sensitivities of 40 and 36% respectively, and were 100% specific.

Pyroglutamate-Modified Amyloid Beta Peptides: Emerging Targets for Alzheimer´s Disease Immunotherapy

Extracellular and intraneuronal accumulation of amyloid-beta (Aβ) peptide aggregates in the brain has been hypothesized to play an important role in the neuropathology of Alzheimer’s Disease (AD). The main Aβ variants detected in the human brain are Aβ1-40 and Aβ1-42, however a significant proportion of AD brain Aβ consists also of N-terminal truncated species. Pyroglutamate-modified Aβ peptides have been demonstrated to be the predominant components among all N-terminal truncated Aβ species in AD brains and represent highly desirable and abundant therapeutic targets. The current review describes the properties and localization of two pyroglutamate-modified Aβ peptides, AβN3(pE) and AβN11(pE), in the brain. The role of glutaminyl cyclase (QC) in the formation of these peptides is also addressed. In addition, two potential therapeutic strategies, the inhibition of QC and immunotherapy approaches, and clinical trials aimed to target these important pathological Aβ species are reviewed.

Further evaluation of the synthetic peptide vaccine S3Pvac against Taenia solium cysticercosis in pigs in an endemic town of Mexico

Taenia solium cysticercosis is a parasitic disease frequently affecting human health and the pig industry in many developing countries. A synthetic peptide vaccine (designated S3Pvac) against porcine cysticercosis has been developed previously as an aid to interrupt transmission and has been shown to be effective. The results of the present study support the effectiveness of the vaccine under endemic field conditions. However, given the time-frame of the vaccination trial, no changes in the local levels of transmission were detectable before and after vaccination using sentinel pigs. Thus, this investigation shows the limited usefulness of single vaccination as the sole means of interrupting Taenia solium transmission in an endemic region.