RESUMEN
D. salina is one of the recognized natural sources to produce beta-carotene, and an useful model for studying the role of inhibitors and enhancers of carotenogenesis. However there is little information in D. salina regarding whether the isoprenoid substrate can be influenced by stress factors (carotenogenic) or selective inhibitors which in turn may further contribute to elucidate the early steps of carotenogenesis and biosynthesis of beta-carotene. In this study, Dunaliella salina (BC02) isolated from La Salina BC Mexico, was subjected to the method of isoprenoids-beta-carotene interference in order to promote the interruption or accumulation of the programmed biosynthesis of carotenoids. When Carotenogenic and non-carotenogenic cells of D. salina BC02 were grown under photoautotrophic growth conditions in the presence of 200 microM fosmidomycin, carotenogenesis and the synthesis of beta-carotene were interrupted after two days in cultured D. salina cells. This result is an indirect consequence of the inhibition of the synthesis of isoprenoids and activity of the recombinant DXR enzyme thereby preventing the conversion of 1-deoxy-D-xylulose 5-phosphate (DXP) to 2-C-methyl-D-erythritol (MEP) and consequently interrupts the early steps of carotenogenesis in D. salina. The effect at the level of proteins and RNA was not evident. Mevinolin treated D. salina cells exhibited carotenogenesis and beta-carotene levels very similar to those of control cell cultures indicating that mevinolin not pursued any indirect action in the biosynthesis of isoprenoids and had no effect at the level of the HMG-CoA reductase, the key enzyme of the Ac/MVA pathway.
Asunto(s)
Carotenoides/biosíntesis , Chlorophyta/aislamiento & purificación , Terpenos/metabolismo , California , Células Cultivadas , Chlorophyta/crecimiento & desarrollo , Chlorophyta/metabolismo , Eritritol/análogos & derivados , Eritritol/metabolismo , Fosfomicina/análogos & derivados , Fosfomicina/farmacología , Lovastatina/farmacología , México , Pentosafosfatos/metabolismo , Fosfatos de Azúcar/metabolismo , beta Caroteno/biosíntesisRESUMEN
Polysaccharide-protein conjugates as vaccines have proven to be very effective in preventing Haemophilus influenzae type b infections in industrialized countries. However, cost-effective technologies need to be developed for increasing the availability of anti-H. influenzae type b vaccines in countries from the developing world. Consequently, vaccine production with partially synthetic antigens is a desirable goal for many reasons. They may be rigidly controlled for purity and effectiveness while at the same time being cheap enough that they may be made universally available. We describe here the antigenicity and immunogenicity of several H. influenzae type b synthetic oligosaccharide-protein conjugates in laboratory animals. The serum of H. influenzae type b-immunized animals recognized our synthetic H. influenzae type b antigens to the same extent as the native bacterial capsular polysaccharide. Compared to the anti-H. influenzae type b vaccine employed, these synthetic versions induced similar antibody response patterns in terms of titer, specificity, and functional capacity. The further development of synthetic vaccines will meet urgent needs in the less prosperous parts of the world and remains our major goal(AU)
Conjugados proteína-polisacárido como vacunas han demostrado ser muy eficaz en la prevención de Haemophilus influenzae tipo b infecciones en los países industrializados. Sin embargo, tecnologías rentables necesitan ser desarrolladas para incrementar la disponibilidad de anti-H. influenzae tipo b, las vacunas en los países del mundo en desarrollo. En consecuencia, la producción de vacunas con antígenos sintéticos en parte es un objetivo deseable por muchas razones. Pueden ser rígidamente controlada por la pureza y la eficacia al mismo tiempo ser lo suficientemente baratas para que sean universalmente disponibles. Se describe aquí la antigenicidad e inmunogenicidad de varias H. influenzae tipo b-oligosacáridos sintéticos conjugados proteína en animales de laboratorio. El suero de H. influenzae tipo b-reconocido nuestros animales inmunizados sintéticas H. influenzae tipo b antígenos en la misma medida que los nativos bacteriana polisacárido capsular. En comparación con el anti-H. influenzae tipo b vacuna empleada, estas versiones sintéticas similares respuesta de anticuerpos inducida por los patrones en términos de títulos, la especificidad y la capacidad funcional. El ulterior desarrollo de vacunas sintéticas satisfacer las necesidades urgentes en las regiones menos prósperas del mundo y sigue siendo nuestro principal objetivo
Asunto(s)
Animales , Ratones , Conejos , Haemophilus influenzae tipo b/inmunología , Pentosafosfatos/inmunología , Polisacáridos Bacterianos/inmunología , Vacunas Sintéticas/inmunologíaRESUMEN
Two genes encoding the enzymes 1-deoxy-D-xylulose-5-phosphate synthase and 1-deoxy-D-xylulose-5-phosphate reductoisomerase have been recently identified, suggesting that isoprenoid biosynthesis in Plasmodium falciparum depends on the methylerythritol phosphate (MEP) pathway, and that fosmidomycin could inhibit the activity of 1-deoxy-D-xylulose-5-phosphate reductoisomerase. The metabolite 1-deoxy-D-xylulose-5-phosphate is not only an intermediate of the MEP pathway for the biosynthesis of isopentenyl diphosphate but is also involved in the biosynthesis of thiamin (vitamin B1) and pyridoxal (vitamin B6) in plants and many microorganisms. Herein we report the first isolation and characterization of most downstream intermediates of the MEP pathway in the three intraerythrocytic stages of P. falciparum. These include, 1-deoxy-D-xylulose-5-phosphate, 2-C-methyl-D-erythritol-4-phosphate, 4-(cytidine-5-diphospho)-2-C-methyl-D-erythritol, 4-(cytidine-5-diphospho)-2-C-methyl-D-erythritol-2-phosphate, and 2-C-methyl-D-erythritol-2,4-cyclodiphosphate. These intermediates were purified by HPLC and structurally characterized via biochemical and electrospray mass spectrometric analyses. We have also investigated the effect of fosmidomycin on the biosynthesis of each intermediate of this pathway and isoprenoid biosynthesis (dolichols and ubiquinones). For the first time, therefore, it is demonstrated that the MEP pathway is functionally active in all intraerythrocytic forms of P. falciparum, and de novo biosynthesis of pyridoxal in a protozoan is reported. Its absence in the human host makes both pathways very attractive as potential new targets for antimalarial drug development.
Asunto(s)
Eritritol/análogos & derivados , Eritritol/metabolismo , Fosfomicina/análogos & derivados , Plasmodium falciparum/metabolismo , Fosfato de Piridoxal/análogos & derivados , Fosfatos de Azúcar/metabolismo , Animales , Antimaláricos/farmacología , Dolicoles/biosíntesis , Eritrocitos/parasitología , Fosfomicina/farmacología , Genes Protozoarios , Humanos , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/parasitología , Estructura Molecular , Pentosafosfatos/biosíntesis , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/genética , Plasmodium falciparum/crecimiento & desarrollo , Fosfato de Piridoxal/biosíntesis , Espectrometría de Masa por Ionización de Electrospray , Ubiquinona/biosíntesisRESUMEN
To determine the immunogenicity of Haemophilus influenzae type b polysaccharide-tetanus protein conjugate vaccine in specific populations at risk, we administered vaccine to children with sickle cell anemia (n = 19; mean age, 18.3 months, malignancies (n = 18; mean age, 43.1 months), or a recent history of systemic H. influenzae type b infection (n = 17; mean age, 11.9 months). After one dose of polyribosylribitol phosphate-tetanus toxoid conjugate vaccine the geometric mean titers for polyribosylribitol phosphate antibody were 4.8 micrograms/ml (14/19 greater than 1 microgram/ml), 1.4 micrograms/ml (9/18 greater than 1 microgram/ml), and 5.6 micrograms/ml (15/17 greater than 1 microgram/ml) in these three groups, respectively. Children with sickle cell anemia or a recent history of systemic H. influenzae type b infection had polyribosylribitol phosphate antibody levels comparable to those of normal children of similar age after one or two doses of polyribosylribitol phosphate-tetanus toxoid conjugate vaccine. We conclude that this vaccine is immunogenic in children with underlying conditions associated with an increased risk of H. influenzae type b infection.
Asunto(s)
Anemia de Células Falciformes/inmunología , Anticuerpos Antibacterianos/biosíntesis , Vacunas Bacterianas/inmunología , Infecciones por Haemophilus/inmunología , Vacunas contra Haemophilus , Haemophilus influenzae/inmunología , Neoplasias/inmunología , Toxoide Tetánico/inmunología , Preescolar , Humanos , Lactante , Neoplasias/tratamiento farmacológico , Pentosafosfatos/inmunología , Polisacáridos Bacterianos/inmunologíaAsunto(s)
Anticuerpos Antibacterianos/inmunología , Proteínas de la Membrana Bacteriana Externa/administración & dosificación , Vacunas Bacterianas/administración & dosificación , Haemophilus influenzae/inmunología , Neisseria meningitidis/inmunología , Pentosafosfatos/administración & dosificación , Polisacáridos Bacterianos/administración & dosificación , Proteínas de la Membrana Bacteriana Externa/inmunología , Vacunas Bacterianas/inmunología , Humanos , Esquemas de Inmunización , Lactante , Pentosafosfatos/inmunología , Polisacáridos Bacterianos/inmunologíaRESUMEN
We studied an immunogen consisting of oligosaccharides derived from Haemophilus influenzae type b capsular polysaccharide (PRP) coupled to CRM197, a nontoxic relative of diphtheria toxin. Subcutaneous injections were given to eight subjects at ages 2, 4, and 6 months, simultaneously with conventional diphtheria-tetanus-pertussis (DTP) vaccine. After the first immunization, total serum anti-PRP antibodies declined in all subjects, but increased in most after the second immunization and after the third in seven of seven subjects analyzed. In these seven infants, the geometric mean level at age 9 months (0.73 micrograms/ml) exceeded by at least 40 times the means of historical control groups given DTP only or DTP plus (uncoupled) PRP vaccine. An isotype-specific assay showed that IgM antibodies increased after the first immunization with the coupled vaccine in all eight infants. Against the background of declining maternal IgG antibody, elevations in IgG antibody were detected after the second or third immunization in six of the eight. These six at age 9 to 11 months were immunized with (uncoupled) PRP vaccine, and a "boost" in anti-PRP antibody, including an IgG component, was found.
Asunto(s)
Vacunas Bacterianas/administración & dosificación , Haemophilus influenzae/inmunología , Pentosafosfatos/administración & dosificación , Polisacáridos Bacterianos/administración & dosificación , Vacunación/métodos , Anticuerpos Antibacterianos/biosíntesis , Proteínas Bacterianas/inmunología , Vacunas Bacterianas/efectos adversos , Infecciones por Haemophilus/prevención & control , Humanos , Inmunoglobulina A/análisis , Inmunoglobulina G/análisis , Inmunoglobulina M/análisis , Lactante , Pentosafosfatos/inmunología , Polisacáridos Bacterianos/inmunología , Factores de TiempoAsunto(s)
Vacunas Bacterianas/administración & dosificación , Haemophilus influenzae/inmunología , Pentosafosfatos/inmunología , Polisacáridos Bacterianos/inmunología , Factores de Edad , Anticuerpos Antibacterianos/biosíntesis , Vacunas Bacterianas/inmunología , Vacunas Bacterianas/uso terapéutico , Niño , Preescolar , Ensayos Clínicos como Asunto , Finlandia , Infecciones por Haemophilus/prevención & control , Humanos , Inmunoglobulina G/biosíntesis , Inmunoglobulina M/biosíntesis , Lactante , Meningitis por Haemophilus/epidemiología , Meningitis por Haemophilus/prevención & control , Factores de Tiempo , Estados Unidos , Vacunación/métodosRESUMEN
Sixty 9- to 15-month-old infants were randomly assigned to receive two doses, 1 month apart, of a Haemophilus influenzae type b capsular polysaccharide-diphtheria toxoid conjugate vaccine (PRP-D) or PRP vaccine, each containing 20 micrograms PRP. There were no significant local or systemic reactions. After one dose of PRP-D, 93% of the subjects attained levels of greater than or equal to 0.15 microgram/ml and 59% achieved greater than or equal to 1 microgram/ml antibody protein. These percentages rose to 100% and 86%, respectively, after the second dose, at which time the geometric mean titer of anti-PRP antibody was 4.8 micrograms/ml. IgG anti-PRP levels were 4.3 times higher than IgM. The proportion of IgG to IgM antibody induced by PRP-D increased with age. After two doses, 33% of the PRP recipients responded with a level of greater than or equal to 0.15 microgram/ml and only 19% responded to a level of greater than or equal to 1.0 microgram/ml. One year later, all of the PRP-D recipients tested still had greater than or equal to 0.15 microgram/ml and more than half had greater than or equal to 1.0 microgram/ml antibody protein.
Asunto(s)
Anticuerpos Antibacterianos/biosíntesis , Vacunas Bacterianas/administración & dosificación , Toxoide Diftérico/administración & dosificación , Infecciones por Haemophilus/inmunología , Pentosafosfatos/administración & dosificación , Polisacáridos Bacterianos/administración & dosificación , Anticuerpos Antibacterianos/análisis , Vacunas Bacterianas/efectos adversos , Vacunas Bacterianas/inmunología , Toxoide Diftérico/efectos adversos , Toxoide Diftérico/inmunología , Infecciones por Haemophilus/prevención & control , Haemophilus influenzae/inmunología , Humanos , Esquemas de Inmunización , Inmunización Secundaria , Inmunoglobulina A/análisis , Inmunoglobulina M/análisis , Lactante , Pentosafosfatos/efectos adversos , Pentosafosfatos/inmunología , Polisacáridos Bacterianos/efectos adversos , Polisacáridos Bacterianos/inmunologíaRESUMEN
Glucose consumption by anaerobic glycolysis and the pentose pathway were studied in two Aymara populations living at different altitudes (3 600 m and 450 m). The measurements were made both with and without methylene blue. We observed a Pasteur effect for both pathways which may explain the increase in 2-3 DPG and ATP levels found in blood samples from people living at high altitudes. The results in the presence of methylene blue showed a reduced activity of the methaemoglobin reductase system in the high altitude group which may be partly responsible for their increased levels of methaemoglobin.