RESUMEN
Merozoite surface protein-1 (MSP-1), a major asexual blood stage antigen, and circumsporozoite protein (CSP), a component of sporozoites that includes a Plasmodium vivax B-cell epitope, are strong candidates for use in a malaria vaccine. A chimeric recombinant gene containing portions of both msp-1 and csp from P. vivax separated by Pro-Gly linker motif was generated. The construct gene was named mlc (msp-1, linker, and csp). The MLC chimeric recombinant protein had a molecular weight of approximately 25 kDa when expressed in Escherichia coli, as determined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The purified chimeric protein reacted with the sera of patients infected with P. vivax but not with the sera of uninfected patients according to western blot analysis. The chimeric protein reacted well with sera of malaria patients (109/115, 94.78%) as assessed with enzyme-linked immunosorbent assay (ELISA). BALB/c mice that were orally immunized with the MLC chimeric recombinant protein successfully produced antigen-specific antibodies. Additionally, levels of the Th1-associated cytokines IL-12(p40), TNF-α, and IFN-γ were significantly increased in the spleens of the BALB/c mice. Therefore, the E. coli-expressed MLC chimeric recombinant protein might be used as a valuable vaccine candidate for oral immunization against vivax malaria.
Asunto(s)
Proteína 1 de Superficie de Merozoito/genética , Plasmodium vivax/genética , Proteínas Protozoarias/genética , Proteínas Recombinantes de Fusión/genética , Secuencia de Aminoácidos , Animales , Anticuerpos Antiprotozoarios/biosíntesis , Anticuerpos Antiprotozoarios/sangre , Secuencia de Bases , Femenino , Expresión Génica , Humanos , Vacunas contra la Malaria , Malaria Vivax/prevención & control , Proteína 1 de Superficie de Merozoito/biosíntesis , Proteína 1 de Superficie de Merozoito/inmunología , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Plasmodium vivax/inmunología , Plasmodium vivax/metabolismo , Proteínas Protozoarias/biosíntesis , Proteínas Protozoarias/inmunología , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/inmunologíaRESUMEN
BACKGROUND: To develop a plant-based vaccine against Plasmodium vivax, two P. vivax candidate proteins were chosen. First, the merozoite surface protein-1 (MSP-1), a major asexual blood stage antigen that is currently considered a strong vaccine candidate. Second, the circumsporozoite protein (CSP), a component of sporozoites that contains a B-cell epitope. METHODS: A synthetic chimeric recombinant 516 bp gene encoding containing PvMSP-1, a Pro-Gly linker motif, and PvCSP was synthesized; the gene, named MLC, encoded a total of 172 amino acids. The recombinant gene was modified with regard to codon usage to optimize gene expression in Brassica napus. The Ti plasmid inducible gene transfer system was used for MLC chimeric recombinant gene expression in B. napus. Gene expression was confirmed by polymerase chain reaction (PCR), beta-glucuronidase reporter gene (GUS) assay, and Western blot. RESULTS: The MLC chimeric recombinant protein expressed in B. napus had a molecular weight of approximately 25 kDa. It exhibited a clinical sensitivity of 84.21% (n=38) and a clinical specificity of 100% (n=24) as assessed by enzyme-linked immunosorbent assay (ELISA). Oral immunization of BALB/c mice with MLC chimeric recombinant protein successfully induced antigen-specific IgG1 production. Additionally, the Th1-related cytokines IL-12 (p40), TNF, and IFN-γ were significantly increased in the spleens of the BALB/c mice. CONCLUSIONS: The chimeric MLC recombinant protein produced in B. napus has potential as both as an antigen for diagnosis and as a valuable vaccine candidate for oral immunization against vivax malaria.
Asunto(s)
Brassica napus/metabolismo , Vacunas contra la Malaria/inmunología , Proteína 1 de Superficie de Merozoito/inmunología , Plasmodium vivax/inmunología , Proteínas Protozoarias/inmunología , Administración Oral , Animales , Anticuerpos Antiprotozoarios/sangre , Antígenos de Protozoos/biosíntesis , Antígenos de Protozoos/genética , Antígenos de Protozoos/inmunología , Brassica napus/genética , Codón , Ensayo de Inmunoadsorción Enzimática/métodos , Expresión Génica , Vectores Genéticos , Inmunoglobulina G/sangre , Vacunas contra la Malaria/administración & dosificación , Vacunas contra la Malaria/biosíntesis , Vacunas contra la Malaria/genética , Masculino , Proteína 1 de Superficie de Merozoito/biosíntesis , Proteína 1 de Superficie de Merozoito/genética , Ratones , Ratones Endogámicos BALB C , Peso Molecular , Plásmidos Inductores de Tumor en Plantas , Plasmodium vivax/genética , Proteínas Protozoarias/biosíntesis , Proteínas Protozoarias/genética , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Sensibilidad y Especificidad , Vacunas Sintéticas/administración & dosificación , Vacunas Sintéticas/biosíntesis , Vacunas Sintéticas/genética , Vacunas Sintéticas/inmunologíaRESUMEN
Magnetic properties of 200 nm ferrimagnetic CoFe2O4 nanoparticles before and after coating with TEOS were explored and compared to soft ferrimagnetic MgFe2O4 nanoparticles (200 nm) to evaluate the feasibility as an in-vitro GMR SV (giant magnetoresistance spin-valve) biosensor agent for single molecular detection (SMD). It was found that the magnetic degradation (or variation) of TEOS coated CoFe2O4 and MgFe2O4 nanoparticles are dominantly affected by the chemical dispersion process, which is carried out in the oleic acid (OA), oleylamine (OL), or OA+OL surfactant, before starting major coating process. In addition, the TEOS coating thickness controlled by TEOS concentration and pH level in the buffer solution prominently influenced on the magnetic degradation of TEOS coated nanoparticles. According to the experimental analysis results, the magnetic degradation of TEOS coated nanoparticles is mainly attributed to the variation of particle dipole interaction caused by the degree of particle aggregation depending on TEOS coating process conditions. The TEOS coated CoFe2O4 nanoparticles exhibited a higher magnetic stability for a GMR biosensor agent, e.g., small variation of remnant magnetization, saturation magnetization and magnetic coercivity, than that of MgFe2O4 nanoparticles at the different coating process conditions. The physical and chemical analysis confirmed that this is primarily due to its higher magnetic anisotropy. The experimentally verified high biocompatibility as well as the stably maintained magnetic properties of TEOS coated CoFe2O4 nanoparticles demonstrate that CoFe2O4 nanoparticles can be considered as one of the promising ferrimagnetic nanoparticle sensor agent for an SMD GMR SV biosensor.
Asunto(s)
Técnicas Biosensibles/métodos , Cobalto/química , Hierro/química , Nanopartículas del Metal/química , Silanos/química , Animales , Línea Celular , Campos Electromagnéticos , Estudios de Factibilidad , Compuestos Férricos/química , Compuestos de Magnesio/química , Microscopía Electrónica de Rastreo , Nanotecnología , Ratas , Espectroscopía Infrarroja por Transformada de FourierRESUMEN
Ocular neuroprotection induced by localized heat shock proteins (HSPs) has been paid considerable attention as an efficacious treatment modality for glaucoma. However, the current clinical approaches to induce HSPs in the retinal ganglion cells (RGCs) are limited due to undesirable side effects. Here, we present that the induction of HSPs by local magnetic hyperthermia using engineered superparamagnetic Mn(0.5)Zn(0.5)Fe(2)O(4) nanoparticle agents (EMZF-SPNPAs) with a 5.5 nm mean particle size is promisingly feasible for a physiologically tolerable ocular neuroprotection modality. The sufficiently high specific absorption rate (SAR) (â¼256.4 W/g in an agar solution) achieved at the biologically safe range of applied AC magnetic field and frequency as well as the superior biocompatibility of EMZF-SPNPA, which were confirmed from both in-vitro and in-vivo animal pilot studies, allowing it to be considered as a potential localized HSPs agent. Furthermore, the successful demonstration of a newly designed infusion technique, which diffuses the EMZF-SPNPAs through the vitreous body to the retina in a rat eye, more strongly verified the promises of this biotechnical approach to the ocular neuroprotection modality in glaucoma clinics.
Asunto(s)
Materiales Biocompatibles/química , Materiales Biocompatibles/uso terapéutico , Glaucoma/tratamiento farmacológico , Glaucoma/metabolismo , Proteínas de Choque Térmico/metabolismo , Magnetismo , Nanopartículas/química , Nanopartículas/uso terapéutico , Animales , Materiales Biocompatibles/efectos adversos , Línea Celular , Masculino , Microscopía Electrónica de Transmisión , Nanopartículas/efectos adversos , Nanopartículas/ultraestructura , Ratas , Ratas Sprague-Dawley , Células Ganglionares de la Retina/citología , Células Ganglionares de la Retina/efectos de los fármacos , Células Ganglionares de la Retina/metabolismo , Cuerpo Vítreo/metabolismoRESUMEN
The biochemical mechanisms of Chlorella vulgaris protection against cadmium (Cd)-induced liver toxicity were investigated in male Sprague-Dawley rats (5 weeks of age, weighing 90-110 g). Forty rats were randomly divided into one control and three groups treated with 10 ppm Cd: one Cd without Chlorella (Cd-0C), one Cd with 5% Chlorella (Cd-5C), and one Cd with 10% Chlorella (Cd-10C) groups. The rats had free access to water and diet for 8 weeks. Body weight gain and relative liver weight were significantly lower in the Cd-0C group than in Cd-5C and Cd-10C groups. Rats in the Cd-0C group had significantly higher hepatic concentrations of Cd and metallothioneins (MTs) than in the Cd-5C or Cd-10C group. The hepatic MT I/II mRNA was expressed in all experimental rats. MT II was more expressed in the Cd-5C and Cd-10C groups than in the Cd-0C group. Morphologically, a higher level of congestion and vacuolation was observed in the livers of the Cd-0C group compared to those of the Cd-5C and Cd-10C groups. Therefore, this study suggests that C. vulgaris has a protective effect against Cd-induced liver damage by reducing Cd accumulation and stimulating the expression of MT II in liver. However, the details of the mechanism of C. vulgaris on liver toxicity remains to be clarified by further studies.