ADN libre plasmático como marcador molecular en lesiones pre-neoplásicas de cuello uterino y su asociación con VPH

Introducción: La concentración de ADN libre en plasma ha sido investigada como un biomarcador tumoral en diferentes tipos de cáncer. Sin embargo, son pocos los estudios que evalúan la concentración ADN libre en pacientes con cáncer cervical y hasta la fecha no hay estudios en pacientes con lesiones pre-cancerosas cervicales. Objetivo: Establecer la asociación entre la concentración de ADN libre y el grado de la neoplasia cervical y evaluar su posible asociación con el tipo viral. Metodología: Estudio de prevalencia de tipo analítico con un muestreo no probabilístico. Se cuantifico el ADN libre en plasma por PCR en tiempo real de 92 pacientes que presentaban algún tipo de lesión intraepitelial cervical, confirmado por biopsia en diferentes instituciones de la ciudad de Bogotá. Adicional a esto se realizó la genotipificación del virus por Reverse Line Blot. Resultados: La concentración de ADN libre en plasma de pacientes con lesiones pre-cancerosas fue 4515 ± 16402 ng/ µl (media ± DS), LIEBG fue de 5188.7 ± 14876.5 ng/ µl (media ± DS), en pacientes con LIEAG fue de 830.3 ± 1515.508 ng/ µl (media ± DS), en pacientes con resultado negativo fue de 7024.7 ± 24107.5 ng/ µl (media ± DS). Los serotipos encontrados en la poblacion de estudio no presentaron asociacion con la concentracion de ADN libre. Discusión: Los resultados demostraron que la concentración absoluta de ADN libre en plasma no tiene un valor predictivo para diferenciar los tipos de lesión pre-neoplasica de cuello uterino, puesto que no se encontraron diferencias significativas en la concentración de ADN libre en plasma de las diferentes etapas progresivas de cáncer de cuello uterino (p;0.57, gl;3 alfa 0.05) de la misma forma el serotipo no contribuye a explicar la concentración de ADN libre.

Characterizing PvARP, a novel Plasmodium vivax antigen

Background Plasmodium vivax continues to be the most widely distributed malarial parasite species in tropical and sub-tropical areas, causing high morbidity indices around the world. Better understanding of the proteins used by the parasite during the invasion of red blood cells is required to obtain an effective vaccine against this disease. This study describes characterizing the P. vivax asparagine-rich protein (PvARP) and examines its antigenicity in natural infection. Methods The target gene in the study was selected according to a previous in silico analysis using profile hidden Markov models which identified P. vivax proteins that play a possible role in invasion. Transcription of the arp gene in the P. vivax VCG-1 strain was here evaluated by RT-PCR. Specific human antibodies against PvARP were used to confirm protein expression by Western blot as well as its subcellular localization by immunofluorescence. Recognition of recombinant PvARP by sera from P. vivax-infected individuals was evaluated by ELISA. Results VCG-1 strain PvARP is a 281-residue-long molecule, which is encoded by a single exon and has an N-terminal secretion signal, as well as a tandem repeat region. This protein is expressed in mature schizonts and is located on the surface of merozoites, having an apparent accumulation towards their apical pole. Sera from P. vivax-infected patients recognized the recombinant, thereby suggesting that this protein is targeted by the immune response during infection.

PvRON2, a new Plasmodium vivax rhoptry neck antigen

Background: Rhoptries are specialized organelles from parasites belonging to the phylum Apicomplexa; they secrete their protein content during invasion of host target cells and are sorted into discrete subcompartments within rhoptry neck or bulb. This distribution is associated with these proteins’ role in tight junction (TJ) and parasitophorous vacuole (PV) formation, respectively. Methods: Plasmodium falciparum RON2 amino acid sequence was used as bait for screening the codifying gene for the homologous protein in the Plasmodium vivax genome. Gene synteny, as well as identity and similarity values, were determined for ron2 and its flanking genes among P. falciparum, P. vivax and other malarial parasite genomes available at PlasmoDB and Sanger Institute databases. Pvron2 gene transcription was determined by RT-PCR of cDNA obtained from the P. vivax VCG-1 strain. Protein expression and localization were assessed by Western blot and immunofluorescence using polyclonal anti-PvRON2 antibodies. Co-localization was confirmed using antibodies directed towards specific microneme and rhoptry neck proteins. Results and discussion: The first P. vivax rhoptry neck protein (named here PvRON2) has been identified in this study. PvRON2 is a 2,204 residue-long protein encoded by a single 6,615 bp exon containing a hydrophobic signal sequence towards the amino-terminus, a transmembrane domain towards the carboxy-terminus and two coiled coil a-helical motifs; these are characteristic features of several previously described vaccine candidates against malaria. This protein also contains two tandem repeats within the interspecies variable sequence possibly involved in evading a host’s immune system. PvRON2 is expressed in late schizonts and localized in rhoptry necks similar to what has been reported for PfRON2, which suggests its participation during target cell invasion. Conclusions: The identification and partial characterization of the first P. vivax rhoptry neck protein are described in the present study. This protein is homologous to PfRON2 which has previously been shown to be associated with PfAMA-1, suggesting a similar role for PvRON2.

Identification, characterization and antigenicity of the Plasmodium vivax rhoptry neck protein 1 (PvRON1)

Background: Plasmodium vivax malaria remains a major health problem in tropical and sub-tropical regions worldwide. Several rhoptry proteins which are important for interaction with and/or invasion of red blood cells, such as PfRONs, Pf92, Pf38, Pf12 and Pf34, have been described during the last few years and are being considered as potential anti-malarial vaccine candidates. This study describes the identification and characterization of the P. vivax rhoptry neck protein 1 (PvRON1) and examine its antigenicity in natural P. vivax infections. Methods: The PvRON1 encoding gene, which is homologous to that encoding the P. falciparum apical sushi protein (ASP) according to the plasmoDB database, was selected as our study target. The pvron1 gene transcription was evaluated by RT-PCR using RNA obtained from the P. vivax VCG-1 strain. Two peptides derived from the deduced P. vivax Sal-I PvRON1 sequence were synthesized and inoculated in rabbits for obtaining anti-PvRON1 antibodies which were used to confirm the protein expression in VCG-1 strain schizonts along with its association with detergent-resistant microdomains (DRMs) by Western blot, and its localization by immunofluorescence assays. The antigenicity of the PvRON1 protein was assessed using human sera from individuals previously exposed to P. vivax malaria by ELISA. Results: In the P. vivax VCG-1 strain, RON1 is a 764 amino acid-long protein. In silico analysis has revealed that PvRON1 shares essential characteristics with different antigens involved in invasion, such as the presence of a secretory signal, a GPI-anchor sequence and a putative sushi domain. The PvRON1 protein is expressed in parasite's schizont stage, localized in rhoptry necks and it is associated with DRMs. Recombinant protein recognition by human sera indicates that this antigen can trigger an immune response during a natural infection with P. vivax. Conclusions: This study shows the identification and characterization of the P. vivax rhoptry neck protein 1 in the VCG-1 strain. Taking into account that PvRON1 shares several important characteristics with other Plasmodium antigens that play a functional role during RBC invasion and, as shown here, it is antigenic, it could be considered as a good vaccine candidate. Further studies aimed at assessing its immunogenicity and protection-inducing ability in the Aotus monkey model are thus recommended.

Prevalencia de enfermedad de Fabry en pacientes en lista de trasplante y posttrasplante renal en fundación cardioinfantil Bogotá

Introducción: La Enfermedad de Fabry (EF), es una enfermedad multisistémica de almacenamiento lisosomal ligada al cromosoma X que afecta principalmente a hombres, pero también puede causar significativa morbilidad en las mujeres heterocigotas (1–5). La deficiencia de la enzyma α-galactosidaseA (α-Gal A,) provoca acumulación de glicosfingolipidos que afectan diferentes tipos celulares entre ellos el endotelio vascular en vasos de pequeño calibre, células epiteliales y Músculo liso en el sistema cardiovascular (cardiomiocitos), sistema nervioso y células epiteliales tubulares del riñón (6,7). Complicaciones como la falla renal es la causa de muerte más frecuente en la EF (7,8). La incidencia se ha calculado en 1 de cada 117.000 nacidos vivos. (9). Objetivos: Determinar la prevalencia de la Enfermedad de Fabry en pacientes con Insuficiencia renal terminal que se encuentren en lista de trasplante y Post-trasplante Renal en Fundación Cardioinfantil Bogotá. Materiales y Métodos: Se realizó un estudio observacional en donde se evaluó la prevalencia de la EF en todos los sujetos mayores de 18 años que se encuentren en lista de trasplante y post-trasplante renal. Resultados: La prevalencia de Enfermedad de Fabry en 98 pacientes con enfermedad renal crónica fue de 7.1% para la muestra general y 12.9% para la muestra con etiología idiopática Conclusiones: La Enfermedad de Fabry es una importante casusa de Enfermedad Renal Crónica Terminal principalmente en el grupo de etiología idiopática. Palabras Clave: Enfermedad de Fabry (FA)