Amino acid sequences of proteins from Leptospira serovar pomona

This report describes a partial amino acid sequences from three putative outer envelope proteins from Leptospira serovar pomona. In order to obtain internal fragments for protein sequencing, enzymatic and chemical digestion was performed. The enzyme clostripain was used to digest the proteins 32 and 45 kDa. In situ digestion of 40 kDa molecular weight protein was accomplished using cyanogen bromide. The 32 kDa protein generated two fragments, one of 21 kDa and another of 10 kDa that yielded five residues. A fragment of 24 kDa that yielded nineteen residues of amino acids was obtained from 45 kDa protein. A fragment with a molecular weight of 20 kDa, yielding a twenty amino acids sequence from the 40 kDa protein.

Acidic ribosomal proteins and histone H3 from Leishmania present a high rate of divergence

Another additional peculiarity in Leishmania will be discussed about of the amino acid divergence rate of three structural proteins: acidic ribosomal P1 and P2b proteins, and histone H3 by using multiple sequence alignment and dendrograms. These structural proteins present a high rate of divergence regarding to their homologous protein in Trypanosoma cruzi. At this regard, L. (V.) peruviana P1 and T. cruzi P1 showed 57.4% of divergence rate. Likewise, L. (V.) braziliensis histone H3 and acidic ribosomal P2 protein exhibited 31.8% and 41.7% respectively of rate of divergence in comparison with their homologous in T. cruzi.

Hepatitis C virus proteins promote mitochondrial bioenergetic dysfunction and nitro-oxidative stress: insights into pathogenesis

HCV-infection induces a state of oxidative stress more pronounced than in many other inflammatory diseases. Here we propose a temporal sequence of events in the HCV-infected cell whereby the primary alteration consists in release of Ca2+ from the ER followed by uptake into mitochondria. This triggers successive mitochondrial dysfunctions leading to generation of ROS and to a progressive metabolic adaptive response. Pathogenetic implications of the model and new opportunities for therapeutic intervention are discussed.

The yellow fever 17D vaccine virus as a vector for the expression of foreign proteins: development of new live flavivirus vaccines

The Flaviviridae is a family of about 70 mostly arthropod-borne viruses many of which are major public health problems with members being present in most continents. Among the most important are yellow fever (YF), dengue with its four serotypes and Japanese encephalitis virus. A live attenuated virus is used as a cost effective, safe and efficacious vaccine against YF but no other live flavivirus vaccines have been licensed. The rise of recombinant DNA technology and its application to study flavivirus genome structure and expression has opened new possibilities for flavivirus vaccine development. One new approach is the use of cDNAs encopassing the whole viral genome to generate infectious RNA after in vitro transcription. This methodology allows the genetic mapping of specific viral functions and the design of viral mutants with considerable potential as new live attenuated viruses. The use of infectious cDNA as a carrier for heterologous antigens is gaining importance as chimeric viruses are shown to be viable, immunogenic and less virulent as compared to the parental viruses. The use of DNA to overcome mutation rates intrinsic of RNA virus populations in conjunction with vaccine production in cell culture should improve the reliability and lower the cost for production of live attenuated vaccines. The YF virus despite a long period ignored by researchers probably due to the effectiveness of the vaccine has made a come back, both in nature as human populations grow and reach endemic areas as well as in the laboratory being a suitable model to understand the biology of flaviviruses in general and providing new alternatives for vaccine development through the use of the 17D vaccine strain.

High throughput sequencing and proteomics to identify immunogenic proteins of a new pathogen: the dirty genome approach.

With the availability of new generation sequencing technologies, bacterial genome projects have undergone a major boost. Still, chromosome completion needs a costly and time-consuming gap closure, especially when containing highly repetitive elements. However, incomplete genome data may be sufficiently informative to derive the pursued information. For emerging pathogens, i.e. newly identified pathogens, lack of release of genome data during gap closure stage is clearly medically counterproductive. We thus investigated the feasibility of a dirty genome approach, i.e. the release of unfinished genome sequences to develop serological diagnostic tools. We showed that almost the whole genome sequence of the emerging pathogen Parachlamydia acanthamoebae was retrieved even with relatively short reads from Genome Sequencer 20 and Solexa. The bacterial proteome was analyzed to select immunogenic proteins, which were then expressed and used to elaborate the first steps of an ELISA. This work constitutes the proof of principle for a dirty genome approach, i.e. the use of unfinished genome sequences of pathogenic bacteria, coupled with proteomics to rapidly identify new immunogenic proteins useful to develop in the future specific diagnostic tests such as ELISA, immunohistochemistry and direct antigen detection. Although applied here to an emerging pathogen, this combined dirty genome sequencing/proteomic approach may be used for any pathogen for which better diagnostics are needed. These genome sequences may also be very useful to develop DNA based diagnostic tests. All these diagnostic tools will allow further evaluations of the pathogenic potential of this obligate intracellular bacterium.

Anemia en la enfermedad inflamatoria intestinal: Prevalencia y factores que influyen.

La anemia es una complicación muy importante de la enfermedad inflamatoria intestinal (EII). Actualmente desconocemos la prevalencia de anemia en los pacientes afectos por dicha enfermedad. Para ello se ha realizado un estudio retrospectivo de revisión del historial clínico de pacientes con EII registrados en la Unidad de Atención Crohn-Colitis, con el objetivo de determinar la prevalencia de anemia en nuestro medio y analizar los factores que influyen en la aparición de la misma. La prevalencia de anemia en la EII en el año 2007 fue del 26,3%. El factor más importante para desarrollarla fue la presencia de actividad clínica.

A novel role for proline- and acid-rich basic region leucine zipper (PAR bZIP) proteins in the transcriptional regulation of a BH3-only proapoptotic gene.

Proline- and acid-rich (PAR) basic region leucine zipper (bZIP) proteins thyrotroph embryonic factor (TEF), D-site-binding protein (DBP), and hepatic leukemia factor have been involved in neurotransmitter homeostasis and amino acid metabolism. Here we demonstrate a novel role for these proteins in the transcriptional control of a BH3-only gene. PAR bZIP proteins are able to transactivate the promoter of bcl-gS. This promoter is particularly responsive to TEF activation and is silenced by NFIL3, a repressor that shares the consensus binding site with PAR bZIP proteins. Consistently, transfection of TEF induces the expression of endogenous bcl-gS in cancer cells, and this induction is independent of p53. A naturally occurring variant of DBP (tDBP), lacking the transactivation domain, has been identified and shown to impede the formation of active TEF dimers in a competitive manner and to reduce the TEF-dependent induction of bcl-gS. Of note, treatment of cancer cells with etoposide induces TEF activation and promotes the expression of bcl-gS. Furthermore, blockade of bcl-gS or TEF expression by a small interfering RNA strategy or transfection with tDBP significantly reduces the etoposide-mediated apoptotic cell death. These findings represent the first described role for PAR bZIP proteins in the regulation of a gene involved in the execution of apoptosis.

Anemia en pacientes de edad avanzada con insuficiencia cardíaca ingresados en un servicio de medicina interna

Introducción: la anemia es una entidad muy prevalente en pacientes con insuficiencia cardíaca (IC) y se asocia a mayor morbimortalidad. Objetivos: analizar las comorbilidades de pacientes hospitalizados de ≥ 65 años con IC. Estudiar el impacto de la anemia sobre esta población y comparar esta comorbilidad respecto a aquéllos que no la presentan en relación a la valoración geriátrica integral (VGI), así como la adecuación del tratamiento farmacológico en la IC. Pacientes y métodos: estudio descriptivo observacional transversal de una cohorte de 150 pacientes ingresados en el Servicio de Medicina Interna del Hospital Vall d’Hebron, entre junio de 2007 y enero de 2010, mediante entrevista clínica y recogida de datos de la historia clínica. Resultados: en la muestra prevalecían las mujeres (62%), los pacientes con hipertensión (84%), los que tenían la FEVI conservada (66,4%) y los pacientes con anemia (61,3%), de los cuales el 60,9% presentaban anemia inflamatoria y el 35,9% anemia ferropénica. Los pacientes con anemia tuvieron peor valor de MNA (p=0,017, con un RR de 2,7), mayor diferencial de Barthel (p=0,021) y peor valor de albuminemia (p=0,001). Asimismo, se observó que 53 pacientes con indicación según las guías clínicas de tratamiento con IECA o ARA II no lo seguían, y hasta 105 pacientes en el caso de los betabloqueantes (BB). Conclusiones: los pacientes con anemia presentaban pero estado nutricional, y mayor empeoramiento del índice de Barthel. Respecto al tratamiento de la IC, destacaba el gran número de pacientes sin tratamiento con IECA, ARA II o BB, que deberían llevarlo según las guías de práctica clínica.

Differential distribution of two microtubule-associated proteins, MAP2 and MAP5, during chick dorsal root ganglion development in situ and in culture.

Microtubule-associated proteins (MAPs) are essential components necessary for the early growth process of axons and dendrites, and for the structural organization within cells. Both MAP2 and MAP5 are involved in these events, MAP2 occupying a role predominantly in dendrites, and MAP5 being involved in both axonal and dendritic growth. In the chick dorsal root ganglia, pseudo-unipolar sensory neurons have a T-shaped axon and are devoid of any dendrites. Therefore, they offer an ideal model to study the differential expression of MAPs during DRG development, specifically during axonal growth. In this study we have analyzed the expression and localization of MAP2 and MAP5 isoforms during chick dorsal root ganglia development in vivo, and in cell culture. In DRG, both MAPs appeared as early as E5. MAP2 consists of the 3 isoforms MAP2a, b and c. On blots, no MAP2a could be found at any stage. MAP2b increased between E6 and E10 and thereafter diminished slowly in concentration, while MAP2c was found between stages E6 and E10 in DRG. By immunocytochemistry, MAP2 isoforms were mainly located in the neuronal perikarya and in the proximal portion of axons, but could not be localized to distal axonal segments, nor in sciatic nerve at any developmental stage. On blots, MAP5 was present in two isoforms, MAP5a and MAP5b. The concentration of MAP5a was highest at E6 and then decreased to a low level at E18. In contrast, MAP5b increased between E6 and E10, and rapidly decreased after E14. Only MAP5a was present in sciatic nerve up to E14. Immunocytochemistry revealed that MAP5 was localized mainly in axons, although neuronal perikarya exhibited a faint immunostaining. Strong staining of axons was observed between E10 and E14, at a time coincidental to a period of intense axonal outgrowth. After E14 immunolabeling of MAP5 decreased abruptly. In DRG culture, MAP2 was found exclusively in the neuronal perikarya and the most proximal neurite segment. In contrast, MAP5 was detected in the neuronal cell bodies and all along their neurites. In conclusion, MAP2 seems involved in the early establishment of the cytoarchitecture of cell bodies and the proximal axon segment of somatosensory neurons, while MAP5 is clearly related to axonal growth.

Estudio retrospectivo de la influencia de la respuesta al tratamiento quimioterápico de primera línea en pacientes con cáncer no microcítico de pulmón sobre la respuesta a la administración de factores estimulantes de la eritropoyesis en la anemia inducida por quimioterapia

S’analitza de manera retrospectiva la relació entre la resposta a la quimioteràpia i la milloria de l’anèmia induïda per la quimioteràpia amb factors estimulants de la eritropoesi en 57 pacients amb carcinoma no microcític de pulmó avançat que presentaren anèmia durant el tractament amb quimioteràpia. Els pacients amb progressió desenvoluparen anèmia significativament abans que els responedors y presentaren de forma significativa una menor tassa de resposta al tractament amb factors estimulants de la eritropoesi. La resposta a la quimioteràpia té un impacte directe en la efectivitat del tractament de la anèmia amb factors estimulant de la eritropoesi. Aquesta correlació pot valdre com marcador subrogat de resposta a la quimioteràpia. Paraules clau: Càncer no microcític de pulmó, anèmia induïda per quimioteràpia, factors estimulants de la eritropoesi, factor predictor de resposta

The E.coli RuvAB proteins branch migrate Holliday junctions through heterologous DNA sequences in a reaction facilitated by SSB.

During genetic recombination a heteroduplex joint is formed between two homologous DNA molecules. The heteroduplex joint plays an important role in recombination since it accommodates sequence heterogeneities (mismatches, insertions or deletions) that lead to genetic variation. Two Escherichia coli proteins, RuvA and RuvB, promote the formation of heteroduplex DNA by catalysing the branch migration of crossovers, or Holliday junctions, which link recombining chromosomes. We show that RuvA and RuvB can promote branch migration through 1800 bp of heterologous DNA, in a reaction facilitated by the presence of E.coli single-stranded DNA binding (SSB) protein. Reaction intermediates, containing unpaired heteroduplex regions bound by SSB, were directly visualized by electron microscopy. In the absence of SSB, or when SSB was replaced by a single-strand binding protein from bacteriophage T4 (gene 32 protein), only limited heterologous branch migration was observed. These results show that the RuvAB proteins, which are induced as part of the SOS response to DNA damage, allow genetic recombination and the recombinational repair of DNA to occur in the presence of extensive lengths of heterology.

Comparative immunological recognition of proteins from New Guinea "C" dengue virus type 2 prototype and from a dengue virus type 2 strain isolated in the State of Rio de Janeiro, Brazil

The protein profiles of the New Guinea "C" dengue virus type 2 (DENV-2)prototype and those of a Brazilian DENV-2 isolated in the State of Rio de Janeiro in 1995 were compared. SDS-PAGE analysis showed that the virus from Rio de Janeiro expresses NS5 (93.0 kDa), NS3 (66.8 kDa) E (62.4 kDa) and NS1 (41.2 kDa) proteins differently from the New Guinea "C" virus. The immunoblot revealed specificity and antigenicity for the NS3 protein from DENV-2 Rio de Janeiro mainly in primary infections, convalescent cases, and in secondary infections in both cases and only antigenicity for E and NS1 proteins for both viruses in primary and secondary infections.