Functional Improvement of Human Cardiotrophin 1 Produced in Tobacco Chloroplasts by Co-Expression with Plastid Thioredoxin m

Human cardiotrophin 1 (CT1), a cytokine with excellent therapeutic potential, was previously expressed in tobacco chloroplasts. However, the growth conditions required to reach the highest expression levels resulted in an impairment of its bioactivity. In the present study, we have examined new strategies to modulate the expression of this recombinant protein in chloroplasts so as to enhance its production and bioactivity. In particular, we assessed the effect of both the fusion and co-expression of Trx m with CT1 on the production of a functional CT1 by using plastid transformation. Our data revealed that the Trx m fusion strategy was useful to increase the expression levels of CT1 inside the chloroplasts, although CT1 bioactivity was significantly impaired, and this was likely due to steric hindrance between both proteins. By contrast, the expression of functional CT1 was increased when co-expressed with Trx m, because we demonstrated that recombinant CT1 was functionally active during an in vitro signaling assay. While Trx m/CT1 co-expression did not increase the amount of CT1 in young leaves, our results revealed an increase in CT1 protein stability as the leaves aged in this genotype, which also improved the recombinant protein’s overall production. This strategy might be useful to produce other functional biopharmaceuticals in chloroplasts

Functional Improvement of Human Cardiotrophin 1 Produced in Tobacco Chloroplasts by Co-Expression with Plastid Thioredoxin m

Human cardiotrophin 1 (CT1), a cytokine with excellent therapeutic potential, was previously expressed in tobacco chloroplasts. However, the growth conditions required to reach the highest expression levels resulted in an impairment of its bioactivity. In the present study, we have examined new strategies to modulate the expression of this recombinant protein in chloroplasts so as to enhance its production and bioactivity. In particular, we assessed the effect of both the fusion and co-expression of Trx m with CT1 on the production of a functional CT1 by using plastid transformation. Our data revealed that the Trx m fusion strategy was useful to increase the expression levels of CT1 inside the chloroplasts, although CT1 bioactivity was significantly impaired, and this was likely due to steric hindrance between both proteins. By contrast, the expression of functional CT1 was increased when co-expressed with Trx m, because we demonstrated that recombinant CT1 was functionally active during an in vitro signaling assay. While Trx m/CT1 co-expression did not increase the amount of CT1 in young leaves, our results revealed an increase in CT1 protein stability as the leaves aged in this genotype, which also improved the recombinant protein’s overall production. This strategy might be useful to produce other functional biopharmaceuticals in chloroplasts

Stable production of peptide antigens in transgenic tobacco chloroplasts by fusion to the p53 tetramerisation domain

7 p., 3 figures and bibliographyThe production of short peptides as single molecules in recombinant systems is often limited by the low stability of the foreign peptide. In the plant expression system this problem has been solved by translational fusions to recombinant proteins that are highly stable or are able to form complex structures. Previously, we demonstrated that the highly immunogenic 21 amino acid peptide 2L21, which is derived from the canine parvovirus (CPV) VP2 protein, did not accumulate in transgenic tobacco chloroplasts. In this report, we translationally fused the 2L21 peptide to the 42 amino acid tetramerisation domain (TD) from the human transcription factor p53. The chimaeric 2L21-TD protein was expressed in tobacco chloroplasts. Leaves accumulated high levels of the recombinant protein (up to 0.4 mg/g fresh weight of leaf material, equivalent to ~6% of total soluble protein; 2% considering only the 2L21 peptide). The 2L21-TD protein was able to form tetramers in the stroma of the chloroplast. Mice immunised intraperitoneally with partially purified leaf extracts containing the 2L21-TD protein developed specific antibodies with titres similar to those elicited by a previously reported fusion between 2L21 and the B subunit of the cholera toxin. Mouse sera were able to detect both the 2L21 synthetic peptide and the CPV VP2 protein, showing that the antigenicity of the 2L21 epitope was preserved in the chimaeric protein. These results demonstrate that the p53 TD can be used as a carrier molecule for the accumulation of short peptides (such as 2L21) in the chloroplast without altering the immunogenic properties of the peptide.This work was partially supported by Grant BIO2005-00155 from the Ministerio de Educación y Ciencia (Spain).Peer reviewe

Jornadas Nacionales de Robótica y Bioingeniería 2023: Libro de actas

Las Jornadas de Robótica y Bioingeniería de 2023 tienen lugar en la Escuela Técnica Superior de Ingeniería Industrial de la Universidad Politécnica de IVIadrid, entre los días 14 y 16 de junio de 2023. En este evento propiciado por el Comité Español de Automática (CEA) tiene lugar la celebración conjunta de las XII Jornadas Nacionales de Robótica y el XIV Simposio CEA de Bioingeniería. Las Jornadas Nacionales de Robótica es un evento promovido por el Grupo Temático de Robótica (GTRob) de CEA para dar visibilidad y mostrar las actividades desarrolladas en el ámbito de la investigación y transferencia tecnológica en robótica. Asimismo, el propósito de Simposio de Bioingeniería, que cumple ahora su decimocuarta dicción, es el de proporcionar un espacio de encuentro entre investigadores, desabolladores, personal clínico, alumnos, industriales, profesionales en general e incluso usuarios que realicen su actividad en el ámbito de la bioingeniería. Estos eventos se han celebrado de forma conjunta en la anualidad 2023. Esto ha permitido aunar y congregar un elevado número de participantes tanto de la temática robótica como de bioingeniería (investigadores, profesores, desabolladores y profesionales en general), que ha posibilitado establecer puntos de encuentro, sinergias y colaboraciones entre ambos. El programa de las jornadas aúna comunicaciones científicas de los últimos resultados de investigación obtenidos, por los grupos a nivel español más representativos dentro de la temática de robótica y bioingeniería, así como mesas redondas y conferencias en las que se debatirán los temas de mayor interés en la actualidad. En relación con las comunicaciones científicas presentadas al evento, se ha recibido un total de 46 ponencias, lo que sin duda alguna refleja el alto interés de la comunidad científica en las Jornadas de Robótica y Bioingeniería. Estos trabajos serán expuestos y presentados a lo largo de un total de 10 sesiones, distribuidas durante los diferentes días de las Jornadas. Las temáticas de los trabajos cubren los principales retos científicos relacionados con la robótica y la bioingeniería: robótica aérea, submarina, terrestre, percepción del entorno, manipulación, robótica social, robótica médica, teleoperación, procesamiento de señales biológicos, neurorehabilitación etc. Confiamos, y estamos seguros de ello, que el desarrollo de las jornadas sea completamente productivo no solo para los participantes en las Jornadas que podrán establecer nuevos lazos y relaciones fructíferas entre los diferentes grupos, sino también aquellos investigadores que no hayan podido asistir. Este documento que integra y recoge todas las comunicaciones científicas permitirá un análisis más detallado de cada una de las mismas

Contemporary use of cefazolin for MSSA infective endocarditis: analysis of a national prospective cohort

Objectives: This study aimed to assess the real use of cefazolin for methicillin-susceptible Staphylococcus aureus (MSSA) infective endocarditis (IE) in the Spanish National Endocarditis Database (GAMES) and to compare it with antistaphylococcal penicillin (ASP). Methods: Prospective cohort study with retrospective analysis of a cohort of MSSA IE treated with cloxacillin and/or cefazolin. Outcomes assessed were relapse; intra-hospital, overall, and endocarditis-related mortality; and adverse events. Risk of renal toxicity with each treatment was evaluated separately. Results: We included 631 IE episodes caused by MSSA treated with cloxacillin and/or cefazolin. Antibiotic treatment was cloxacillin, cefazolin, or both in 537 (85%), 57 (9%), and 37 (6%) episodes, respectively. Patients treated with cefazolin had significantly higher rates of comorbidities (median Charlson Index 7, P <0.01) and previous renal failure (57.9%, P <0.01). Patients treated with cloxacillin presented higher rates of septic shock (25%, P = 0.033) and new-onset or worsening renal failure (47.3%, P = 0.024) with significantly higher rates of in-hospital mortality (38.5%, P = 0.017). One-year IE-related mortality and rate of relapses were similar between treatment groups. None of the treatments were identified as risk or protective factors. Conclusion: Our results suggest that cefazolin is a valuable option for the treatment of MSSA IE, without differences in 1-year mortality or relapses compared with cloxacillin, and might be considered equally effective