58 research outputs found
Understanding the effect of increased cell specific productivity on galactosylation of monoclonal antibodies produced using Chinese hamster ovary cells
No full textPhysiological Assessment of ECCO<sub>2</sub>R on the Work of Breathing in Exacerbations of COPD
No full textRationaleThe impact of extracorporeal carbon dioxide removal (ECCO2R) on work of breathing and aeration in exacerbations of chronic obstructive pulmonary disease (AECOPD) is poorly understood. ObjectivesThe study explores the impact of non-invasive ventilation (NIV) and ECCO2R on respiratory drive, effort and distribution of ventilation in AECOPD. MethodsPatients enrolled in a randomised controlled study of the addition of ECCO2R to NIV compared with NIV underwent oesophageal pressure measurement, electrical impedance tomography and parasternal electromyography. Measurements and main results18 patients were enrolled, nine in each arm. Of these, eight in the NIV arm and seven in the ECCO2R arm underwent physiological assessment. Patients randomised to ECCO2R, had a period of NIV combined with ECCO2R and, after removal of NIV, a period of ECCO2R alone. The removal of NIV whilst remaining on ECCO2R resulted in a respiratory acidosis (pH 7.34 (7.31–7.34) vs. 7.31 (7.31–7.34), p < 0.0001), increased work of breathing (7.43 (6.08–10.19) vs. 11.1 (8.11–15.15) J/min, p < 0.0001) and increased neural drive (884.4 (684.7–967.3) vs. 1321.1 (903.3–1575.3) AU, p = 0.0005). On day 1, the work of breathing was lower in the NIV than the ECCO2R group (4.38 (2.76–7.27) vs. 8.03 (4.8–15.94) J/min, p < 0.0001), minute ventilation was higher (15.54 (13.14–18.48) vs. 12.24 (8.51–13.9) L/min, p < 0.0001) and neural drive was the same (1,163.8 (1,085.5–1,325.5) vs. 1,093.8 (885.7–1,258.7) AU, p = 0.5556). ConclusionsThe combination of NIV and ECCO2R results in lower work of breathing and improved neuro-ventilatory coupling. NIV fully supports ventilation early whilst ECCO2R improves neuro-ventilatory coupling and work of breathing over time. Trial registrationClinicaltrials.gov; NCT02086084; registered 1 December 2015; https://clinicaltrials.gov/study/NCT02086084?cond=copd&term=ecco2r&rank=4.</p
Two related neurokinin-1 receptor antagonists have overlapping but different binding sites
No full textMolecular Characterization of the Melanin-Concentrating Hormone/Receptor Complex: Identification of Critical Residues Involved in Binding and Activation
No full textImproving Antibody Production in Stably Transfected CHO Cells by CRISPR‐Cas9‐Mediated Inactivation of Genes Identified in a Large‐Scale Screen with Chinese Hamster‐Specific siRNAs
No full textEngineering the substrate specificity of glutathione reductase toward that of trypanothione reduction
No full textGlutathione reductase (EC 1.6.4.2; CAS registry number 9001-48-3) and trypanothione reductase (CAS registry number 102210-35-5), which are related flavoprotein disulfide oxidoreductases, have marked specificities for glutathione and trypanothione, respectively. A combination of primary sequence alignments and molecular modeling, together with the high-resolution crystal structure of human glutathione reductase, identified certain residues as potentially being responsible for substrate discrimination. Site-directed mutagenesis of Escherichia coli glutathione reductase was used to test these predictions. The mutation of Asn-21 to Arg demonstrated that this single change was insufficient to generate the greater discrimination against trypanothione shown by human glutathione reductase compared with the E. coli enzyme. However, the mutation of Ala-18, Asn-21, and Arg-22 to the amino acid residues (Glu, Trp, and Asn, respectively) in corresponding positions in Trypanosoma congolense trypanothione reductase confirmed that this region of polypeptide chain is intimately involved in substrate recognition. It led to a mutant form of E. coli glutathione reductase that possessed essentially no activity with glutathione but that was able to catalyze trypanothione reduction with a kcat/Km value that was 10% of that measured for natural trypanothione reductases. These results should be of considerable importance in the design of trypanocidal drugs targeted at the differences between glutathione and trypanothione metabolism in trypanosomatids and their hosts
Systems biology approach in the development of chemically-defined media for production of protein therapeutics in Chinese hamster ovary cells
No full textCell culture medium plays a critical role on mammalian cell growth, protein expression and quality. Typical cell culture medium formulations consist of \u3e50 components which include amino acids, vitamins, trace metals, lipids and proteins. Chinese Hamster Ovary (CHO) cells that produce biotherapeutics are propagated in specific cell culture media to ensure robust productivity and product quality.
Systems biology has been applied to multiple areas of biological research to gain a better understanding of disease origins and to identify potential new drug targets. Although CHO cells are simpler systems, they share similar biochemistry and cellular pathways. Therefore, leveraging the systems biology knowledge from animal systems and applying these strategic systems biological tools to bioprocess development can be valuable in gaining better understanding of CHO cell culture performance, optimizing cell culture media, and subsequently resulting in better control of the overall production processes.
In this presentation, we will present several case studies of various ‘omics tools applied to (1) optimize cell culture medium formulation for improve cell growth and productivity via metabolomics, (2) understand effects of medium components on cellular gene expression via transcriptomics, and on product quality via glycomics, and (3) identify potential cellular protein targets that are affected by stress imposed during production process via proteomics. The development of a statistical model that aims to highlight key metabolites and a machine learning model that identifies significantly important genes which are involved in monoclonal antibody production will also be discussed
Il borgo alpino di Contrada Bianchi in Valmalenco. Fragilità, inneschi e scenari di riuso
No full textLAUREA MAGISTRALEIl lavoro di tesi assume come punto di partenza uno studio avviato nel 2007 presso l’allora Dipartimento di Ingegneria Civile e ambientale del Politecnico di Milano in collaborazione con la Comunità Montana della Valmalenco, avente come tema lo studio della vulnerabilità del costruito storico diffuso. Da tale studio iniziava ad emergere uno sguardo multidisciplinare attento a far cogliere le dinamiche complesse che hanno contribuito a generare il fenomeno dell’abbandono dei piccoli borghi storici in Valmalenco.
Un contributo significativo per la stesura del lavoro di tesi, avente come tema la valutazione degli scenari di rischio alle differenti scale (da quella territoriale a quella costruttiva) è rappresentato dalla Tesi di Laurea Magistrale “Borghi dismessi in area alpina : scenari di riuso. Il caso studio di Torre di Santa Maria, Contrada Bianchi (SO)” discussa nel 2013 dal Dott. Arch. Lorenzo Murgolo presso la Scuola di Architettura Civile del Politecnico di Milano. Un più recente incontro con l’autore ha consentito di procedere con una analisi delle trasformazioni in atto dal 2009 fino ad oggi, cercando di cogliere le ricadute di alcune scelte politiche, economiche, sociali, ambientali che diverse Amministrazioni Pubbliche ed enti preposti alla tutela del territorio hanno perseguito.
A partire da tali premesse, il lavoro di tesi indaga la realtà contemporanea di Contrada Bianchi in Valmalenco con l’obiettivo di (ri)percorrere e valutare criticamente i fenomeni che hanno caratterizzato il borgo negli ultimi dieci anni e valutare l’opportunità di un ripensamento delle politiche e delle modalità di intervento considerando i recenti eventi, al fine di far emergere le risorse potenziali inespresse ma insite del territorio, nel borgo, negli edifici.
Di forte rilevanza risulta essere il dialogo con enti, associazioni ed operatori che agiscono sul territorio della Valmalenco (stakeholder). Gli incontri preliminari al progetto hanno fatto emergere la possibilità di sviluppare occasioni di incontri partecipativi finalizzati al far emergere: in fase iniziale i bisogni e le risorse, in fase metaprogettuale le potenzialità dei luoghi e gli scenari di riuso, in fase progettuale la condivisione delle soluzioni architettoniche a partire da (ri)costruzioni in sito.The thesis takes as a starting point a study launched in 2007 at the then Department of Civil and Environmental Engineering of the Polytechnic of Milan in collaboration with the Comunità Montana della Valmalenco, whose theme was the study of the vulnerability of the widespread historical building. From this study a multidisciplinary gaze began to emerge, careful to capture the complex dynamics that contributed to generating the phenomenon of the abandonment of the small historic villages in Valmalenco.
A significant contribution to the drafting of the thesis work, whose theme is the evaluation of risk scenarios at different scales (from the territorial to the constructive one) is represented by the Master's Degree Thesis “Disused villages in the Alpine area: scenarios of reuse. The case study of Torre di Santa Maria, Contrada Bianchi (SO) "discussed in 2013 by Dr. Arch. Lorenzo Murgolo at the School of Civil Architecture of the Politecnico di Milano. A more recent meeting with the author has made it possible to proceed with an analysis of the transformations taking place from 2009 until today, trying to grasp the repercussions of some political, economic, social and environmental choices that various Public Administrations and bodies in charge of protecting territory they pursued.
Starting from these premises, the thesis work investigates the contemporary reality of Contrada Bianchi in Valmalenco with the aim of (re) tracing and critically evaluating the phenomena that have characterized the village in the last ten years and evaluating the opportunity for a rethinking policies and methods of intervention considering recent events, in order to bring out the potential unexpressed but inherent resources of the territory, in the village, in the buildings.
The dialogue with bodies, associations and operators acting on the Valmalenco territory (stakeholders) is of great importance. The preliminary meetings of the project revealed the possibility of developing opportunities for participatory meetings aimed at bringing out: in the initial phase the needs and resources, in the metaprojective phase the potentialities of the places and the scenarios of reuse, in the design phase the sharing of solutions architectural from (re) construction on sit
Engineering the substrate specificity of glutathione reductase toward that of trypanothione reduction
No full textGlutathione reductase (EC 1.6.4.2; CAS registry number 9001-48-3) and trypanothione reductase (CAS registry number 102210-35-5), which are related flavoprotein disulfide oxidoreductases, have marked specificities for glutathione and trypanothione, respectively. A combination of primary sequence alignments and molecular modeling, together with the high-resolution crystal structure of human glutathione reductase, identified certain residues as potentially being responsible for substrate discrimination. Site-directed mutagenesis of Escherichia coli glutathione reductase was used to test these predictions. The mutation of Asn-21 to Arg demonstrated that this single change was insufficient to generate the greater discrimination against trypanothione shown by human glutathione reductase compared with the E. coli enzyme. However, the mutation of Ala-18, Asn-21, and Arg-22 to the amino acid residues (Glu, Trp, and Asn, respectively) in corresponding positions in Trypanosoma congolense trypanothione reductase confirmed that this region of polypeptide chain is intimately involved in substrate recognition. It led to a mutant form of E. coli glutathione reductase that possessed essentially no activity with glutathione but that was able to catalyze trypanothione reduction with a kcat/Km value that was 10% of that measured for natural trypanothione reductases. These results should be of considerable importance in the design of trypanocidal drugs targeted at the differences between glutathione and trypanothione metabolism in trypanosomatids and their hosts
Mapping and characterization of the epitope(s) of Sch 55700, a humanized mAb, that inhibits human IL-5
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