Recombinant protein production provoked accumulation of ATP, fructose-1,6-bisphosphate and pyruvate in E. coli K12 strain TG1

Background: Recently it was shown that production of recombinant proteins in E. coli BL21(DE3) using pET based expression vectors leads to metabolic stress comparable to a carbon overfeeding response. Opposite to original expectations generation of energy as well as catabolic provision of precursor metabolites were excluded as limiting factors for growth and protein production. On the contrary, accumulation of ATP and precursor metabolites revealed their ample formation but insufficient withdrawal as a result of protein production mediated constraints in anabolic pathways. Thus, not limitation but excess of energy and precursor metabolites were identified as being connected to the protein production associated metabolic burden. Results: Here we show that the protein production associated accumulation of energy and catabolic precursor metabolites is not unique to E. coli BL21(DE3) but also occurs in E. coli K12. Most notably, it was demonstrated that the IPTG-induced production of hFGF-2 using a tac-promoter based expression vector in the E. coli K12 strain TG1 was leading to persistent accumulation of key regulatory molecules such as ATP, fructose-1,6-bisphosphate and pyruvate. Conclusions: Excessive energy generation, respectively, accumulation of ATP during recombinant protein production is not unique to the BL21(DE3)/T7 promoter based expression system but also observed in the E. coli K12 strain TG1 using another promoter/vector combination. These findings confirm that energy is not a limiting factor for recombinant protein production. Moreover, the data also show that an accelerated glycolytic pathway flux aggravates the protein production associated “metabolic burden”. Under conditions of compromised anabolic capacities cells are not able to reorganize their metabolic enzyme repertoire as required for reduced carbon processing. © 2021, The Author(s)

Attitudes Toward Assisted Suicide and Life-Prolonging Measures in Swiss ALS Patients and Their Caregivers

Objectives: In Switzerland, assisted suicide (AS) is legal, provided that the person seeking assistance has decisional capacity and the person assisting is not motivated by reasons of self-interest. However, in this particular setting nothing is known about patients' and their caregivers' attitudes toward AS and life-prolonging measures. Methods: Data was retrieved through validated questionnaires and personal interviews in 33 patients and their caregivers covering the following domains: physical function according to the revised Amyotrophic Lateral Sclerosis Functional Rating Scale (ALSFRS-R), demographic data, quality of life, anxiety, depression, social situation, spirituality, burden of disease, life-prolonging, and life-shortening acts. Results: In patients the median time after diagnosis was 9 months (2-90) and the median Amyotrophic Lateral Sclerosis (ALS) FRS-R score was 37 (22-48). The majority of patients (94%; n = 31) had no desire to hasten death. Patients' and caregivers' attitudes toward Percutaneous Endoscopic Gastrostomy (PEG) and Non-Invasive Ventilation (NIV) differed. Significantly more patients than caregivers (21.2 versus 3.1%) stated that they were against NIV (p = 0.049) and against PEG (27.3 versus 3.1%; p = 0.031). Answers regarding tracheotomy were not significantly different (p = 0.139). Caregivers scored significantly higher levels of "suffering" (p = 0.007), "loneliness" (p = 0.006), and "emotional distress" answering the questionnaires (p > 0.001). Suffering (p > 0.026) and loneliness (p > 0.016) were related to the score of the Hospital Anxiety and Depression Scale (HADS) in patients. Conclusion: A liberal legal setting does not necessarily promote the wish for AS. However, the desire to discuss AS is prevalent in ALS patients. There is a higher level of suffering and loneliness on the caregivers' side. A longitudinal study is warranted

Wissensstandsanalyse zu Qualität, Verbraucherschutz und Verarbeitung ökologischer Lebensmittel

Zielsetzung des Projektes war es, den aktuellen Stand des Wissens zur Qualität und Verarbeitung ökologischer Lebensmittel sowie zum Verbraucherschutz aufzuzeigen und zu bewerten. Das Projektteam setzte sich aus Wissenschaftlern des Forschungsinstituts für biologischen Landbau (Deutschland und Schweiz), des Büros für Lebensmittelkunde, des Forschungsrings für Biologisch-Dynamische Wirtschaftsweise e.V. sowie des Fachgebiets Ökologische Lebensmittelqualität und Ernährungskultur der Universität Kassel zusammen. In dem zeitlich sehr begrenzten Projekt fokussierte die Arbeitsgruppe die Betrachtung auf folgende Themenkomplexe und Produktgruppen: Bereich Qualität - Ernährung - Sensorik - Ökospezifische Qualitäten - Authentizität und Rückverfolgbarkeit Bereich Verarbeitung - Rohwaren/Lagerung und Technologien für die Produktgruppen: -- Getreide -- Milch -- Fleisch -- Obst und Gemüse und Erzeugnisse aus diesen - Nachhaltigkeit im Unternehmen und entlang der Prozesskette - Verpackung Vorhandenes Wissen, aktuelle Fragestellungen und Entwicklungsrichtungen wurden nach thematisch angepassten Vorgehensweisen identifiziert und anhand internationaler Literatur und Experteninterviews diskutiert. Zu allen Themenkomplexen wurden kurze und allgemeinverständliche Reports erstellt, die einen schnellen und fundierten Überblick zum aktuellen Stand des Wissens und zu eventuellen Wissenslücken geben. Der Abschlussreport wurde in deutscher und englischer Sprache veröffentlicht. Insgesamt wurde festgestellt, dass in den betrachteten Themenkomplexen erheblicher Bedarf an Forschung und Entwicklung sowie an Wissenstransfer besteht. Neben einer Reihe von Detailfragen sind grundlegende Themen, wie beispielsweise das Anforderungsprofil an eine „ökologische Verarbeitung“, nicht hinreichend erforscht und geklärt. Diese Arbeiten sind jedoch Voraussetzung für die Bearbeitung von Detailthemen, da hier Aufgabenstellungen, Schwerpunktsetzungen und Methoden definiert werden

Renal developmental genes are differentially regulated after unilateral ureteral obstruction in neonatal and adult mice

Congenital obstructive nephropathy hinders normal kidney development. The severity and the duration of obstruction determine the compensatory growth of the contralateral, intact opposite kidney. We investigated the regulation of renal developmental genes, that are relevant in congenital anomalies of the kidney and urinary tract (CAKUT) in obstructed and contralateral (intact opposite) kidneys after unilateral ureteral obstruction (UUO) in neonatal and adult mice. Newborn and adult mice were subjected to complete UUO or sham-operation, and were sacrificed 1, 5, 12 and 19 days later. Quantitative RT-PCR was performed in obstructed, intact opposite kidneys and sham controls for Gdnf, Pax2, Six4, Six2, Dach1, Eya1, Bmp4, and Hnf-1 beta. Neonatal UUO induced an early and strong upregulation of all genes. In contrast, adult UUO kidneys showed a delayed and less pronounced upregulation. Intact opposite kidneys of neonatal mice revealed a strong upregulation of all developmental genes, whereas intact opposite kidneys of adult mice demonstrated only a weak response. Only neonatal mice exhibited an increase in BMP4 protein expression whereas adult kidneys strongly upregulated phosphatidylinositol 3 kinase class III, essential for compensatory hypertrophy. In conclusion, gene regulation differs in neonatal and adult mice with UUO. Repair and compensatory hypertrophy involve different genetic programs in developing and adult obstructed kidneys

Aspekte von Verpackungsmaterialien bei Ökoprodukten

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Personality traits and physical complaints in patients with acromegaly : a cross sectional multi-center study with analysis of influencing factors

Objective: Acromegalic patients display a distinct neuropsychological profile and suffer from chronic physical complaints. We aimed to investigate in more detail these aspects in acromegalic patients, dependent on influencing factors like disease activity, age, sex, chronic medication, surgery, pituitary radiation, pituitary insufficiency and comorbidities. Design: Cross sectional, multicentric. Methods: 129 patients (M/W 65/64, 58.3±12.7 ys., 53/76 with active/controlled disease). Acromegalic patients completed the following inventories: NEO-FFI, IIP-D, and the Giessen Complaints List (GBB-24), after written informed consent. Age, sex, IGF-1 concentrations, comorbidities, treatment modalities and pituitary insufficiency were documented. Results: Acromegalic patients or specific patient-subgroups were more agreeable, neurotic, exploitable/permissive, introverted/socially avoidant, non-assertive/insecure, nurturant and less open to experience, cold/denying, domineering, compared to normal values from the healthy population (controls). Multivariable analysis demonstrated that these overall results were due to the specific patient subgroups as patients on chronic medication, with arthrosis and pituitary insufficiency. Disease activity was only associated with the trait nurturant. Higher scores for introversion were associated with arthrosis. Lower domineering was independent of any disease- or treatment related variable or comorbidity. The GBB inventory showed overall higher scores in patients, with higher scores for exhaustion and general complaints being associated with pituitary insufficiency, coronary heart disease and history of malignancy in the multivariable analysis. Joint complaints were independent of any disease- or treatment- related variable. Conclusions: We define new aspects of a distinct neuropsychological profile in patients with acromegaly, which are largely independent of disease activity. Chronic physical complaints are more pronounced in patients than in controls, with exhaustion and general complaints showing no association with disease activity