Search

To evaluate the prevalence and clinical correlates of lifetime migraine among patients with bipolar disorder (BD).In a cross-sectional study, we evaluated 721 adults with BD from the Mayo Clinic Bipolar Disorder Biobank and compared clinical correlates of those with and without a lifetime history of migraine. A structured clinical interview (DSM-IV) and a clinician-assessed questionnaire were utilized to establish a BD diagnosis, lifetime history of migraine, and clinical correlates.Two hundred and seven (29%) BD patients had a lifetime history of migraine. BD patients with migraine were younger and more likely to be female as compared to those without migraine (p values0.01). In a multivariate logistic regression model, younger age (OR=0.98, p0.01), female sex (OR=2.02, p0.01), higher shape/weight concern (OR=1.04, p=0.02), greater anxiety disorder comorbidities (OR=1.24, p0.01), and evening chronotype (OR=1.65, p=0.03) were associated with migraine. In separate regression models for each general medical comorbidity (controlled for age, sex, and site), migraines were significantly associated with fibromyalgia (OR=3.17, p0.01), psoriasis (OR=2.65, p=0.03), and asthma (OR=2.0, p0.01). Participants with migraine were receiving ADHD medication (OR=1.53, p=0.05) or compounds associated with weight loss (OR=1.53, p=0.02) at higher rates compared to those without migraine.Study design precludes determination of causality. Migraine subtypes and features were not assessed.Migraine prevalence is high in BD and is associated with a more severe clinical burden that includes increased comorbidity with pain and inflammatory conditions. Further study of the BD-migraine phenotype may provide insight into common underlying neurobiological mechanisms.

Real-time, detailed online information on cell cultures is essential for understanding modern biopharmaceutical production processes. The determination of key parameters, such as cell density and viability, is usually based on the offline sampling of bioreactors. Gathering offline samples is invasive, has a low time resolution, and risks altering or contaminating the production process. In contrast, measuring process parameters online provides more safety for the process, has a high time resolution, and thus can aid in timely process control actions. We used online double differential digital holographic microscopy (D3HM) and machine learning to perform non-invasive online cell concentration and viability monitoring of insect cell cultures in bioreactors. The performance of D3HM and the machine learning model was tested for a selected variety of baculovirus constructs, products, and multiplicities of infection (MOI). The results show that with online holographic microscopy insect cell proliferation and baculovirus infection can be monitored effectively in real time with high resolution for a broad range of process parameters and baculovirus constructs. The high-resolution data generated by D3HM showed the exact moment of peak cell densities and temporary events caused by feeding. Furthermore, D3HM allowed us to obtain information on the state of the cell culture at the individual cell level. Combining this detailed, real-time information about cell cultures with methodical machine learning models can increase process understanding, aid in decision-making, and allow for timely process control actions during bioreactor production of recombinant proteins.

The potential of sponge-derived chemicals for pharmaceutical applications remains largely unexploited due to limited available biomass. Although many have attempted to culture marine sponge cells in vitro to create a scalable production platform for such biopharmaceuticals, these efforts have been mostly unsuccessful. We recently showed that Geodia barretti sponge cells could divide rapidly in M1 medium. In this study we established the first continuous marine sponge cell line, originating from G. barretti. G. barretti cells cultured in OpM1 medium, a modification of M1, grew more rapidly and to a higher density than in M1. Cells in OpM1 reached 1.74 population doublings after 30 min, more than twofold higher than the already rapid growth rate of 0.74 population doublings in 30 min in M1. The maximum number of population doublings increased from 5 doublings in M1 to at least 98 doublings in OpM1. Subcultured cells could be cryopreserved and used to inoculate new cultures. With these results, we have overcome a major obstacle that has blocked the path to producing biopharmaceuticals with sponge cells at industrial scale for decades.

BACKGROUND: The prevalence of atrial fibrillation(AF) and diabetes mellitus is rising to epidemic proportions. We aimed to assess the impact of diabetes on the management and outcomes of patients with AF.METHODS: The EORP-AF General Long-Term Registry is a prospective, observational registry from 250 centres across 27 European countries. Outcomes of interest were as follows: i)rhythm control interventions; ii)quality of life; iii)healthcare resource utilisation; and iv)major adverse events.RESULTS: Of 11,028 patients with AF, the median age was 71 (63-77) years and 2537 (23.0%) had diabetes. Median follow-up was 24 months. Diabetes was related to increased use of anticoagulation but less rhythm control interventions. Using multivariable analysis, at 2-year follow-up, patients with diabetes were associated with greater levels of anxiety (p = 0.038) compared to those without diabetes. Overall, diabetes was associated with worse health during follow-up, as indicated by Health Utility Score and Visual Analogue Scale. Healthcare resource utilisation was greater with diabetes in terms of length of hospital stay (8.1 (±8.2) vs. 6.1 (±6.7) days); cardiology and internal medicine/general practitioner visits; and emergency room admissions. Diabetes was an independent risk factor of major adverse cardiovascular event (MACE; HR 1.26 [95% CI, 1.04-1.52]), all-cause mortality (HR 1.28 [95% CI, 1.08-1.52]), and cardiovascular mortality (HR 1.41 [95% CI, 1.09-1.83]).CONCLUSION: In this contemporary AF cohort, diabetes was present in 1 in 4 patients and it served as an independent risk factor for reduced quality of life, greater healthcare resource utilisation and excess MACE, all-cause mortality and cardiovascular mortality. There was increased use of anticoagulation therapy in diabetes but with less rhythm control interventions.

Karen Kaiser,1 Susan E Yount,1 Christa E Martens,1 Kimberly A Webster,1 Sara Shaunfield,1 Amy Sparling,1 John Devin Peipert,1 David Cella,1 Scott T Rottinghaus,2 Bonnie MK Donato,2 Richard Wells,3 Ioannis Tomazos2 1Department of Medical Social Sciences, Northwestern University Feinberg School of Medicine, Chicago, IL, USA; 2Alexion Pharmaceuticals Inc, Boston, MA, USA; 3Sunnybrook Health Sciences Centre, Toronto, ON, CanadaCorrespondence: Karen KaiserDepartment of Medical Social Sciences, Northwestern University Feinberg School of Medicine, 625 N. Michigan Ave. Suite 2700, Chicago, IL 60611, USATel +1 312-503-3521Fax +1 312-503-4800Email k-kaiser@northwestern.eduPurpose: To develop a patient preference questionnaire (PPQ) assessing eculizumab and ravulizumab treatment for paroxysmal nocturnal hemoglobinuria (PNH).Patients and Methods: The development of the PNH-PPQ© was consistent with Food and Drug Administration guidelines for patient-reported outcome measure development, and included 1) a targeted literature review; 2) PNH expert clinician input on treatment preferences; 3) review of existing qualitative data on the PNH treatment and disease experience; 4) concept elicitation interviews with 8 PNH patients who received eculizumab and/or ravulizumab; 5) translatability review; and 6) cognitive debriefing with 5 patients. Interview participants were recruited through a United Kingdom PNH patient advocacy group and a Canadian clinical site involved in clinical trial ALXN1210-PNH-302.Results: Six themes were identified as most relevant to the PNH treatment experience from the concept elicitation interviews: disease symptoms (n=8/8); treatment frequency (n=7/8); quality of life impact of treatment/disease (n=7/8); treatment burden (n=7/8); treatment efficacy (n=5/8); and treatment side effects (n=5/8). An initial list of 88 preference questions was reduced to 11 highly relevant and non-redundant questions reflecting the 6 themes. Cognitive interview participants unanimously agreed that the PNH-PPQ instructions were clear; response options were understandable, easy to use, and provided enough choices; and the questions captured the factors that inform treatment preferences.Discussion: When new drugs have similar efficacy to existing medications, documenting patient preferences is important for confirming patient benefit from the new medication. Understanding what matters most to patients is essential for delivering patient-centered care and may play a particularly significant role in treatment decision making. The availability of such a tool may be especially important as new orphan drugs are developed and patients with rare diseases have more than one treatment option to consider.Conclusion: The PNH-PPQ provides a patient-centered approach for evaluating preferences for the treatment of PNH. The PNH-PPQ has subsequently assessed patient preference in the clinical trial sub-study ALXN1210-PNH-302s.Keywords: paroxysmal nocturnal hemoglobinuria, questionnaire development, ravulizumab, eculizumab, treatment experience

BACKGROUND: Cardiac troponins (cTn) have been reported to be predictors for adverse outcomes in atrial fibrillation (AF), patients, but their actual use is still unclear.AIM: To assess the factors associated with cTn testing in routine practice and evaluate the association with outcomes.METHODS: Patients enrolled in the ESC-EHRA EORP-AF General Long-Term Registry were stratified into 3 groups according to cTn levels as (i) cTn not tested, (ii) cTn in range (≤99th percentile), (iii) cTn elevated (>99th percentile). The composite outcome of any thromboembolism /any acute coronary syndrome/cardiovascular (CV) death, defined as Major Adverse Cardiovascular Events (MACE) and all-cause death were the main endpoints.RESULTS: Among 10 445 AF patients (median age 71 years, 40.3% females) cTn were tested in 2834 (27.1%). cTn was elevated in 904/2834 (31.9%) and in-range in 1930/2834 (68.1%) patients. Female sex, in-hospital enrollment, first-detected AF, CV risk factors, history of coronary artery disease, and atypical AF symptoms were independently associated with cTn testing. Elevated cTn were independently associated with a higher risk for MACE (Model 1, hazard ratio [HR] 1.74, 95% confidence interval [CI] 1.40-2.16, Model 2, HR 1.62, 95% CI 1.28-2.05; Model 3 HR 1.76, 95% CI 1.37-2.26) and all-cause death (Model 1, HR 1.45, 95% CI 1.21-1.74; Model 2, HR 1.36, 95% CI 1.12-1.66; Model 3, HR 1.38, 95% CI 1.12-1.71).CONCLUSIONS: Elevated cTn levels were associated with an increased risk of all-cause mortality and adverse CV events. Clinical factors that might enhance the need to rule out CAD were associated with cTn testing.

Chikungunya virus (CHIKV) is a rapidly emerging mosquito-borne virus that causes a severe febrile illness with long-lasting arthralgia in humans. As there is no vaccine to protect humans and limit CHIKV epidemics, the virus continues to be a global public health concern. The CHIKV envelope glycoproteins E1 and E2 are important immunogens; therefore, the aim of this study is to produce trimeric CHIKV spikes in insect cells using the baculovirus expression system. The CHIKV E1 and E2 ectodomains were covalently coupled by a flexible linker that replaces the 6K transmembrane protein. The C-terminal E1 transmembrane was replaced by a Strep-tag II for the purification of secreted spikes from the culture fluid. After production in Sf9 suspension cells (product yields of 5.8–7.6 mg/L), the CHIKV spikes were purified by Strep-Tactin affinity chromatography, which successfully cleared the co-produced baculoviruses. Bis(sulfosuccinimidyl)suberate cross-linking demonstrated that the spikes are secreted as trimers. PNGase F treatment showed that the spikes are glycosylated. LC–MS/MS-based glycoproteomic analysis confirmed the glycosylation and revealed that the majority are of the mannose- or hybrid-type N-glycans and

Process intensification is increasingly used in the mammalian biomanufacturing industry. The key driver of this trend is the need for more efficient and flexible production strategies to cope with the increased demand for biotherapeutics predicted in the next years. Therefore, such intensified production strategies should be designed, established, and characterized. We established a CHO cell process consisting of an intensified fed-batch (iFB), which is inoculated by an N-1 perfusion process that reaches high cell concentrations (100 × 106 c ml−1). We investigated the impact of butyric acid (BA) supplementation in this iFB process. Most prominently, higher cellular productivities of more than 33% were achieved, thus 3.5 g L−1 of immunoglobulin G (IgG) was produced in 6.5 days. Impacts on critical product quality attributes were small. To understand the biological mechanisms of BA in the iFB process, we performed a detailed transcriptomic analysis. Affected gene sets reflected concurrent inhibition of cell proliferation and impact on histone modification. These translate into subsequently enhanced mechanisms of protein biosynthesis: enriched regulation of transcription, messenger RNA processing and transport, ribosomal translation, and cellular trafficking of IgG intermediates. Furthermore, we identified mutual tackling points for optimization by gene engineering. The presented strategy can contribute to meet future requirements in the continuously demanding field of biotherapeutics production.

Purpose Liver steatosis is a frequently reported condition in patients with inflammatory bowel disease (IBD). Different factors, both metabolic and IBD-associated, are believed to contribute to the pathogenesis. The aim of our study was to calculate the prevalence of liver steatosis and fibrosis in IBD patients and to evaluate which factors influence changes in steatosis and fibrosis during follow-up. Methods From June 2017 to February 2018, demographic and biochemical data was collected at baseline and after 6-12 months. Measured by transient elastography (FibroScan), liver steatosis was defined as Controlled Attenuation Parameter (CAP) >= 248 and fibrosis as liver stiffness value (Emed) >= 7.3 kPa. IBD disease activity was defined as C-reactive protein (CRP) >= 10 mg/l and/or fecal calprotectin (FCP) >= 150 mu g/g. Univariate and multivariate regression analysis was performed; a p-value of

Outer membrane vesicles (OMVs) are nanoparticles produced by Gram-negative bacteria that can be used as vaccines. The application of OMVs as vaccine component can be expanded by expressing heterologous antigens on OMVs, creating an OMV-based antigen presenting platform. This study aims to develop a production process for such OMV-based vaccines and studies a production method based on meningococcal OMVs that express heterologous antigens on their surface. As a proof of concept, the Borrelia burgdorferi antigens OspA and OspC were expressed on Neisseria meningitidis OMVs to create a concept anti-Lyme disease vaccine. Production of OMVs released in the culture supernatant was induced by high dissolved oxygen concentrations and purification was based on scalable unit operations. A crude recovery of 90 mg OMV protein could be obtained per liter culture. Expressing heterologous antigens on the OMVs did result in minor reduction of bacterial growth, while OMV production remained constant. The antigen expression did not alter the OMV characteristics. This study shows that production of well characterized OMVs containing heterologous antigens is possible with high yields by combining high oxygen concentrations with an optimized purification process. It is concluded that heterologous OMVs show potential as a vaccine platform. Unit Licence Agreement

Funding Acknowledgements Type of funding sources: Private company. Main funding source(s): Biotronik SE & Co.KG OnBehalf BIO|CONCEPT.BIOMONITOR III study group, BIO|MASTER.BIOMONITOR III study group, BIO|STREAM-ICM study group Background Implantable Cardiac Monitors (ICM) provide continuous long-term heart rhythm monitoring. The new ICM BIOMONITOR III / IIIm (BM III) is provided with a single-step insertion tool. Purpose To report on the insertion procedure of the BM III in a large real-world patient population. Methods The BM III combines a low cross-section (4.5 x 8.5 mm) with an extended ICM length (77 mm, including flexible antenna). It is inserted into subcutaneous tissue with an ‘injection’ tool that forms the pocket and delivers the device in a single step. We report results of the insertion procedure from a pooled data set from the BIO|CONCEPT BM III (completed) and the BIO|MASTER BM III and BIO|STREAM-ICM (ongoing) studies. Results From 54 investigational sites in 11 countries, 455 insertions were reported (including 39 BM IIIm). The patients were 63 ± 16 years old, had a BMI of 27.6 ± 5.4, and 43% were women. The indications were syncope or pre-syncope (57%), cryptogenic stroke (23%), management of AF (11%) or other (9%). Insertions took 1.7 ± 1.8 minutes until removal of the insertion tool, 4.7 ± 3.4 minutes until wound closure, and 7.1 ± 5.6 minutes including wound cleaning. The wound was sutured (79%) or closed with staples (10%) or adhesive strips (10%). General anaesthesia was used in 8% of the patients and antibiotic prophylaxis in 50% (44% systemic and 6% local). Insertions took place in the catheter laboratory (62%), operating theatre (22%) or in a consultation room (16%) without specific precautional equipment. The insertion site was parallel to the heart"s long axis (56%), parasternal (39%), in the 2nd/3rd intercostal space (3.5%), axillary (0.9%) or at the clavicula (0.7%). The device was repositioned in one case (0.2%). 13 adverse events were reported in connection to the insertion procedure. 5 cases of device pocket bleeding or hematoma occurred. In 5 further cases, the device migrated, posing the risk of extrusion, or actually extruded. Three of these cases used only adhesive strips or no wound closure at all. In two cases, an incorrect usage of the incision tool and substantial subcutaneous fatty tissue may have contributed. One device was damaged by a 200 J defibrillation shock with a shock electrode placed over the device. One patient suffered from dyspnoea, possibly due to psychogenic hyperventilation. One patient had a vasovagal syncope due to pain after an insertion with insufficient local anaesthesia. No infections were reported until the day of analysis, which was more than 30 days after insertion in 92% of all cases. Conclusion The new BM III was inserted in typically less than 5 minutes until wound closure. A relevant number of insertions took place in a consultation room. Prophylactic antibiotics may be unnecessary, because no pocket infections were reported, although no antibiotic prophylaxis was used in one half of all cases (N = 229). In summary, the insertion with the new tool is fast and has a low risk of complications.

Besides being considered pathogens, viruses are important drivers of evolution and they can shape large ecological and biogeochemical processes, by influencing host fitness, population dynamics, and community structures. Moreover, they are simple systems that can be used and manipulated to be beneficial and useful for biotechnological applications. In this context, microalgae biotechnology is a growing field of research, which investigated the usage of photosynthetic microorganisms for the sustainable production of food, fuel, chemical, and pharmaceutical sectors. Viruses infecting microalgae have become important subject of ecological studies related to marine and aquatic environments only four decades ago when virus-like-particles associated with bloom-forming algae were discovered. These first findings have opened new questions on evolution and identity. To date, 63 viruses that infect eukaryotic microalgae have been isolated and cultured. In this short review we briefly summarize what is known about viruses infecting eukaryotic microalgae, and how acknowledging their importance can shape future research focussed not only on marine ecology and evolutionary biology but also on biotechnological applications related to microalgae cell factories.

Current CHO cell production processes require an optimized space-time-yield. Process intensification can support achieving this by enhancing the productivity and improving facility utilization. The use of perfusion at the last stage of the seed train (N-1) for high cell density inoculation of the fed-batch N-stage production culture is a relatively new approach with few industry applicable examples. Within this work, the impact of the cell-specific perfusion rate (CSPR) of the N-1 perfusion and the relevance of its control for the quality of generated inoculation cells was evaluated using an automated perfusion rate (PR) control based on online biomass measurements. Precise correlations (R² = 0.99) between permittivity and viable cell counts were found up to the high densities of 100⋅106 c·mL−1. Cells from N-1 perfusion were cultivated at a high and low CSPR with 50 and 20 pL·(c·d)−1, respectively. Lowered cell growth and an increased apoptotic reaction was found as a consequence of the latter due to nutrient limitations and reduced uptake rates. Subsequently, batch cultivations (N-stage) from the different N-1 sources were inoculated to evaluate the physiological state of the inoculum. Successive responses resulting from the respective N-1 condition were uncovered. While cell growth and productivity of approaches inoculated from high CSPR and a conventional seed were comparable, low CSPR inoculation suffered significantly in terms of reduced initial cell growth and impaired viability. This study underlines the importance to determine the CSPR for the design and implementation of an N-1 perfusion process in order to achieve the desired performance at the crucial production stage.

Currently, the mammalian biomanufacturing industry explores process intensification (PI) to meet upcoming demands of biotherapeutics while keeping production flexible but, more importantly, as economic as possible. However, intensified processes often require more development time compared with conventional fed-batches (FBs) preventing their implementation. Hence, rapid and efficient, yet straightforward strategies for PI are needed. In this study we demonstrate such a strategy for the intensification of an N-stage FB by implementing N-1 perfusion cell culture and high inoculum cell densities resulting in a robust intensified FB (iFB). Furthermore, we show successful combination of such an iFB with the addition of productivity enhancers, which has not been reported so far. The conventional CHO cell FB process was step-wise improved and intensified rapidly in multi-parallel small-scale bioreactors using N-1 perfusion. The iFBs were performed in 15 and 250 ml bioreactors and allowed to evaluate the impact on key process indicators (KPI): the space–time yield (STY) was successfully doubled from 0.28 to 0.55 g/L d, while product quality was maintained. This gain was generated by initially increasing the inoculation density, thus shrinking process time, and second supplementation with butyric acid (BA), which reduced cell growth and enhanced cell-specific productivity from ~25 to 37 pg/(cell d). Potential impacts of PI on cell metabolism were evaluated using flux balance analysis. Initial metabolic differences between the standard and intensified process were observed but disappeared quickly. This shows that PI can be achieved rapidly for new as well as existing processes without introducing sustained changes in cellular and metabolic behavior.

Vaccines pave the way out of the SARS-CoV-2 pandemic. Besides mRNA and adenoviral vector vaccines, effective protein-based vaccines are needed for immunization against current and emerging variants. We have developed a virus-like particle (VLP)-based vaccine using the baculovirus-insect cell expression system, a robust production platform known for its scalability, low cost, and safety. Baculoviruses were constructed encoding SARS-CoV-2 spike proteins: full-length S, stabilized secreted S, or the S1 domain. Since subunit S only partially protected mice from SARS-CoV-2 challenge, we produced S1 for conjugation to bacteriophage AP205 VLP nanoparticles using tag/catcher technology. The S1 yield in an insect-cell bioreactor was similar to 11 mg/liter, and authentic protein folding, efficient glycosylation, partial trimerization, and ACE2 receptor binding was confirmed. Prime-boost immunization of mice with 0.5 mu g S1-VLPs showed potent neutralizing antibody responses against Wuhan and UK/B.1.1.7 SARS-CoV-2 variants. This two-component nanoparticle vaccine can now be further developed to help alleviate the burden of COVID-19.IMPORTANCE Vaccination is essential to reduce disease severity and limit the transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Protein-based vaccines are useful to vaccinate the world population and to boost immunity against emerging variants. Their safety profiles, production costs, and vaccine storage temperatures are advantageous compared to mRNA and adenovirus vector vaccines. Here, we use the versatile and scalable baculovirus expression vector system to generate a two-component nanoparticle vaccine to induce potent neutralizing antibody responses against SARS-CoV-2 variants. These nanoparticle vaccines can be quickly adapted as boosters by simply updating the antigen component.

Introduction: Student-staff partnerships as a concept to improve medical education have received a growing amount of attention. Such partnerships are collaborations in which students and teachers seek to improve education by each adding their unique contribution to decision-making and implementation processes. Although previous research has demonstrated that students are favourable to this concept, teachers remain hesitant. The present study investigated teachers’ conceptions of student-staff partnerships and of the prerequisites that are necessary to render such partnerships successful and enhance educational quality. Method: We conducted semi-structured interviews with 14 course coordinators who lead course design teams and also teach in 4 bachelor health programmes, using Bovill and Bulley’s levels of student participation as sensitising concepts during data analysis. Results: The results pointed to three different conceptions of student-staff partnerships existing among teachers: Teachers teach and students study; teachers teach and value students’ feedback; and teachers and students co-create. The prerequisites for effective co-creation teachers identified were: Teachers must be open to involve students and create dialogues; students must be motivated and have good communication skills; the organisation must be supportive; and teachers should have the final say. Conclusion: We conclude that teachers’ conceptions are consistent with Bovill and Bulley’s levels of student participation. Under certain conditions, teachers are willing to co-create and reach the highest levels of student participation.

Genetic transformation of microalgae remains a challenge due to poor intracellular delivery of exogenous molecules. This limitation is caused by the structure and composition of the cell wall and cell membrane of each species. Moreover, successful delivery of proteins or nucleic acids cannot be assessed by determining transformability since their functionality is not always known in the studied microorganism. We propose a quick and effective screening tool for the prediction and optimization of electroporation settings by monitoring cell permeability and viability using Sytox Green and propidium iodide respectively. We determined voltage settings for the microalgae Chlamydomonas reinhardtii, Chlorella vulgaris, Neochloris oleoabundans and Acutodesmus obliquus. To evaluate the predicted settings, we delivered labelled DNA and proteins into the cells. We demonstrated that high transformation efficiencies can be accomplished when predicted values were applied with functional plasmids. Additionally, we increased transformation efficiencies by testing cell concentrations, light intensities and fragment sizes. This method can be used to determine suitable transformation conditions for non-transformed microalgae species and to increase the insight on established transformation protocols.

This study explored the experiences of healthy elderly regularly visiting the allotment gardens (AGs) in a large city of Norway. Semistructured interviews with eight elderly visitors were analyzed ...

Background. Students are ever more involved in the design of educational practices, which is reflected in the growing body of literature about approaches to student participation. Similarities and differences between these approaches often remain vague since the terms are used interchangeably. This confusing and fragmented body of literature hampers our understanding the process and outcomes of student participation and choosing the most suitable approach for it.Method. We identified the three most frequently used terms related to the design of learning and teaching – design-based research (DBR), participatory design (PD), and co-creation – and disentangled the terminology by focusing on relevant definitions, aims, involvement of students, outcomes, and related terminology.Results. Differences between the approaches to student participation can be found in the degree to which students are the central actors and the degree to which the design is informed by educational theory.Conclusion. It is important to align the level of student participation with the purpose of the approach.

Normally, the growth profile of a CHO cell fed-batch process can be divided into two main phases based on changes in cell concentration, being an exponential growth phase and a stationary (non-growth) phase. In this study, an additional phase is observed during which the cell division comes to a halt but the cell growth continues in the form of an increase in cell size. The cell size increase (SI) phase occurs between the exponential proliferation phase (also called the number increase or NI phase) and the stationary phase. During the SI phase, the average volume and dry weight per cell increase threefold linearly with time. The average mAb specific productivity per cell increases linearly with the cell volume and therefore is on average two times higher in the SI phase than in the NI phase. The specific essential amino acids consumption rates per cell remain fairly constant between the NI and the SI phase, which agrees with the similar biomass production rate per cell between these two phases. Accumulation of fatty acids and formation of lipid droplets in the cells are observed during the SI phase, indicating that the fatty acids synthesis rate exceeds the demand for the synthesis of membrane lipids. A metabolic comparison between NI and SI phase shows that the cells with a larger size produce more mAb per unit of O2 and nutrient consumed, which can be used for further process optimization. Electronic supplementary material The online version of this article (10.1007/s00253-017-8531-y) contains supplementary material, which is available to authorized users.

To investigate apoptosis as a mechanism of sulfur mustard (SM) inhalation injury in animals, we studied different caspases (caspase-8, -9, -3, and -6) in the lungs from a ventilated rat SM aerosol inhalation model. SM activated all four caspases in cells obtained from bronchoalveolar lavage fluid (BALF) as early as 6 h after exposure. Caspase-8, which is known to initiate the extrinsic Fas-mediated pathway of apoptosis, was increased fivefold between 6 and 24 h, decreasing to the unexposed-control level at 48 h. The initiator, caspase-9, in the intrinsic mitochondrial pathway of apoptosis as well as the executioner caspases, caspase-3 and -6, all peaked ( P < 0.01) at 24 h; caspase-3 and -6 remained elevated, but caspase-9 decreased to unexposed-control level at 48 h. To study further the Fas pathway, we examined soluble as well as membrane-bound Fas ligand (sFas-L and mFas-L, respectively) and Fas receptor (Fas-R) in both BALF cells and BALF. At 24 h after SM exposure, sFas-L increased significantly in both BALF cells ( P < 0.01) and BALF ( P < 0.05). However, mFas-L increased only in BALF cells between 24 and 48 h ( P < 0.1 and P < 0.001, respectively). Fas-R increased only in BALF cells by 6 h ( P < 0.01) after SM exposure. Apoptosis in SM-inhaled rat lung specimens was also confirmed by both immunohistochemical staining using cleaved caspase-3 and -9 antibodies and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining as early as 6 h in the proximal trachea and bronchi, but not before 48 h in distal airways. These findings suggest pathogenic mechanisms at the cellular and molecular levels and logical therapeutic target(s) for SM inhalation injury in animals.

The authors compared medication-induced mood switch risk (primary outcome), as well as treatment response and side effects (secondary outcomes) with three acute-phase treatments for bipolar II depression.In a 16-week, double-blind, multisite comparison study, 142 participants with bipolar II depression were randomly assigned to receive lithium monotherapy (N=49), sertraline monotherapy (N=45), or combination treatment with lithium and sertraline (N=48). At each visit, mood was assessed using standardized rating scales. Rates of switch were compared, as were rates of treatment response and the presence and severity of treatment-emergent side effects.Twenty participants (14%) experienced a switch during the study period (hypomania, N=17; severe hypomania, N=3). Switch rates did not differ among the three treatment groups, even after accounting for dropout. No patient had a manic switch or was hospitalized for a switch. Most switches occurred within the first 5 weeks of treatment. The treatment response rate for the overall sample was 62.7% (N=89), without significant differences between groups after accounting for dropout. The lithium/sertraline combination group had a significantly higher overall dropout rate than the monotherapy groups but did not have an accelerated time to response.Lithium monotherapy, sertraline monotherapy, and lithium/sertraline combination therapy were associated with similar switch and treatment response rates in participants with bipolar II depression. The dropout rate was higher in the lithium/sertraline combination treatment group, without any treatment acceleration advantage.

Contains fulltext : 225487.pdf (Publisher’s version ) (Open Access) BACKGROUND: Eculizumab has transformed management of paroxysmal nocturnal hemoglobinuria (PNH) since its approval. However, its biweekly dosing regimen remains a high treatment burden. Ravulizumab administered every 8 weeks demonstrated noninferiority to eculizumab in two phase 3 trials. In regions where two PNH treatment options are available, it is important to consider patient preference. OBJECTIVE: The aim of this study was to assess patient preference for ravulizumab or eculizumab. METHODS: Study 302s (ALXN1210-PNH-302s) enrolled PNH patients who participated in the extension period of phase 3 study ALXN1210-PNH-302. In the parent study, eculizumab-experienced adult PNH patients received ravulizumab or eculizumab during a 26-week primary evaluation period. All patients in the extension period received ravulizumab. In study 302s, patient treatment preference was evaluated using an 11-item PNH-specific Patient Preference Questionnaire (PNH-PPQ©). Of 98 patients, 95 completed PNH-PPQ© per protocol for analysis. RESULTS: Overall, 93% of patients preferred ravulizumab whereas 7% of patients either had no preference (6%) or preferred eculizumab (1%) (P < 0.001). For specific aspects of treatment, ravulizumab was preferred (in comparison to no preference or eculizumab) on infusion frequency (98% vs. 0% vs. 2%), ability to plan activities (98% vs. 0% vs. 2%), and overall quality of life (88% vs. 11% vs. 1%), among other aspects. Most participants selected frequency of infusions as the most important factor determining preference (43%), followed by overall quality of life (23%). CONCLUSION: This study shows that a substantial proportion of patients preferred ravulizumab over eculizumab and provides an important patient perspective on PNH treatment when there is more than one treatment option.

Baculovirus expression vectors are successfully used for the commercial production of complex (glyco)proteins in eukaryotic cells. The genome engineering of single-copy baculovirus infectious clones (bacmids) in E. coli has been valuable in the study of baculovirus biology, but bacmids are not yet widely applied as expression vectors. An important limitation of first-generation bacmids for large-scale protein production is the rapid loss of gene of interest (GOI) expression. The instability is caused by the mini-F replicon in the bacmid backbone, which is non-essential for baculovirus replication in insect cells, and carries the adjacent GOI in between attTn7 transposition sites. In this paper, we test the hypothesis that relocation of the attTn7 transgene insertion site away from the mini-F replicon prevents deletion of the GOI, thereby resulting in higher and prolonged recombinant protein expression levels. We applied lambda red genome engineering combined with SacB counterselection to generate a series of bacmids with relocated attTn7 sites and tested their performance by comparing the relative expression levels of different GOIs. We conclude that GOI expression from the odv-e56 (pif-5) locus results in higher overall expression levels and is more stable over serial passages compared to the original bacmid. Finally, we evaluated this improved next-generation bacmid during a bioreactor scale-up of Sf9 insect cells in suspension to produce enveloped chikungunya virus-like particles as a model vaccine.

Sponges and their associated microorganisms are the most prolific source of marine natural products, and many attempts have been made at creating a marine sponge cell line to produce these products efficiently. However, limited knowledge on the nutrients sponge cells require to grow and poor genetic accessibility have hampered progress toward this goal. Recently, a new sponge-specific nutrient medium M1 has been shown to stimulate sponge cells in vitro to divide rapidly. In this study, we demonstrate for the first time that sponge cells growing in M1 can be genetically modified using a CRISPR/Cas12a gene editing system. A short sequence of scrambled DNA was inserted using a single-stranded oligodeoxynucleotide donor template to disrupt the 2′,5′-oligoadenylate synthetase gene of cells from the boreal deep-sea sponge Geodia barretti. A blue fluorescent marker gene appeared to be inserted in an intron of the same gene and expressed by a small number of G. barretti cells. Our results represent an important step toward developing an optimized continuous sponge cell line to produce bioactive compounds.

Cryopreservation is a commonly used method for the long-term storage of cell lines and provides a stable source of cells for experiments, allowing researchers to study species that are not geographically nearby, and useful to progress studies on sponge cell biotechnology.The marine sponge Dysidea etheria was chosen as our model organism to evaluate the impact and effectiveness of two commonly used cryoprotectants, dimethyl sulfoxide (DMSO) and glycerol.By testing a range of concentrations (3-10% DMSO, 10-50% glycerol), we determined the optimal cryoprotectant for D. etheria based on its ability to preserve viable cells and optimize recovery after cryopreservation.Cells cryopreserved in DMSO had significantly higher viability after cryopreservation than those cryopreserved in glycerol. Cells cryopreserved in glycerol had irregular morphology as well as lower recovery of viable cells than those from DMSO treatments.Our results demonstrate that the optimal cryoprotectant for sponge cells, without a significant loss of viability, is 5-8% DMSO. This approach can be used to optimize cryopreservation methods for cells of other marine invertebrate species.

In this study the influence of day length on circadian rhythms of N. oleoabundans was investigated. N. oleoabundans was grown in a photobioreactor continuously operated (turbidostat), under various day/night (D/N) cycles; 20D4N, 16D8N and 12D12N. The following variables showed to be regulated by the circadian clock: maximum growth rate, start of starch synthesis and DNA replication to 4 and 8 copies. Timing of these processes was not influenced by day length. The length of the photoperiod caused changes in biomass composition, especially due to variations in starch content. In longer days, more starch was accumulated. Starch was used for cell division probably also when cell division occurred in the light. Therefore, also the timing of cell division should be considered in the production of microalgae biomass. This timing mechanism can be explored to produce biomass with a desired concentration of protein, lipids, carbohydrates or pigments.

The aim of this study is to investigate the effect of robot assisted laparoscopic sacrohysteropexy (RALS), with preservation of the uterus, in patients with pelvic organ prolapse on short and long term outcome. We report on (anatomical) status of the prolaps and the associated health related quality of life of women treated with RALS before and five years after surgery.A prospective cohort study in a teaching hospital in The Netherlands was performed. Quality of life was assessed pre-operative, post-operative and five years after RALS using the UDI/IIQ validated self-questionnaire designed for Dutch-speaking patients. Clinical and operative data were prospectively collected up to five years. Statistical analysis of categorical data was performed with the paired T-test. Descriptive statistics were computed with the use of standard methods for means, median and proportions.Hundred women with utero vaginal prolapse were treated with RALS with preservation of the uterus. The overall success rate of pelvic organ prolapse (POP) was 89.2%. After surgery the quality of life improved (P0.05) Overall health status, based on a 0-100% visual analogue scale (VAS), improved from 72.6% pre-operative to 82.2% six weeks postoperative (P0.05). Postoperative patients experienced less feelings of nervousness (P=0.01), shame (P0.05) and frustration (P0.05). The positive effects on these feelings remained present after five years. The learning curve shows a decrease in operating time with gained experience.RALS has proven to be a safe and effective treatment for uterine preserving surgery in cases of pelvic organ prolapse. The long term anatomical outcomes and quality of life after RALS compare favorably with laparoscopic and open hysteropexy.

Patient-centered communication (PCC) is an essential component of patient-centered care and contributes to patient satisfaction, health-related quality of life, and other important patient outcomes. The aim of this study was to develop and test survey questions to assess patients’ experiences with PCC in cancer care. We used a conceptual model developed by the National Cancer Institute as our framework. The survey questions align with the six core functions of PCC defined in the model: Exchanging Information, Managing Uncertainty, Enabling Patient Self-Management, Fostering Healing Relationships, Making Decisions, and Responding to Emotions. The study focused on colorectal cancer patients. We conducted two rounds of cognitive interviewing to evaluate patients’ ability to understand and provide valid answers to the PCC questions. Interviews were conducted in Maryland and North Carolina in 2014. We involved a patient advocacy group, Fight Colorectal Cancer, and a multidisciplinary panel of stakeholders throughout the measurement development process to ensure that the survey questions capture aspects of PCC that are important to patients and meet the needs of potential end users, including researchers, healthcare organizations, and health professionals. Patient and other stakeholder input informed revisions of draft survey questions, including changes to survey instructions, frame of reference for questions, response scales, and language. This study demonstrated the feasibility and value of engaging patients and other stakeholders in a measurement development study. The Patient-Centered Outcomes Research Institute (PCORI) conceptual model of patient-centered outcomes research provides a useful guide for patient engagement in research. Research funders should call for meaningful roles for patients and other stakeholders in health research, including in the development of patient-centered outcomes.

Rodenticides and pesticides pose a significant threat not only to the environment but also directly to humans by way of accidental and/or intentional exposure. Metal phosphides, such as aluminum, magnesium, and zinc phosphides, have gained popularity owing to ease of manufacture and application. These agents and their hydrolysis by-product phosphine gas (PH3 ) are more than adequate for eliminating pests, primarily in the grain storage industry. In addition to the potential for accidental exposures in the manufacture and use of these agents, intentional exposures must also be considered. As examples, ingestion of metal phosphides is a well-known suicide route, especially in Asia; and intentional release of PH3 in a populated area cannot be discounted. Metal phosphides cause a wide array of effects that include cellular poisoning, oxidative stress, cholinesterase inhibition, circulatory failure, cardiotoxicity, gastrointestinal and pulmonary toxicity, hepatic damage, neurological toxicity, electrolyte imbalance, and overall metabolic disturbances. Mortality rates often exceed 70%. There are no specific antidotes against metal phosphide poisoning. Current therapeutic intervention is limited to supportive care. The development of beneficial medical countermeasures will rely on investigative mechanistic toxicology; the ultimate goal will be to identify specific treatments and therapeutic windows for intervention.

Tetradesmus obliquus is an oleaginous microalga with high potential for triacylglycerol production. We characterized the biochemical composition and the transcriptional landscape of T. obliquus wild-type and the starchless mutant (slm1), adapted to 16:8 h light dark (LD) cycles under nitrogen limitation. In comparison to the nitrogen replete conditions, the diurnal RNA samples from both strains also displayed a cyclic pattern, but with much less variation which could be related to a reduced transcription activity in at least the usually highly active processes. During nitrogen limitation, the wild-type continued to use starch as the preferred storage compound to store energy and carbon. Starch was accumulated to an average content of 0.25 g·gDW−1, which is higher than the maximum observed under nitrogen replete conditions. Small oscillations were observed, indicating that starch was being used as a diurnal energy storage compound, but to a lesser extent than under nitrogen replete conditions. For the slm1 mutant, TAG content was higher than for the wild-type (average steady state value was 0.26 g·gDW−1 for slm1 compared to 0.06 g·gDW−1 for the wild-type). Despite the higher TAG content in the slm1, the conversion efficiency of photons into biomass components for the slm1 was only half of the one obtained for the wild-type. This is related to the observed decrease in biomass productivity (from 1.29 gDW·L−1·day−1 for the wild-type to 0.52 gDW·L−1·day−1 for the slm1). While the transcriptome of slm1 displayed clear signs of energy generation by degrading TAG and amino-acids during the dark period, no significant variation of these metabolites could be measured. When looking through the diurnal cycle, the photosynthetic efficiency was lower for the slm1 mutant compared to the wild-type especially during the second half of the light period, where starch accumulation occurred in the wild-type. Paid Open Access