Application of alternative cell separation systems for the harvest of mammalian cell culture processes in a fully disposable single-use facility

In a fully disposable facility where the use of continuous disk-stack centrifuges are not preferred, harvest processes based on conventional depth filtration become more challenging with increasing single-use bioreactor (SUB) size and higher density culture. Lower filterability due to high volume and high cell density diminishes the efficiency of the depth filtration. In addition, the use of larger depth filtration systems is constrained by the facility footprint. To address the challenge described, several alternative single-use harvest technologies were evaluated. A disposable centrifuge with a small foot print, low consumable cost, and ease of operation was tested. In a study using a 1kL SUB for harvest, the disposable centrifuge was compared to a continuous disk-stack centrifuge, both followed by conventional depth filtration. The best operating conditions, as well as the edges of failures, were found for the disposable centrifuge. When compared to the disk-stack centrifuge results showed that the operation can be performed in a wide range without impacting depth filtration area requirements and less depth filter area was needed. In addition to testing a disposable centrifuge, flocculation was evaluated and compared, followed by the depth filtration. This was found to be another low-cost alternative process, while also occupying less facility space. The flocculation technology was tested with different cell lines and results showed significant improvement in filterability and reduction of depth filter area compared to full traditional depth filtration train

Vitamin D Deficiency in Adult Patients with Schizophreniform and Autism Spectrum Syndromes: A One-Year Cohort Study at a German Tertiary Care Hospital

Introduction: Vitamin D has many immunomodulatory, anti-inflammatory, and neuroprotective functions, and previous studies have demonstrated an association between vitamin D deficiency and neuropsychiatric disease. The aim of our study was to analyze the prevalence of vitamin D deficiency in a one-year cohort of adult inpatients with schizophreniform and autism-spectrum syndromes in a naturalistic in-patient setting in Germany. Participants and methods: Our study was comprised of 60 adult schizophreniform and 23 adult high-functioning autism spectrum patients who were hospitalized Page: 2between January and December of 2015. We compared our findings with a historical German reference cohort of 3,917 adults using Pearson’s two-sided chi-squared test. The laboratory measurements of 25-hydroxyvitamin D2/3 (25(OH)vitamin D) were obtained using a chemiluminescence immunoassay. Results: In the schizophreniform group, we found decreased ( 30 ng/ml were observed in only 5% of the schizophreniform patients, 8.7% of the autism spectrum patients, and 21.9% of the healthy controls. Discussion: We found very high rates of 25(OH)vitamin D deficiency in both patient groups, and have discussed whether our findings might be related to alterations in the immunological mechanisms. Irrespective of the possible pathophysiological links between vitamin D deficiency and schizophrenia or autism spectrum disorders, a more frequent measurement of vitamin D levels seems to be justified in these patient groups. Further prospective, controlled, blinded, and randomized research should be conducted to analyze the effectiveness of vitamin D supplementation on the improvement of psychiatric symptoms

Alterations in Cerebrospinal Fluid in Patients with Bipolar Syndromes

Bipolar disorder (BD) is a severe and lifelong condition. Primary endogenic polygenetic forms are common. Secondary organic forms have received increasing interest recently due to the detection of immunological encephalopathies that mimic various psychiatric syndromes, including bipolar disorder. However, only limited data about routine findings of cerebrospinal fluid (CSF) analyses in bipolar disorder are available. Therefore, we investigated the frequency of alterations in the CSF in patients with BD and the association with autoantibodies, cerebral magnetic resonance imaging, and electroencephalography findings.CSF samples of patients with BD collected from January 1998 until December 2015 were analyzed retrospectively. Patients with preexisting causes for alterations in the CSF (e.g., patients with obvious past or current neurological disorders) were excluded. In total, 63 patients with BD fulfilled the inclusion criteria for the study. In 1.6% of the patients with BD, an increased white blood cell count was found in the CSF. Increased albumin quotients were found in 12.9% of the patients, oligoclonal bands (OCBs) in 1.6%, and increased immunoglobulin (Ig) G indices in 3.2% (OCBs were not measured in case of increased IgG indices). No significant differences in CSF findings were found between patients with manic and depressive episodes. The main findings of this open uncontrolled study are that alterations in the CSF may be found in a small but potentially relevant subgroup of patients with BD. These findings are discussed in light of the new concepts of mild encephalitis and immunological encephalopathy. The detection of patients with possibly secondary organic bipolar syndromes could open up new causal treatment options with immunomodulatory medication

Systemic Lupus Erythematosus With Isolated Psychiatric Symptoms and Antinuclear Antibody Detection in the Cerebrospinal Fluid

Background: Organic psychiatric disorders can be caused by immunological disorders, such as autoimmune encephalitis or systemic lupus erythematosus (SLE). SLE can affect most organs, as well as the central nervous system (CNS). In this paper, we describe a patient with an isolated psychiatric syndrome in the context of SLE and discuss the role of antibody detection in the cerebrospinal fluid (CSF).Case presentation: The 22-year-old German male high school graduate presented with obsessive–compulsive and schizophreniform symptoms. He first experienced obsessive–compulsive symptoms at the age of 14. At the age of 19, his obsessive thoughts, hallucinations, diffuse anxiety, depressed mood, severe dizziness, and suicidal ideation became severe and did not respond to neuroleptic or antidepressant treatment. Due to increased antinuclear antibodies (ANAs) with anti-nucleosome specificity in serum and CSF, complement activation, multiple bilateral white matter lesions, and inflammatory CSF alterations, we classified the complex syndrome as an isolated psychiatric variant of SLE. Immunosuppressive treatment with two times high-dose steroids, methotrexate, and hydroxychloroquine led to a slow but convincing improvement.Conclusion: Some patients with psychiatric syndromes and increased ANA titers may suffer from psychiatric variants of SLE, even if the American College of Rheumatology criteria for SLE are not met. Whether the psychiatric symptoms in our patient represent a prodromal stage with the later manifestation of full-blown SLE or a subtype of SLE with isolated CNS involvement remains unclear. Regardless, early diagnosis and initiation of immunosuppressive treatment are essential steps in preventing further disease progression and organ damage. Intrathecal ANAs with extractable nuclear antigen differentiation may be a more sensitive marker of CNS involvement compared with serum analyses alone

A peripheral epigenetic signature of immune system genes is linked to neocortical thickness and memory

Increasing age is tightly linked to decreased thickness of the human neocortex. The biological mechanisms that mediate this effect are hitherto unknown. The DNA methylome, as part of the epigenome, contributes significantly to age-related phenotypic changes. Here, we identify an epigenetic signature that is associated with cortical thickness (P=3.86 × 10(-8)) and memory performance in 533 healthy young adults. The epigenetic effect on cortical thickness was replicated in a sample comprising 596 participants with major depressive disorder and healthy controls. The epigenetic signature mediates partially the effect of age on cortical thickness (P<0.001). A multilocus genetic score reflecting genetic variability of this signature is associated with memory performance (P=0.0003) in 3,346 young and elderly healthy adults. The genomic location of the contributing methylation sites points to the involvement of specific immune system genes. The decomposition of blood methylome-wide patterns bears considerable potential for the study of brain-related traits

New Variant of MELAS Syndrome With Executive Dysfunction, Heteroplasmic Point Mutation in the MT-ND4 Gene (m.12015T>C; p.Leu419Pro) and Comorbid Polyglandular Autoimmune Syndrome Type 2

Background: Mitochondrial diseases are caused by dysfunctions in mitochondrial metabolic pathways. MELAS syndrome is one of the most frequent mitochondrial disorders; it is characterized by encephalopathy, myopathy, lactic acidosis, and stroke-like episodes. Typically, it is associated with a point mutation with an adenine-to-guanine transition at position 3243 of the mitochondrial DNA (mtDNA; m.3243A>G) in the mitochondrially encoded tRNA leucine 1 (MT-TL1) gene. Other point mutations are possible and the association with polyglandular autoimmune syndrome type 2 has not yet been described.Case presentation: We present the case of a 25-year-old female patient with dysexecutive syndrome, muscular fatigue, and continuous headache. Half a year ago, she fought an infection-triggered Addison crisis. As the disease progressed, she had two epileptic seizures and stroke-like episodes with hemiparesis on the right side. Cerebral magnetic resonance imaging showed a substance defect of the parieto-occipital left side exceeding the vascular territories with a lactate peak. The lactate ischemia test was clearly positive, and a muscle biopsy showed single cytochrome c oxidase-negative muscle fibers. Genetic testing of blood mtDNA revealed a heteroplasmic base exchange mutation in the mitochondrially encoded NADH:ubiquinone oxidoreductase core subunit 4 (MT-ND4) gene (m.12015T>C; p.Leu419Pro; heteroplasmy level in blood 12%, in muscle tissue: 15%). The patient suffered from comorbid autoimmune polyglandular syndrome type 2 with Hashimoto's thyroiditis, Addison's disease, and autoimmune gastritis. In addition, we found increased anti-glutamic acid decarboxylase 65, anti-partial cell, anti-intrinsic factor, and anti-nuclear antibodies.Conclusion: We present an atypical case of MELAS syndrome with predominant symptoms of a dysexecutive syndrome, two stroke-like episodes, and fast-onset fatigue. The symptoms were associated with a not yet described base and aminoacid exchange mutation in the MT-ND4 gene (m.12015T>C to p.Leu419Pro). The resulting changed protein complex in our patient is part of the respiratory chain multicomplex I and might be the reason for the mitochondriopathy. However, different simulations for pathogenetic relevance are contradictory and rather speak for a benign variant. To our knowledge this case report is the first reporting MELAS syndrome with comorbid polyglandular autoimmune syndrome type 2. Screening for autoimmune alterations in those patients is important to prevent damage to end organs

Inferior Frontal Gyrus Volume Loss Distinguishes Between Autism and (Comorbid) Attention-Deficit/Hyperactivity Disorder—A FreeSurfer Analysis in Children

Objective: Autism spectrum (ASD) and attention-deficit/hyperactivity disorder (ADHD) are neurodevelopmental disorders with a high rate of comorbidity. To date, diagnosis is based on clinical presentation and distinct reliable biomarkers have been identified neither for ASD nor ADHD. Most previous neuroimaging studies investigated ASD and ADHD separately.Method: To address the question of structural brain differences between ASD and ADHD, we performed FreeSurfer analysis in a sample of children with ADHD (n = 30), with high-functioning ASD (n = 14), with comorbid high-functioning ASD and ADHD (n = 15), and of typically developed controls (TD; n = 36). With FreeSurfer, an automated brain imaging processing and analyzing suite, we reconstructed the cerebral cortex and calculated gray matter volumes as well as cortical surface parameters in terms of cortical thickness and mean curvature.Results: A significant main effect of the factor ADHD was detected for the left inferior frontal gyrus (Pars orbitalis) volume, with the ADHD group exhibiting smaller Pars orbitalis volumes. Dimensional measures of autism (SRS total raw score) and ADHD (DISYPS-II FBB-ADHD score) had no significant influence on the left Pars orbitalis volume. Both, ASD and ADHD tended to have an effect on cortical thickness or mean curvature, which did not survive correction for multiple comparisons.Conclusion: Our results underline that ADHD rather than ASD is associated with volume loss in the left inferior frontal gyrus (Pars orbitalis). This area might play a relevant role in modulating symptoms of inattention and/or impulsivity in ADHD. The effect of comorbid ADHD in ASD samples and vice versa, on cortical thickness and mean curvature, requires further investigation in larger samples

M19 Modulates Skeletal Muscle Differentiation and Insulin Secretion in Pancreatic β-Cells through Modulation of Respiratory Chain Activity

Mitochondrial dysfunction due to nuclear or mitochondrial DNA alterations contributes to multiple diseases such as metabolic myopathies, neurodegenerative disorders, diabetes and cancer. Nevertheless, to date, only half of the estimated 1,500 mitochondrial proteins has been identified, and the function of most of these proteins remains to be determined. Here, we characterize the function of M19, a novel mitochondrial nucleoid protein, in muscle and pancreatic β-cells. We have identified a 13-long amino acid sequence located at the N-terminus of M19 that targets the protein to mitochondria. Furthermore, using RNA interference and over-expression strategies, we demonstrate that M19 modulates mitochondrial oxygen consumption and ATP production, and could therefore regulate the respiratory chain activity. In an effort to determine whether M19 could play a role in the regulation of various cell activities, we show that this nucleoid protein, probably through its modulation of mitochondrial ATP production, acts on late muscle differentiation in myogenic C2C12 cells, and plays a permissive role on insulin secretion under basal glucose conditions in INS-1 pancreatic β-cells. Our results are therefore establishing a functional link between a mitochondrial nucleoid protein and the modulation of respiratory chain activities leading to the regulation of major cellular processes such as myogenesis and insulin secretion

High-resolution CT phenotypes in pulmonary sarcoidosis: a multinational Delphi consensus study

One view of sarcoidosis is that the term covers many different diseases. However, no classification framework exists for the future exploration of pathogenetic pathways, genetic or trigger predilections, patterns of lung function impairment, or treatment separations, or for the development of diagnostic algorithms or relevant outcome measures. We aimed to establish agreement on high-resolution CT (HRCT) phenotypic separations in sarcoidosis to anchor future CT research through a multinational two-round Delphi consensus process. Delphi participants included members of the Fleischner Society and the World Association of Sarcoidosis and other Granulomatous Disorders, as well as members' nominees. 146 individuals (98 chest physicians, 48 thoracic radiologists) from 28 countries took part, 144 of whom completed both Delphi rounds. After rating of 35 Delphi statements on a five-point Likert scale, consensus was achieved for 22 (63%) statements. There was 97% agreement on the existence of distinct HRCT phenotypes, with seven HRCT phenotypes that were categorised by participants as non-fibrotic or likely to be fibrotic. The international consensus reached in this Delphi exercise justifies the formulation of a CT classification as a basis for the possible definition of separate diseases. Further refinement of phenotypes with rapidly achievable CT studies is now needed to underpin the development of a formal classification of sarcoidosis

CCWORK protocol: a longitudinal study of Canadian Correctional Workers' Well-being, Organizations, Roles and Knowledge.

IntroductionKnowledge about the factors that contribute to the correctional officer's (CO) mental health and well-being, or best practices for improving the mental health and well-being of COs, have been hampered by the dearth of rigorous longitudinal studies. In the current protocol, we share the approach used in the Canadian Correctional Workers' Well-being, Organizations, Roles and Knowledge study (CCWORK), designed to investigate several determinants of health and well-being among COs working in Canada's federal prison system.Methods and analysis CCWORK is a multiyear longitudinal cohort design (2018-2023, with a 5-year renewal) to study 500 COs working in 43 Canadian federal prisons. We use quantitative and qualitative data collection instruments (ie, surveys, interviews and clinical assessments) to assess participants' mental health, correctional work experiences, correctional training experiences, views and perceptions of prison and prisoners, and career aspirations. Our baseline instruments comprise two surveys, one interview and a clinical assessment, which we administer when participants are still recruits in training. Our follow-up instruments refer to a survey, an interview and a clinical assessment, which are conducted yearly when participants have become COs, that is, in annual 'waves'. Ethics and dissemination CCWORK has received approval from the Research Ethics Board of the Memorial University of Newfoundland (File No. 20190481). Participation is voluntary, and we will keep all responses confidential. We will disseminate our research findings through presentations, meetings and publications (e.g., journal articles and reports). Among CCWORK's expected scientific contributions, we highlight a detailed view of the operational, organizational and environmental stressors impacting CO mental health and well-being, and recommendations to prison administrators for improving CO well-being