A lifestyle score in childhood and adolescence was positively associated with subsequently measured fluid intelligence in the DONALD cohort study.

PURPOSE: Lifestyle scores which combine single factors such as diet, activity, or sleep duration showed associations with cognitive decline in adults. However, the role of a favourable lifestyle in younger age and the build-up of cognitive reserve is less clear, which is why we investigated longitudinal associations between a lifestyle score in childhood and adolescence and fluid intelligence obtained on average 6 years later. METHODS: In the DONALD cohort, a lifestyle score of 0 to 4 points including healthy diet and duration of moderate-to-vigorous physical activity, sedentary behaviour and sleep was repeatedly assessed in participants aged 5 and 19 years. Data on fluid intelligence were assessed via a German version of the culture fair intelligence test (CFT), using CFT 1-R in children 8.5 years of age or younger (n = 62) or CFT 20-R in participants older than 8.5 years (n = 192). Multivariable linear regression models were used to investigate prospective associations between the lifestyle score and the fluid intelligence score. RESULTS: Mean lifestyle score of all participants was 2.2 (0.7-4) points. A one-point increase in the lifestyle score was associated with a higher fluid intelligence score (4.8 points [0.3-7.3], p = 0.0343) for participants completing the CFT 20-R. Furthermore, each additional hour of sedentary behaviour was associated with a lower fluid intelligence score (- 3.0 points [- 5.7 to - 0.3], p = 0.0313). For younger participants (CFT 1-R), no association was found in any analysis (p > 0.05). CONCLUSION: A healthy lifestyle was positively associated with fluid intelligence, whereby sedentary behaviour itself seemed to play a prominent role.

Changes in patterns of eating habits and food intake during the first German COVID-19 lockdown: results of a cross-sectional online survey.

PURPOSE: The COVID-19 pandemic and public measures have a direct impact on the nutrition situation; studies show changes in food consumption, eating behavior or body weight but complex pattern analyses of changes rarely exist. METHODS: During the first German lockdown, a web-based survey was conducted among adults. It included 33 questions about changes in food intake, eating habits and physical activity, as well as anthropometrics and sociodemographic factors. Patterns of change were calculated based on changes in food intake and eating habits using two-step cluster analysis. To identify influencing factors for assignment to the patterns of change, binary logistic regression analyses were performed. RESULTS: Data from 2103 participants (81% female, 40 ± 14 years) were considered for analysis. Increased stockpiling, cooking, and variation in preparation was reported by 50-70%. The constant pattern (C-P, 36%) reported little change besides the above. The health-oriented pattern (HO-P; 37%) reported eating more healthy foods, avoiding unhealthy foods, and eating less and less frequently. The emotional-driven pattern (ED-P; 28%) exhibits higher influence of emotions on eating behavior, less avoidance of unhealthy foods, and increased consumption of sweets, pastries, and alcohol. The odds of changing eating behavior either to HO-P or ED-P were higher in women, people with migration background, younger participants, and increased with BMI categories. CONCLUSION: Both, the ED-P and HO-P, exhibit distinctive reactions in eating habits and food intake when dealing with a distressing experience. In subgroups, these may lead to disturbances in eating behavior and increase the risk for eating disorders and obesity.

Low leptin levels are associated with elevated physical activity among lean school children in rural Tanzania.

BACKGROUND: In Sub-Saharan African countries, rapid urbanization and increasing socio-economic status are associated with a transition to decreased physical activity (PA). A more sedentary lifestyle is linked to increased body fat leading to increments in leptin levels. Since rodent and human studies in high-income countries have shown that starvation-induced hypoleptinemia triggers high PA, efforts are warranted to pursue the hypothesis that low leptin levels in lean children of low- and middle-income countries (LMIC) are also associated with high PA. METHODS: In this cross-sectional study, we assessed seven-day PA with triaxial accelerometry (ActiGraph GT3X) among 223 primary school children (9 to 12 years of age) in rural Tanzania. Moderate-to-vigorous PA (MVPA) and total accelerometer counts per day were outcome variables. Leptin was determined using enzyme linked immunosorbent assay tests from dried blood spots. Anthropometric assessments were conducted and food insecurity and socio-demographic data were gathered using semi-structured interviews. RESULTS: In this sample of school children in rural Tanzania, leptin concentrations (median: 0.91 ng/mL, P25: 0.55, P75: 1.69), body mass index z-scores (median: -1.35, P25: -1.93, P75: -0.82), and height-for-age-z-scores (median: -1.16, P25: -1.96, P75: -0.61) were low. In contrast, PA levels were high with a median MVPA time of 119 min/day. Linear regression confirmed that leptin levels were negatively associated with MVPA (beta: -18.1; 95%CI: -29.7; -6.5; p = 0.002) and total accelerometer counts (beta: -90,256; 95%CI: -154,146; -26,365; p = 0.006). Children residing in areas with better infrastructure had lower MVPA levels (p < 0.001) and tended to have higher leptin levels (p = 0.062) than children residing in areas only reachable via dirt roads. CONCLUSION: Our cross-sectional field study is the first that supports the hypothesis of low leptin levels as a potential endocrine trigger of high PA in lean children of a LMIC. We observed early signs of a PA transition towards a less active lifestyle in a subgroup residing in areas with better infrastructure that concomitantly tended to have higher leptin concentrations. Considering that area-dependent PA differences were more pronounced among girls than boys, whereas differences in leptin levels were less pronounced, not only biological, but also external factors explain PA transition.

Rapid amelioration of anorexia nervosa in a male adolescent during metreleptin treatment including recovery from hypogonadotropic hypogonadism.

With this case report we support our medical hypothesis that metreleptin treatment ameliorates starvation related emotional, cognitive and behavioral symptomatology of anorexia nervosa (AN) and show for the first time strong effects in a male patient with AN. A 15.9 year old adolescent with severe AN of eight-month duration was treated off-label with metreleptin. Hyperactivity was assessed with accelerometry. Visual analogue scales (VAS), validated self- and clinician rating scales and lab results tracked changes from baseline to end of the 24-day dosing period and a five-month follow-up. Substantial improvements of mood and eating disorder related cognitions and hyperactivity set in after two days of treatment. During dosing, sub-physiological testosterone and TT3 levels normalized; clinically libido reemerged. Weight did not increase substantially during the dosing period. During follow-up target weight was attained; mood did not deteriorate; hyperactivity ceased. The results substantiate the strong effects seen in female cases and underscore the need for a double-blind placebo-controlled trial to confirm the observed strong, multiple and rapid onset beneficial effects of metreleptin in AN.

A mendelian randomization study on causal effects of 25(OH)vitamin D levels on attention deficit/hyperactivity disorder.

BACKGROUND: While observational studies revealed an inverse association between serum 25(OH)vitamin D (25(OH)D) and the risk of attention deficit/hyperactivity disorder (ADHD), the causality of this relationship remains unclear. METHODS: We conducted a bidirectional two-sample Mendelian Randomization (MR) study to examine whether 25(OH)D has an effect on the risk to develop ADHD or vice versa. Information on single nucleotide polymorphisms (SNP) associated with serum 25(OH)D was obtained from a genome-wide association study (GWAS) considering phenotype data from 79,366 individuals of European ancestry. Data on risk for ADHD were derived from a GWAS analysis with 20,183 individuals diagnosed with ADHD and 35,191 controls. For our analysis, we considered effect sizes based on the European participants (19,099 cases and 34,194 controls). RESULTS: Single SNP analyses showed a causal effect of vitamin D on ADHD risk for only one SNP (rs12785878, p = 0.024). The overall MR estimates did not reveal a causal effect of 25(OH)D on risk for ADHD. In the reverse analysis, neither any single nor the multi-SNP MR analyses showed a causal effect of ADHD on 25(OH)D. CONCLUSION: Results from this two-sample MR study did not confirm a causal effect of 25(OH)D on ADHD or vice versa. Accordingly, our study does not provide evidence that improving 25(OH)D via supplementation could reduce the risk of developing ADHD.

The impact of intragenic CpG content on gene expression.

The development of vaccine components or recombinant therapeutics critically depends on sustained expression of the corresponding transgene. This study aimed to determine the contribution of intragenic CpG content to expression efficiency in transiently and stably transfected mammalian cells. Based upon a humanized version of green fluorescent protein (GFP) containing 60 CpGs within its coding sequence, a CpG-depleted variant of the GFP reporter was established by carefully modulating the codon usage. Interestingly, GFP reporter activity and detectable protein amounts in stably transfected CHO and 293 cells were significantly decreased upon CpG depletion and independent from promoter usage (CMV, EF1 alpha). The reduction in protein expression associated with CpG depletion was likewise observed for other unrelated reporter genes and was clearly reflected by a decline in mRNA copy numbers rather than translational efficiency. Moreover, decreased mRNA levels were neither due to nuclear export restrictions nor alternative splicing or mRNA instability. Rather, the intragenic CpG content influenced de novo transcriptional activity thus implying a common transcription-based mechanism of gene regulation via CpGs. Increased high CpG transcription correlated with changed nucleosomal positions in vitro albeit histone density at the two genes did not change in vivo as monitored by ChIP.

Uncoupling human immunodeficiency virus type 1 Gag and Pol reading frames: role of the transframe protein p6* in viral replication.

Apart from its regulatory role in protease (PR) activation, little is known about the function of the human immunodeficiency virus type 1 transframe protein p6* in the virus life cycle. p6* is located between the nucleocapsid and PR domains in the Gag-Pol polyprotein precursor and is cleaved by PR during viral maturation. We have recently reported that the central region of p6* can be extensively mutated without abolishing viral infectivity and replication in vitro. However, mutagenesis of the entire p6*-coding sequence in the proviral context is not feasible without affecting the superimposed frameshift signal or the overlapping p1-p6(gag) sequences. To overcome these limitations, we created a novel NL4-3-derived provirus by displacing the original frameshift signal to the 3' end of the gag gene, thereby uncoupling the p6* gene sequence from the p1-p6(gag) reading frame. The resulting virus (AL) proved to be replication competent in different cell cultures and thus represents an elegant tool for detailed analysis of p6* function. Hence, extensive deletions or substitutions were introduced into the p6* gene sequence of the AL provirus, and effects on particle release, protein processing, and viral infectivity were evaluated. Interestingly, neither the deletion of 63% of all p6* residues nor the partial substitution by a heterologous sequence affected virus growth and infectivity, suggesting that p6* is widely dispensable for viral in vitro replication. However, the insertion of a larger reporter sequence interfered with virus production and maturation, implying that the length or conformation of this spacer region might be critical for p6* function.

Importance of protease cleavage sites within and flanking human immunodeficiency virus type 1 transframe protein p6* for spatiotemporal regulation of protease activation.

The human immunodeficiency virus type 1 (HIV-1) protease (PR) has recently been shown to be inhibited by its propeptide p6* in vitro. As p6* itself is a PR substrate, the primary goal of this study was to determine the importance of p6* cleavage for HIV-1 maturation and infectivity. For that purpose, short peptide variants mimicking proposed cleavage sites within and flanking p6* were designed and analyzed for qualitative and quantitative hydrolysis in vitro. Proviral clones comprising the selected cleavage site mutations were established and analyzed for Gag and Pol processing, virus maturation, and infectivity in cultured cells. Amino-terminal cleavage site mutation caused aberrant processing of nucleocapsid proteins and delayed replication kinetics. Blocking the internal cleavage site resulted in the utilization of a flanking site at a significantly decreased hydrolysis rate in vitro, which however did not affect Gag-Pol processing and viral replication. Although mutations blocking cleavage at the p6* carboxyl terminus yielded noninfectious virions exhibiting severe Gag processing defects, mutations retarding hydrolysis of this cleavage site neither seemed to impact viral infectivity and propagation in cultured cells nor seemed to interfere with overall maturation of released viruses. Interestingly, these mutants were shown to be clearly disadvantaged when challenged with wild-type virus in a dual competition assay. In sum, we conclude that p6* cleavage is absolutely essential to allow complete activation of the PR and subsequent processing of the viral precursors.

Clinical Trials Required to Assess Potential Benefits and Side Effects of Treatment of Patients With Anorexia Nervosa With Recombinant Human Leptin.

The core phenotype of anorexia nervosa (AN) comprises the age and stage dependent intertwining of both its primary and secondary (i.e., starvation induced) somatic and mental symptoms. Hypoleptinemia acts as a key trigger for the adaptation to starvation by affecting diverse brain regions including the reward system and by induction of alterations of the hypothalamus-pituitary-"target-organ" axes, e.g., resulting in amenorrhea as a characteristic symptom of AN. Particularly, the rat model activity-based anorexia (ABA) convincingly demonstrates the pivotal role of hypoleptinemia in the development of starvation-induced hyperactivity. STAT3 signaling in dopaminergic neurons in the ventral tegmental area (VTA) plays a crucial role in the transmission of the leptin signal in ABA. In patients with AN, an inverted U-shaped relationship has been observed between their serum leptin levels and physical activity. Albeit obese and therewith of a very different phenotype, humans diagnosed with rare congenital leptin deficiency have starvation like symptoms including hypothalamic amenorrhea in females. Over the past 20 years, such patients have been successfully treated with recombinant human (rh) leptin (metreleptin) within a compassionate use program. The extreme hunger of these patients subsides within hours upon initiation of treatment; substantial weight loss and menarche in females ensue after medium term treatment. In contrast, metreleptin had little effect in patients with multifactorial obesity. Small clinical trials have been conducted for hypothalamic amenorrhea and to increase bone mineral density, in which metreleptin proved beneficial. Up to now, metreleptin has not yet been used to treat patients with AN. Metreleptin has been approved by the FDA under strict regulations solely for the treatment of generalized lipodystrophy. The recent approval by the EMA may offer, for the first time, the possibility to treat extremely hyperactive patients with AN off-label. Furthermore, a potential dissection of hypoleptinemia-induced AN symptoms from the primary cognitions and behaviors of these patients could ensue. Accordingly, the aim of this article is to review the current state of the art of leptin in relation to AN to provide the theoretical basis for the initiation of clinical trials for treatment of this eating disorder.

Vitamin D and the Risk of Depression: A Causal Relationship? Findings from a Mendelian Randomization Study.

While observational studies show an association between 25(OH)vitamin D concentrations and depressive symptoms, intervention studies, which examine the preventive effects of vitamin D supplementation on the development of depression, are lacking. To estimate the role of lowered 25(OH)vitamin D concentrations in the etiology of depressive disorders, we conducted a two-sample Mendelian randomization (MR) study on depression, i.e., "depressive symptoms" (DS, n = 161,460) and "broad depression" (BD, n = 113,769 cases and 208,811 controls). Six single nucleotide polymorphisms (SNPs), which were genome-wide significantly associated with 25(OH)vitamin D concentrations in 79,366 subjects from the SUNLIGHT genome-wide association study (GWAS), were used as an instrumental variable. None of the six SNPs was associated with DS or BD (all p > 0.05). MR analysis revealed no causal effects of 25(OH)vitamin D concentration, either on DS (inverse variance weighted (IVW); b = 0.025, SE = 0.038, p = 0.52) or on BD (IVW; b = 0.020, SE = 0.012, p = 0.10). Sensitivity analyses confirmed that 25(OH)vitamin D concentrations were not significantly associated with DS or BD. The findings from this MR study indicate no causal relationship between vitamin D concentrations and depressive symptoms, or broad depression. Conflicting findings from observational studies might have resulted from residual confounding or reverse causation.

Virus-like particles-universal molecular toolboxes.

Virus-like particles (VLPs) are highly organised spheres that self-assemble from virus-derived structural antigens. These stable and versatile subviral particles possess excellent adjuvant properties capable of inducing innate and cognate immune responses. Commercialised VLP-based vaccines have been successful in protecting humans from hepatitis B virus (HBV) and human papillomavirus (HPV) infection and are currently explored for their potential to combat other infectious diseases and cancer. Much insight into VLP-mediated immune stimulation and optimised VLP design has been gained from human immunodeficiency virus (HIV)-derived VLPs presenting promising components of current AIDS vaccine approaches. Owing to their unique features, VLPs and virosomes, the in vitro-reconstituted VLP counterparts, have recently gained ground in the field of nanobiotechnology as organic templates for the development of new biomaterials.

High CRP Levels After Critical Illness are Associated With an Increased Risk of Rehospitalization.

PURPOSE: Chronic inflammation, even at subclinical levels, is associated with adverse long-term outcome. PATIENTS AND METHODS: In this prospective, observational study, 66 critically ill patients surviving to hospital discharge were included. C-reactive protein (CRP) levels were determined at hospital discharge, 1, 2, and 6 weeks after hospital discharge. All the patients were repeatedly screened for adverse events resulting in rehospitalization or death for 1.5 years. RESULTS: After hospital discharge, over two-thirds of the patients exhibited elevated CRP levels (>2.0 mg/L). During the first week, CRP decreased compared with hospital discharge (P < 0.001) but did not change after week 1 (P = 0.67). Age (P = 0.24), surgical status (P = 0.95), or sepsis (P = 0.77) did not influence the CRP course. The latter differed between patients with (n = 15) and without (n = 51) adverse events (P = 0.003). CRP levels of patients without adverse events persistently decreased after hospital discharge (P = 0.03), whereas those of patients with adverse events did not (P = 0.86) but rebounded early. CONCLUSIONS: Plasma CRP levels in critically ill patients decreased during the first week after hospital discharge but remained unchanged during the subsequent 5 weeks. Over two-thirds of the patients exhibited elevated CRP levels compatible with chronic sub-clinical inflammation. Persistently elevated CRP levels after hospital discharge are associated with higher risk of rehospitalization.

Bitter gourd reduces elevated fasting plasma glucose levels in an intervention study among prediabetics in Tanzania.

ETHNOPHARMACOLOGICAL RELEVANCE: Impaired glucose tolerance and diabetes mellitus have become major health issues even in non-industrialized countries. As access to clinical management is often poor, dietary interventions and alternative medicines are required. For bitter gourd, Momordica charantia L., antidiabetic properties have been claimed. AIM OF THE STUDY: The main objective of the intervention study was to assess antidiabetic effects of daily bitter gourd consumption of 2.5g powder over the course of eight weeks among prediabetic individuals. MATERIALS AND METHODS: In a randomized placebo-controlled single blinded clinical trial, 52 individuals with prediabetes were studied after consuming a bitter gourd or a cucumber juice. For reducing the impact of between subject differences in the study population, a crossover design was chosen with eight weeks for each study period and four weeks washout in between. Fasting plasma glucose was chosen as the primary outcome variable. RESULTS: Comparing the different exposures, the CROS analysis (t=-2.23, p=0.031, r=0.326) revealed a significant difference in the change of FPG of 0.31mmol/L (5.6mg/dL) with a trend (R2=0,42387). The number of 44 finally complete data sets achieved a power of 0.82, with a medium-to-large effect size (Cohen's d 0.62). The effect was also proven by a general linear mixed model (estimate 0.31; SE: 0.12; p: 0.01; 95%CI: 0.08; 0.54). Not all participants responded, but the higher the initial blood glucose levels were, the more pronounced the effect was. No serious adverse effects were observed. CONCLUSIONS: Bitter gourd supplementation appeared to have benefits in lowering elevated fasting plasma glucose in prediabetes. The findings should be replicated in other intervention studies to further investigate glucose lowering effects and the opportunity to use bitter gourd for dietary self-management, especially in places where access to professional medical care is not easily assured.

Momordica charantia and type 2 diabetes: from in vitro to human studies.

Type 2 diabetes is a growing health problem worldwide that is particularly severe in India and China. In these areas, bitter gourd (Momordica charantia) is a popular vegetable which is traditionally known to have health beneficial effects not only, but mainly, on diabetes. Bitter gourd could be a cheap possibility to help the poor in these and other countries to control their blood glucose levels. This review describes anti-diabetic effects of bitter gourd reported in the literature and discusses what still needs to be clarified for developing an evidence-based and safe use of the bitter gourd for diabetes. Analyses of bioactive compounds have shown that bitter gourd is rich in nutrients and phytochemicals of which some have anti-diabetic effects. Juices, powders, extracts, and isolated compounds have been tested in vitro and in vivo. Bitter gourd increases insulin secretion of the pancreas, decreases intestinal glucose uptake, and increases uptake and utilization of glucose in peripheral tissues. Although human studies with type 2 diabetics are weak in their design and/or results, some of the studies do indicate anti-diabetic effects in patients and safety for bitter gourd treatment in humans. In the future, well designed studies with rodents will help to understand what kind of bitter gourd variety, dosage, preparation, and duration of administration is optimal. Such results will help to design human studies which are necessary to prove the effectiveness of bitter gourd in patients.

Comprehensive analysis of interactions between the Src-associated protein in mitosis of 68 kDa and the human Src-homology 3 proteome.

The protein Sam68 is involved in many cellular processes such as cell-cycle regulation, RNA metabolism, or signal transduction. Sam68 comprises a central RNA-binding domain flanked by unstructured tails containing docking sites for signalling proteins including seven proline-rich sequences (denoted P0 to P6) as potential SH3-domain binding motifs. To comprehensively assess Sam68-SH3-interactions, we applied a phage-display screening of a library containing all approx. 300 human SH3 domains. Thereby we identified five new (from intersectin 2, the osteoclast stimulating factor OSF, nephrocystin, sorting nexin 9, and CIN85) and seven already known high-confidence Sam68-ligands (mainly from the Src-kinase family), as well as several lower-affinity binders. Interaction of the high-affinity Sam68-binders was confirmed in independent assays in vitro (phage-ELISA, GST-pull-down) and in vivo (FACS-based FRET-analysis with CFP- and YFP-tagged proteins). Fine-mapping analyses with peptides established P0, P3, P4, and P5 as exclusive docking-sites for SH3 domains, which showed varying preferences for these motifs. Mutational analyses identified individual residues within the proline-rich motifs being crucial for the interactions. Based on these data, we generated a Sam68-mutant incapable of interacting with SH3 domains any more, as subsequently demonstrated by FRET-analyses. In conclusion, we present a thorough characterization of Sam68's interplay with the SH3 proteome. The observed interaction between Sam68 and OSF complements the known Sam68-Src and OSF-Src interactions. Thus, we propose, that Sam68 functions as a classical scaffold protein in this context, assembling components of an osteoclast-specific signalling pathway.

Multiparameter RNA and codon optimization: a standardized tool to assess and enhance autologous mammalian gene expression.

Autologous expression of recombinant human proteins in human cells for biomedical research and product development is often hampered by low expression yields limiting subsequent structural and functional analyses. Following RNA and codon optimization, 50 candidate genes representing five classes of human proteins--transcription factors, ribosomal and polymerase subunits, protein kinases, membrane proteins and immunomodulators--all showed reliable, and 86% even elevated expression. Analysis of three representative examples showed no detrimental effect on protein solubility while unaltered functionality was demonstrated for JNK1, JNK3 and CDC2 using optimized constructs. Molecular analysis of a sequence-optimized transgene revealed positive effects at transcriptional, translational, and mRNA stability levels. Since improved expression was consistent in HEK293T, CHO and insect cells, it was not restricted to distinct mammalian cell systems. Additionally, optimized genes represent powerful tools in functional genomics, as demonstrated by the successful rescue of an siRNA-mediated knockdown using a sequence-optimized counterpart. This is the first large-scale study addressing the influence of multiparameter optimization on autologous human protein expression.

Influence of extended mutations of the HIV-1 transframe protein p6 on Nef-dependent viral replication and infectivity in vitro.

The HIV-1 transframe protein p6 known to modulate HIV-1 protease activation has been suggested to interact with the viral pathogenicity factor Nef. However, a potential interaction site in p6 has not been mapped so far. To evaluate effects of p6 modification on viral replication in light of Nef function, clustered substitutions were introduced into the central p6 region of the infectious provirus NL4-3 and virus growth and composition of the various mutants was analyzed in different cell cultures in the presence or absence of Nef. Whereas clustered p6 substitutions did neither affect particle incorporation of Nef, nor precursor maturation or viral infectivity, a simultaneous substitution of 40 of the total 56 p6 residues significantly diminished viral infectivity and replication in a Nef-independent manner. Furthermore, this extended modification was not capable of rescuing the negative effects of a transdominant Nef mutant on particle production suggesting that the proposed target for Nef interaction in Gag-Pol is located outside the modified p6 region. In sum these data strongly argue against a functional connection of the central p6 region and Nef during viral life cycle.