Associations between breastfeeding duration and adherence to complementary feeding recommendations in Scotland.

We aimed to describe how breastfeeding relates to adherence to complementary feeding (CF) recommendations, diet diversification and feeding skills development and whether sociodemographic factors explain any differences observed. The Scottish Maternal Infant and Nutrition Survey for infants aged 8-12 months collected breastfeeding history, CF practices, diet and sociodemographic data using a self-completion questionnaire. Non-healthful CF practices were starting CF < 6 months, any consumption of sugar-sweetened beverages (SSBs), sweet or salty snacks (treats) or unmodified cow's milk and regular consumption of commercial baby foods. Diet diversification and feeding skills were assessed by amount of self-feeding and number of food groups, meals and snacks eaten daily. Of the 2730 mothers, 20% were solely infant formula fed (IFF) and 48% continued breastfeeding ≥6 months. Compared to IFF babies, mothers who gave any breast milk ≥6 months were more likely to start CF ≥ 6 months compared to those IFF (66% vs. 37%) and less likely to give treats (15% vs. 45%), SSBs (11% vs. 20%) and commercial baby foods (31% vs. 53%). These associations remained highly significant (p < 0.001) even after sociodemographic factor adjustment. Despite starting CF later, infants breastfed ≥6 months ate the same number of food groups and meals as those IFF, were just as likely to self-feed purees and more likely to self-feed finger foods daily (87% vs. 81% p < 0.001). Mothers who breastfeed beyond 6 months adhere more to CF recommendations and start CF later compared to IFF, but their babies eat a similarly diverse diet and have similar feeding skills.

Does planning to mixed feed undermine breastfeeding?

Continued breastfeeding is important for infants' health, but it is unclear whether mixed feeding increases the risk of breastfeeding cessation. We aimed to explore associations of mixed feeding and lactation problems with early cessation of breastfeeding. We analysed data from mothers who completed the Scottish National Maternal and Infant Feeding Survey and had previously breastfed their infants. At age 8-12 weeks, mothers (N = 1974) reported their feeding history and intentions, lactation problems and reasons for giving formula milk. The main outcome measure was cessation of breastfeeding before 6-8 weeks and time to cessation. By 6 weeks, 65% had mixed fed at some point, 32% had ceased breastfeeding, 22% were currently mixed feeding and 46% were exclusively breastfeeding. Lactation problems before 2 weeks were common (65%), and strongly associated with stopping breastfeeding (relative risk [RR]: 3.23, 95% confidence interval [CI]: 2.0-5.3) and with mixed feeding (RR: 3.14, 95% CI: 2.5-4.0). However, even after adjustment for breastfeeding problems mothers who planned to mixed feed (RR: 3.39, 95% CI: 2.4-4.9) and those who introduced formula for practicalities (RR: 3.21, 95% CI: 2.3-4.4) were more likely to stop breastfeeding. These variables also predicted later lactation insufficiency (planned mixed feeding RR: 1.39, 95% CI: 1.0-2.0; formula for practicalities RR: 1.76, 95% CI: 1.3-2.3). Mothers who received specialist lactation support were less likely to cease breastfeeding (RR: 0.63, 95% CI: 0.5-0.9) but nonspecialist input was unrelated to risk of cessation (RR: 1.06, 95% CI: 0.2-4.9). In conclusion, choosing to mix feed an infant is strongly associated with stopping breastfeeding, even in the absence of lactation problems.

Unveiling Sticholysin II and plasmid DNA interaction: Implications for developing non-viral vectors.

Non-viral gene delivery systems offer significant potential for gene therapy due to their versatility, safety, and cost advantages over viral vectors. However, their effectiveness can be hindered by the challenge of efficiently releasing the genetic cargo from endosomes to prevent degradation in lysosomes. To overcome this obstacle, functional components can be incorporated into these systems. Sticholysin II (StII) is one of the pore-forming proteins derived from the sea anemone Stichodactyla helianthus, known for its high ability to permeabilize cellular and model membranes. In this study, we aimed to investigate the interaction between StII, and a model plasmid (pDNA) as an initial step towards designing an improved vector with enhanced endosomal escape capability. The electrophoretic mobility shift assay (EMSA) confirmed the formation of complexes between StII and pDNA. Computational predictions identified specific residues involved in the StII-DNA interaction interface, highlighting the importance of electrostatic interactions and hydrogen bonds in mediating the binding. Atomic force microscopy (AFM) of StII-pDNA complexes revealed the presence of nodular fiber and toroid shapes. These complexes were found to have a predominantly micrometer size, as confirmed by dynamic light scattering (DLS) measurements. Despite increase in the overall charge, the complexes formed at the evaluated nitrogen-to-phosphorus (N/P) ratios still maintained a negative charge. Moreover, StII retained its pore-forming capacity regardless of its binding to the complexes. These findings suggest that the potential ability of StII to permeabilize endosomal membranes could be largely maintained when combined with nucleic acid delivery systems. Additionally, the still remaining negative charge of the complexes would enable the association of another positively charged component to compact pDNA. However, to minimize non-specific cytotoxic effects, it is advisable to explore methods to regulate the protein's activity in response to the microenvironment.

PREVALENCE OF SARS-CoV-2 RNA AND ANTIBODY DETECTION, AND VACCINATION STATUS IN PATIENTS WITH OCULAR VASCULAR OCCLUSION.

PURPOSE: To analyze the annual prevalence of ocular vascular occlusion in relation to COVID-19 infection and vaccination status in a prospective study. METHODS: All patients were examined for an active severe acute respiratory syndrome coronavirus 2 infection by RNA detection and for a previous infection by virus-specific antibody detection, and their vaccination status was documented. Data from pandemic year 2020 and previous years, before COVID-19 (2019, 2018, 2017), were retrospectively analyzed. RESULTS: In 2021, a total of 103 patients with the first diagnosis of ocular vascular occlusion were treated. Most frequent subdiagnoses were central retinal vein occlusion (20.4%), nonarteritic anterior ischemic optic neuropathy (18.4%), central retinal artery occlusion (13.6%), and branch retinal artery occlusion (12.6%). Thereof, only three patients (2.9%) presented with virus-specific severe acute respiratory syndrome coronavirus 2 antibodies, and none was PCR positive. Patients with preceded severe acute respiratory syndrome coronavirus 2 vaccination (59.2%) presented with comparable characteristics as unvaccinated patients with vascular occlusion regarding age, gender distribution, systemic risk factors, duration of symptoms, visual acuity, and the present subdiagnoses ( P > 0.05). The total number of cases in 2021 (103 cases) was comparable with the pandemic year 2020, at which no vaccination was available (114 cases), and to earlier years 2017, 2018, and 2019 without COVID-19 pandemic (100, 120, and 119 cases). Furthermore, we did not reveal any differences between pandemic and reference years regarding patients' characteristics ( P > 0.05). CONCLUSION: Our study did not reveal an increased annual prevalence of ocular vascular occlusions during COVID-19 pandemic years 2020 and 2021. Patients with previous COVID-19 vaccination did not present differences regarding the risk profile nor symptoms, compared with unvaccinated individuals.

Bioavailability of orange juice (poly)phenols: ß-glucan-rich oat bran decreases urinary excretion of flavanone phase II metabolites and enhances excretion of microbiota-derived phenolic catabolites.

The impact of ß-glucan-rich oat bran on the bioavailability of orange juice (OJ) flavanones was investigated. Volunteers consumed 500 mL of OJ with and without 22 g of oat bran containing 6 g of ß-glucan (OB-6). Urine collected 12 h prior to and over a 0-24 h period post-supplementation was analysed by UHPLC-HRMS. Sixteen flavanone metabolites and thirty-nine colon-derived phenolic catabolites were identified and quantified. The major compounds were hesperetin-3'-glucuronide, along with hippuric acids and the C6-C3 phenolic acids 3-(3'-hydroxy-4'-methoxyphenyl)hydracrylic acid and 3-(4'-hydroxy-3'-methoxyphenyl)propanoic acid. A marked reduction in the 0-24 h excretion of flavanone metabolites from 29.7 µmol (9.3% recovery) to 9.3 µmol (2.9% recovery), occurred following consumption of OB-6 compared to OJ. This appeared not to be an effect of fiber on the rate of transport in the upper gut. After consumption of OJ there was a 163 ± 15 µmol excretion of colon-derived phenolic catabolites, equivalent to 43% of (poly)phenol intake and following OB-6 intake there was a further significant 30% increase. The ß-oat bran in OB-6 contained 5.8 µmol of free and 52 µmol of bound phenolic derivatives compared to 371 µmol of OJ (poly)phenols. The elevated excretion of phenolics after OB-6 consumption appears not to be due to bound phenolics in the bran, rather it is consequence, principally, of a bran-mediated increase in the quantities of flavanones passing from the upper to the lower bowel where they were subjected to microbiota-mediated catabolism. CLINICAL TRIAL REGISTRATION NUMBER: This trial was registered at clinicaltrials.gov as NCT04867655.

Biomarkers of Fibrosis in Patients with COVID-19 One Year After Hospital Discharge: A Prospective Cohort Study.

Beyond the acute infection of coronavirus disease (COVID-19), concern has arisen about long-term effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. The aim of our study was to analyze if there is any biomarker of fibrogenesis in patients with COVID-19 pneumonia capable of predicting post-COVID-19 pulmonary sequelae. We conducted a multicenter, prospective, observational cohort study of patients admitted to a hospital with bilateral COVID-19 pneumonia. We classified patients into two groups according to severity, and blood sampling to measure matrix metalloproteinase 1 (MMP-1), MMP-7, periostin, and VEGF and respiratory function tests and high-resolution computed tomography were performed at 2 and 12 months after hospital discharge. A total of 135 patients were evaluated at 12 months. Their median age was 61 (interquartile range, 19) years, and 58.5% were men. We found between-group differences in age, radiological involvement, length of hospital stay, and inflammatory laboratory parameters. Differences were found between 2 and 12 months in all functional tests, including improvements in predicted forced vital capacity (98.0% vs. 103.9%; P = 0.001) and DlCO <80% (60.9% vs. 39.7%; P = 0.001). At 12 months, 63% of patients had complete high-resolution computed tomography resolution, but fibrotic changes persisted in 29.4%. Biomarker analysis demonstrated differences at 2 months in periostin (0.8893 vs. 1.437 ng/ml; P < 0.001) and MMP-7 (8.7249 vs. 15.2181 ng/ml; P < 0.001). No differences were found at 12 months. In multivariable analysis, only 2-month periostin was associated with 12-month fibrotic changes (odds ratio, 1.0013; 95% confidence interval, 1.0006-1.00231; P = 0.003) and 12-month DlCO impairment (odds ratio, 1.0006; 95% confidence interval, 1.0000-1.0013; P = 0.047). Our data suggest that early periostin postdischarge could predict the presence of fibrotic pulmonary changes.

Association between Early Feeding Patterns and Neonatal Outcomes in Very Preterm Infants: A Retrospective Cohort Study.

OBJECTIVE: Mother's own milk (MOM) is the optimal feed for premature infants but may not always be sufficiently available. Alternative feeding includes donor human milk (DONOR), with or without fortification and preterm formula. This study evaluated the association between early feeding with exclusively and predominantly MOM (MAINLY-MOM) versus MOM supplemented with fortified DONOR (MOM + DONOR) or preterm formula (MOM + FORMULA) and in-hospital growth and neonatal morbidities. METHOD: This was a multicentre (n = 13 units) cohort study of infants born at <32 weeks' gestation. Data captured at the point of care were extracted from the UK National Neonatal Research Database. The study groups were defined based on feeding patterns within the first 2 weeks of life using predefined cut-offs. The primary outcome was the in-hospital growth rate. RESULTS: Data from 1,272 infants were analysed. Infants fell into two groups: extremely preterm (EPT) infants and very preterm (VPT) infants, born after <28 weeks and 28 to <32 weeks of gestation, respectively. Only 11 of 365 EPT infants received formula supplements, precluding a useful comparison of MOM + DONOR and MOM + FORMULA. There was no difference in median (25th-75th centile) growth velocity over the first 30 days of life between the MAINLY-MOM (n = 248) and MOM + DONOR (n = 106) groups: 10 (8-13) versus 10 (7-13) g/kg/day. Similarly, for VPT infants, there was no difference in growth velocities between MAINLY-MOM (n = 407), MOM + DONOR (N = 196), and MOM + FORMULA (N = 304): 11 (8-14) versus 11 (8-14) versus 11 (8-14) g/kg/day. Head growth did not differ (p value = 0.670). Cox regression analysis showed no difference in time to discharge between feeding types or any difference in major neonatal morbidities. In both EPT and VPT infants, growth velocity from the time of regaining birth weight to discharge was significantly lower in the MAINLY-MOM group compared to the MOM-DONOR group (EPT: 12.5 [11-14.2] vs. 14 [12.3-15.9] p = 0.45, VPT 13.5 [11-15.7] vs. 14.5 [12.6-16.8] p = 0.015). CONCLUSION: Early feeding with fortified DONOR, in comparison to formula, to supplement MOM was not associated with any differences in short-term growth, length of stay, and neonatal morbidities. However, early feeding with mainly maternal milk, compared to maternal milk supplemented with DONOR, was associated with significantly lower overall weight gain.

Down-Regulation-Resistant STAT4 Risk Haplotype Contributes to Lupus Nephritis Through CD4+ T Cell Interferon-γ Production.

OBJECTIVE: Variants in STAT4 are associated with systemic lupus erythematosus (SLE) and other autoimmune diseases. We undertook this study to investigate how disease-associated variants affect STAT4 expression, in particular in CD4+ T cells where STAT4 plays an essential role. METHODS: We compared Th1 differentiation between naive CD4+ T cells from healthy donors homozygous for the risk (R/R) or nonrisk (NR/NR) alleles. We analyzed epigenetic marks in STAT4 and evaluated the relevance of its third intron, assessed the consequences of Stat4 overexpression in vivo in mice, and analyzed the effects of the STAT4 genotype in patients with lupus nephritis. RESULTS: Naive CD4+ T cells from NR/NR healthy donors down-regulated STAT4 in response to interleukin-12 (IL-12). In contrast, cells from R/R healthy donors maintained high levels. R/R cells exhibited a higher abundance of transcriptionally active STAT4 and increased interferon-γ production. Accordingly, R/R healthy donors exhibited a stronger induction of local active enhancer marks. Genetic editing confirmed the presence of a negative regulatory region in the STAT4 third intron, where most of the SLE-associated STAT4 single-nucleotide polymorphisms (SNPs) are located. In vivo forced expression demonstrated that increases in Stat4 levels in T cells enhanced glomerulonephritis in mice. Accordingly, the R/R genotype was associated with suboptimal response to treatment and with worse clinical outcomes in patients with proliferative lupus nephritis. CONCLUSION: The SLE-associated STAT4 haplotype correlates with an abnormal IL-12-mediated STAT4 transcriptional regulation. Carriers of the risk variant exhibit exaggerated CD4+ proinflammatory capacities that, in the context of SLE, contribute to more severe disease. R/R patients may benefit from blockade of the IL-12/STAT4 pathway.

Strength of hip muscles and bradykinesia contribute to walking speed over a long distance, but not a short distance, in people with mild Parkinson's disease: An observational study.

BACKGROUND: Bradykinesia and postural instability contribute to walking limitations in people with Parkinson's disease (PD), but the contribution of muscle strength to walking speed has not been examined extensively. RESEARCH QUESTION: Does strength of the major lower limb muscles contribute to walking speed over short and long distances in people with PD? METHOD: na. DESIGN: A cross-sectional, observational study. PARTICIPANTS: 50 people with mild PD. OUTCOME MEASURES: Maximum isometric strength of the flexors and extensors of the lower limb was measured using hand-held dynamometry. Flexed posture, postural instability and bradykinesia were measured using the MDS-UPDRS part III. Walking speed was measured during the 10-m Walk Test and 6-min Walk Test. RESULTS: Univariate analysis revealed that age, bradykinesia, flexed posture, and hip, knee and ankle weakness were all significantly correlated with walking speed over short and long distances. Multiple regression analysis revealed that age, bradykinesia, flexed posture and weakness of the hip and knee muscles together accounted for 47% of the variance in walking speed over a short distance, with age (R2 = 0.11, p = 0.003) making a significant independent contribution. Age, bradykinesia, flexed posture and strength of the hip and knee muscles together accounted for 56% of the variance in walking speed over a long distance. Age (R2 = 0.12, p = 0.001), weakness of the hip muscles (R2 = 0.12, p = 0.001) and bradykinesia (R2 = 0.04, p = 0.04) made significant, independent contributions to the variance in walking speed over a long distance. SIGNIFICANCE: In people with mild PD, weakness of the hip muscles and bradykinesia only make a significant relative contribution to walking speed over a long distance. Therefore, in people with normal walking speed over a short distance the 6-min Walk Test could be measured to more effectively identify the impact of motor impairments on walking.

Lung function and radiological findings 1 year after COVID-19: a prospective follow-up.

BACKGROUND: The coronavirus disease (COVID-19) pandemic has already affected more than 400 million people, with increasing numbers of survivors. These data indicate that a myriad of people may be affected by pulmonary sequelae of the infection. The aim of this study was to evaluate pulmonary sequelae in patients with bilateral COVID-19 pneumonia according to severity 1 year after hospital discharge. METHODS: COVID-FIBROTIC is a multicenter prospective observational cohort study for admitted patients with bilateral COVID-19 pneumonia. Pulmonary functional outcomes and chest computed tomography sequelae were analyzed 12 months after hospital discharge and we classified patients into three groups according to severity. A post hoc analysis model was designed to establish how functional test changed between groups and over time. A multivariable logistic regression model was created to study prognostic factors for lung diffusion impairment and radiological fibrotic-like changes at 12 months. RESULTS: Among 488 hospitalized patients with COVID-19 pneumonia, 284 patients had completed the entire evaluation at 12 months. Median age was 60.5 ± 11.9 and 55.3% were men. We found between-group differences in male sex, length of hospital stay, radiological involvement and inflammatory laboratory parameters. The functional evaluation of pulmonary sequelae showed that severe patients had statistically worse levels of lung diffusion at 2 months but no between group differences were found in subsequent controls. At 12-month follow up, however, we found impaired lung diffusion in 39.8% unrelated to severity. Radiological fibrotic-like changes at 12 months were reported in 22.7% of patients (102/448), only associated with radiological involvement at admission (OR: 1.55, 95% CI 1.06-2.38; p = 0.02) and LDH (OR: 0.99, 95% CI 0.98-0.99; p = 0.046). CONCLUSION: Our data suggest that a significant percentage of individuals would develop pulmonary sequelae after COVID 19 pneumonia, regardless of severity of the acute process. Trial registration clinicaltrials.gov NCT04409275 (June 1, 2020).

Prevalence and overlap of known undernutrition risk factors in children in Nairobi Kenya.

We aimed to describe the co-occurrence of known risk factors for undernutrition and the prevalence of modifiable risks in wasted, stunted and healthy children. Quota sampling was used to recruit healthy [weight for age Z scores (WAZ) > -2 SD] and undernourished [weight for length (WLZ) or WAZ scores ≤ -2 SD] children aged 6-24 months from seven clinics in low-income areas of Nairobi. Structured interviews were used to identify exposure to socioeconomic, water and hygiene, infant feeding, dietary and behavioural risks (low interest in food, high food refusal and force feeding). We recruited 92 wasted WLZ ≤ -2 SD, 133 stunted (length for age Z scores LAZ ≤ -2 SD) and 172 healthy (LAZ and WLZ > 2SD) children. Nearly all children were exposed to hygiene risks (90%) and low dietary diversity (95%) regardless of nutritional status. Stunted children were more likely to be exposed to socio-economic risks (54% healthy, 64% wasted and 72% stunted; P = 0.001). Compared with healthy children, wasted and stunted children were more likely to be exposed to infant feeding (25% healthy, 40% wasted and 41% stunted; P = 0.02) and behaviour risks (24% healthy, 49% wasted, and 44% stunted; P = 0.004). Overall, wasted and stunted children were twice as likely to be exposed to more than three risks (23% healthy, 48% wasted, and 50% stunted; P = <0.001). They were also more likely to be exposed to more than three modifiable risks (dietary, handwashing and behaviour risks). Wasting and stunting are associated with exposure to multiple risk factors, many of which are potentially modifiable using targeted advice.

Acute effect of oat ß-glucan on the bioavailability of orange juice flavanones.

The impact of ß-glucan on the bioavailability of orange juice (OJ) flavanones was investigated in a randomised controlled trial. Volunteers consumed 500 mL of OJ without or with either 3 g (OB-3) or 6 g (OB-6) of ß-glucan. Urine samples, collected 12 h before and over a 0-24 h period post-supplementation, were analysed by high-performance liquid chromatography-high resolution mass spectrometry. The overall 0-24 h urinary excretion of the 17 flavanone metabolites identified and quantified in urine after OJ ingestion corresponded to 29.7 µmol, and 25.0 and 9.3 µmol, respectively, after OB-3 and OB-6 intake. This corresponds to 9.3, 7.9, and 2.9% recoveries of the 318 µmol of the ingested flavanones. The acute ingestion of OJ with 6 g, but not 3 g of ß-glucan led to a significant reduction (p < 0.05) in the excretion of flavanone metabolites compared with consumption of OJ alone.

Interactions between dietary fibre and the gut microbiota.

Research characterising the gut microbiota in different populations and diseases has mushroomed since the advent of next-generation sequencing techniques. However, there has been less emphasis on the impact of dietary fibres and other dietary components that influence gut microbial metabolic activities. Dietary fibres are the main energy source for gut bacteria. However, fibres differ in their physicochemical properties, their effects on the gut and their fermentation characteristics. The diversity of carbohydrates and associated molecules in fibre-rich foods can have a major influence on microbiota composition and production of bioactive molecules, for example SCFAs and phenolic acids. Several of these microbial metabolites may influence the functions of body systems including the gut, liver, adipose tissues and brain. Dietary fibre intake recommendations have recently been increased (to 30 g daily) in response to growing obesity and other health concerns. Increasing intakes of specific fibre and plant food sources may differentially influence the bacteria and their metabolism. However, in vitro studies show great individual variability in the response of the gut microbiota to different fibres and fibre combinations, making it difficult to predict which foods or food components will have the greatest impact on levels of bioactive molecules produced in the colon of individuals. Greater understanding of individual responses to manipulation of the diet, in relation to microbiome composition and production of metabolites with proven beneficial impact on body systems, would allow the personalised approach needed to best promote good health.

Development of a Scale to Measure Infant Eating Behaviour Worldwide.

In order to create a short, internationally valid scale to assess eating behaviour (EB) in young children at risk of undernutrition, we refined 15 phrases describing avidity or food refusal (avoidance). In study one, 149 parents matched phrases in English, Urdu, Cantonese, Indonesian or Greek to videos showing avidity and avoidance; 82-100% showed perfect agreement for the avidity phrases and 73-91% for the avoidant phrases. In study two, 575 parents in the UK, Cyprus and Indonesia (healthy) and in Kenya, Pakistan and Guatemala (healthy and undernourished) rated their 6-24 months old children using the same phrases. Internal consistency (Cronbach's α) was high for avidity (0.88) and moderate for avoidance (0.72). The best-performing 11 items were entered into a principal components analysis and the two scales loaded separately onto 2 factors with Eigen values > 1. The avidity score was positively associated with weight (r = 0.15 p = 0.001) and body mass index (BMI) Z scores (r = 0.16 p = 0.001). Both high and low avoidance were associated with lower weight and BMI Z scores. These scales are internationally valid, relate to nutritional status and can be used to inform causes and treatments of undernutrition worldwide.

Interphase epichromatin: last refuge for the 30-nm chromatin fiber?

"Interphase epichromatin" describes the surface of chromatin located adjacent to the interphase nuclear envelope. It was discovered in 2011 using a bivalent anti-nucleosome antibody (mAb PL2-6), now known to be directed against the nucleosome acidic patch. The molecular structure of interphase epichromatin is unknown, but is thought to be heterochromatic with a high density of "exposed" acidic patches. In the 1960s, transmission electron microscopy of fixed, dehydrated, sectioned, and stained inactive chromatin revealed "unit threads," frequently organized into parallel arrays at the nuclear envelope, which were interpreted as regular helices with ~ 30-nm center-to-center distance. Also observed in certain cell types, the nuclear envelope forms a "sandwich" around a layer of closely packed unit threads (ELCS, envelope-limited chromatin sheets). Discovery of the nucleosome in 1974 led to revised helical models of chromatin. But these models became very controversial and the existence of in situ 30-nm chromatin fibers has been challenged. Development of cryo-electron microscopy (Cryo-EM) gave hope that in situ chromatin fibers, devoid of artifacts, could be structurally defined. Combining a contrast-enhancing phase plate and cryo-electron tomography (Cryo-ET), it is now possible to visualize chromatin in a "close-to-native" situation. ELCS are particularly interesting to study by Cryo-ET. The chromatin sheet appears to have two layers of ~ 30-nm chromatin fibers arranged in a criss-crossed pattern. The chromatin in ELCS is continuous with adjacent interphase epichromatin. It appears that hydrated ~ 30-nm chromatin fibers are quite rare in most cells, possibly confined to interphase epichromatin at the nuclear envelope.

DNA Replication: An in vivo Space-Time Continuum in the Ciliate Replication Band.

The replication band in the macronucleus of ciliated protozoa has fascinated microscopists since the 19th Century. It migrates through the nucleus, corresponding to a region of DNA replication and nascent chromatin assembly. A new study shows that calcium and actin filaments may participate in the formation and migration of the replication band.

Men's Reflections on Their Experiences of Gender-Based Violence.

Using Multicultural and Gender Case Conceptualization (MGCC) epistemology, the aim of this phenomenological study was to explore adult male victims' perceptions of gender and reflections on their experiences of gender-based violence (GBV). Data were analyzed using interpretative phenomenological analysis (IPA). Results indicated that, regardless of sexual orientation, participants experienced verbal and physical abuse due to their feminine gender expression, beginning in primary school and continuing through university and into the workplace. To reduce victimization, the men developed a range of strategies, such as policing their gender and avoiding social situations. The experiences of violence affected the participants' social relationships, choice of occupation, self-esteem, and ability to trust. However, after entering university, heterosexual, gay, and bisexual participants interpreted their experiences differently, whereby heterosexual participants linked the abuse to gender, and gay and bisexual participants viewed it as homophobia. As well, gay and bisexual participants found respite in the gay and bisexual communities, while heterosexual participants remained silent, isolated, and afraid to discuss their experiences. Implications for future research are discussed.

A Nursery-Based Cooking Skills Programme with Parents and Children Reduced Food Fussiness and Increased Willingness to Try Vegetables: A Quasi-Experimental Study.

Children's fussy eating is associated with a reduced vegetable intake. This quasi-experimental study evaluated "Big Chef Little Chef" (BCLC), a nursery-based cooking skills programme aimed at reducing food fussiness and increasing willingness to try green vegetables by incorporating repeated exposure and sensory learning. Parent and child (3-5 years) dyads attended BCLC for four/1.5 h weekly sessions. A comparison group was recruited after BCLC completion and attended a single education session at week 1. A questionnaire measured food fussiness at week 1 and week 4. At week 4, all children were offered six green vegetables (raw and cooked) and an average score (1 = did not try; 2 = tried it/ate some; 3 = ate it all) was calculated for willingness to try vegetables. In total, 121 dyads (intervention: n = 64; comparison: n = 57) participated. The food fussiness score (1 min-5 max) in the intervention group decreased significantly from 3.0 to 2.6 (p < 0.01) between time points, while there was no change in the comparison group (3.1 (week 1) and 3.0 (week 4)). The intervention group was more willing to try green vegetables with significantly higher (p < 0.001) median scores for raw and cooked vegetables (2.5 for both) compared with the comparison group (2.0 and 1.7, respectively). The BCLC reduced food fussiness and increased willingness to try green vegetables.

Plasma pharmacokinetics of (poly)phenol metabolites and catabolites after ingestion of orange juice by endurance trained men.

The health benefits of orange juice (OJ) consumption are attributed in part to the circulating flavanone phase II metabolites and their microbial-derived ring fission phenolic catabolites. The present study investigated these compounds in the bloodstream after acute intake of 500 mL of OJ. Plasma samples obtained at 0, 1, 2, 3, 4, 5, 6, 7, 8 and 24 h after OJ intake were analysed by HPLC-HR-MS. Eleven flavanone metabolites and 36 phenolic catabolites were identified and quantified in plasma. The main metabolites were hesperetin-3'-sulfate with a peak plasma concentration (Cmax) of 80 nmol/L, followed by hesperetin-7-glucuronide (Cmax 24 nmol/L), hesperetin-3'-glucuronide (Cmax 18 nmol/L) and naringenin-7-glucuronide (Cmax 21 nmol/L). Among the main phenolic catabolites to increase in plasma after OJ consumption were 3'-methoxycinnamic acid-4'-sulfate (Cmax 19 nmol/L), 3-hydroxy-3-(3'-hydroxy-4'-methoxyphenyl)propanoic acid (Cmax 20 nmol/L), 3-(3'-hydroxy-4'-methoxyphenyl)propanoic acid (Cmax 19 nmol/L), 3-(4'-hydroxyphenyl)propanoic acid (Cmax 25 nmol/L), and 3-(phenyl)propanoic acid (Cmax 19 nmol/L), as well as substantial amounts of phenylacetic and hippuric acids. The comprehensive plasma pharmacokinetic profiles that were obtained are of value to the design of future ex vivo cell studies, aimed at elucidating the mechanisms underlying the potential health benefits of OJ consumption. CLINICAL TRIAL REGISTRATION NUMBER: This trial was registered at clinicaltrials.gov as NCT02627547.

Linker histone epitopes are hidden by in situ higher-order chromatin structure.

BACKGROUND: Histone H1 is the most mobile histone in the cell nucleus. Defining the positions of H1 on chromatin in situ, therefore, represents a challenge. Immunoprecipitation of formaldehyde-fixed and sonicated chromatin, followed by DNA sequencing (xChIP-seq), is traditionally the method for mapping histones onto DNA elements. But since sonication fragmentation precedes ChIP, there is a consequent loss of information about chromatin higher-order structure. Here, we present a new method, xxChIP-seq, employing antibody binding to fixed intact in situ chromatin, followed by extensive washing, a second fixation, sonication and immunoprecipitation. The second fixation is intended to prevent the loss of specifically bound antibody during washing and subsequent sonication and to prevent antibody shifting to epitopes revealed by the sonication process. In many respects, xxChIP-seq is comparable to immunostaining microscopy, which also involves interaction of the primary antibody with fixed and permeabilized intact cells. The only epitopes displayed after immunostaining are the "exposed" epitopes, not "hidden" by the fixation of chromatin higher-order structure. Comparison of immunoprecipitated fragments between xChIP-seq versus xxChIP-seq should indicate which epitopes become inaccessible with fixation and identify their associated DNA elements. RESULTS: We determined the genomic distribution of histone variants H1.2 and H1.5 in human myeloid leukemia cells HL-60/S4 and compared their epitope exposure by both xChIP-seq and xxChIP-seq, as well as high-resolution microscopy, illustrating the influences of preserved chromatin higher-order structure in situ. We found that xChIP and xxChIP H1 signals are in general negatively correlated, with differences being more pronounced near active regulatory regions. Among the intriguing observations, we find that transcription-related regions and histone PTMs (i.e., enhancers, promoters, CpG islands, H3K4me1, H3K4me3, H3K9ac, H3K27ac and H3K36me3) exhibit significant deficiencies (depletions) in H1.2 and H1.5 xxChIP-seq reads, compared to xChIP-seq. These observations suggest the existence of in situ transcription-related chromatin higher-order structures stabilized by formaldehyde. CONCLUSION: Comparison of H1 xxChIP-seq to H1 xChIP-seq allows the development of hypotheses on the chromosomal localization of (stabilized) higher-order structure, indicated by the generation of "hidden" H1 epitopes following formaldehyde crosslinking. Changes in H1 epitope exposure surrounding averaged chromosomal binding sites or epigenetic modifications can also indicate whether these sites have chromatin higher-order structure. For example, comparison between averaged active or inactive promoter regions suggests that both regions can acquire stabilized higher-order structure with hidden H1 epitopes. However, the H1 xChIP-seq comparison cannot define their differences. Application of the xxChIP-seq versus H1 xChIP-seq method is particularly relevant to chromatin-associated proteins, such as linker histones, that play dynamic roles in establishing chromatin higher-order structure.