Search

Ninna Hinchely Ebdrup,1– 3 Anders Hammerich Riis,4,5 Cecilia Høst Ramlau-Hansen,2 Bjørn Bay,1,6 Julie Lyngsø,7 Dorte Rytter,2 Marianne Johansson Jørgensen,4 Ulla Breth Knudsen1,3 1Department of Obstetrics and Gynecology, Fertility Clinic, Horsens Regional Hospital, Horsens, Denmark; 2Department of Public Health, Aarhus University, Aarhus, Denmark; 3Department of Clinical Medicine, Aarhus University, Aarhus, Denmark; 4Department of Research, Regional Hospital Horsens, Horsens, Denmark; 5Enversion A/S, Aarhus, Denmark; 6Maigaard Fertility Clinic, Aarhus, Denmark; 7Department of Obstetrics and Gynecology, Aarhus University Hospital, Aarhus, DenmarkCorrespondence: Ninna Hinchely Ebdrup, Department of Obstetrics and Gynecology, Fertility Clinic, Horsens Regional Hospital, Sundvej 30, Horsens, 8700, Denmark, Tel +45 28 47 21 11, Email ninebd@clin.au.dkPurpose: Infertility may affect somatic and mental health later in life. Nevertheless, health status before diagnosed infertility is sparsely studied in women. We aimed to describe healthcare use in primary and secondary care before a first infertility diagnosis and compare use between cases and controls.Materials and Methods: The case–control study was based on register data and used incidence density sampling. From the CROSS-TRACKS Cohort, we included women residing in the Horsens area in Denmark in 2012– 2018 (n = 54,175). Eligible women were aged 18– 40 years, nulliparous, and living in heterosexual relationships. Cases were women with a first infertility diagnosis in the Danish National Patient Registry (index date). Five controls were matched on age, birth year, and calendar time. Through linkage to Danish national health registries, we identified general practitioner (GP) attendance, paraclinical examinations, hospital contacts, diagnoses, and redeemed prescriptions. Healthcare use from one year to five years before index date was compared with conditional logistic regression.Results: We identified 711 cases and 3555 controls. At one year before index date, cases consulted their GP (odds ratio (OR) = 5.2, 95% confidence interval (CI): 3.2, 8.3) and visited hospital (OR = 1.2, 95% CI: 1.0, 1.4) and redeemed prescriptions (OR = 2.3 95% CI: 1.9, 2.7) more often compared to controls. Cases more often had blood and hemoglobin tests performed, redeemed more drugs related to genitourinary and hormonal diseases, and were more often diagnosed with endocrine and genitourinary diseases in the year before a first infertility diagnosis compared to controls. Cases and controls had comparable healthcare use from five years to one year before a first infertility diagnosis.Conclusion: Cases and controls had similar healthcare use from five years to one year before a first infertility diagnosis. However, cases had a higher healthcare use in the year preceding a first infertility diagnosis compared to controls.Keywords: behavior, health status, incidence density, fertility, preconception health, medically assisted reproduction

Ninna Hinchely Ebdrup,1– 3 Ulla Breth Knudsen,1,3 Jörg Schullehner,2,4 Linn Håkonsen Arendt,2,5 Zeyan Liew,6,7 Julie Lyngsø,2,5 Bjørn Bay,1,8 Pernille Jul Clemmensen,2 Torben Sigsgaard,2,9,10 Birgitte Hansen,4 Cecilia Høst Ramlau-Hansen2 1Department of Obstetrics and Gynecology, Horsens Regional Hospital, Horsens, Denmark; 2Department of Public Health, Aarhus University, Aarhus, Denmark; 3Department of Clinical Medicine, Aarhus University, Aarhus, Denmark; 4Geological Survey of Denmark and Greenland, Aarhus, Denmark; 5Department of Obstetrics and Gynecology, Aarhus University Hospital, Aarhus, Denmark; 6Department of Environmental Health Sciences, Yale School of Public Health, New Haven, CT, USA; 7Yale Center for Perinatal, Pediatric, and Environmental Epidemiology, Yale School of Public Health, New Haven, CT, USA; 8Maigaard Fertility Clinic, Aarhus, Denmark; 9Centre for Integrated Register-based Research at Aarhus University, Aarhus, Denmark; 10Danish Big Data Centre for Environment and Health (BERTHA), Aarhus University, Aarhus, DenmarkCorrespondence: Ninna Hinchely Ebdrup, Department of Obstetrics and Gynecology, Horsens Regional Hospital, Regionshospitalet Horsens, Sundvej 30, Horsens, 8700, Denmark, Tel +4528472111, Email ninebd@rm.dkPurpose: No studies have investigated if drinking water nitrate affects human fecundity. Experimental studies point at detrimental effects on fetal development and on female and male reproduction. This cohort study aimed to explore if female and male preconception and long-term exposure to nitrate in drinking water was associated with fecundability measured as time to pregnancy (TTP) or use of medically assisted reproduction (MAR) treatment.Methods: The study population consisted of pregnant women recruited in their first trimester in 1996– 2002 to the Danish National Birth Cohort. Preconception drinking-water nitrate exposure was estimated for the pregnant women (89,109 pregnancies), and long-term drinking water nitrate exposure was estimated from adolescence to conception for the pregnant women (77,474 pregnancies) and their male partners (62,000 pregnancies) by linkage to the national drinking water quality-monitoring database Jupiter. Difference in risk of TTP > 12 months or use of MAR treatment between five exposure categories and log-transformed continuous models of preconception and long-term nitrate in drinking water were estimated. Binominal regression models for risk ratios (RR) were adjusted for age, occupation, education, population density, and lifestyle factors.Results: Nitrate in drinking water (median preconception exposure: 1.9 mg/L; median long-term exposure: 3.3 mg/L) was not associated with TTP > 12 months or use of MAR treatment, neither in the categorical nor in the continuous models.Conclusion: We found no association between preconception or long-term exposure to drinking water nitrate and fecundability.Keywords: environmental pollutant, subfecundity, time to pregnancy, public health, groundwater

Julie Lyngsø,1 Cecilia Høst Ramlau-Hansen,1 Bjørn Bay,2 Hans Jakob Ingerslev,3 Adam Hulman,1,4 Ulrik Schiøler Kesmodel5 1Department of Public Health, Section for Epidemiology, Aarhus University, Aarhus, 2The Fertility Clinic, Regional Horsens Hospital, Horsens, 3Department of Obstetrics and Gynaecology, Aarhus University Hospital, Aarhus, 4Danish Diabetes Academy, Odense, 5Department of Obstetrics and Gynaecology, Herlev and Gentofte Hospital, Herlev, Denmark Objective: The aim was to investigate whether coffee or caffeine consumption is associated with reproductive endpoints among women with natural fertility (ie, time to pregnancy [TTP] and spontaneous abortion [SAB]) and among women in fertility treatment (ie, clinical pregnancy rate or live birth rate). Design: This study was a systematic review and dose–response meta-analysis including data from case–control and cohort studies. Methods: An extensive literature search was conducted in MEDLINE and Embase, with no time and language restrictions. Also, reference lists were searched manually. Two independent reviewers assessed the manuscript quality using the Newcastle–Ottawa Scale (NOS). A two-stage dose–response meta-analysis was applied to assess a potential association between coffee/caffeine consumption and the outcomes: TTP, SAB, clinical pregnancy, and live birth. Heterogeneity between studies was assessed using Cochrane Q-test and I2 statistics. Publication bias was assessed using Egger’s regression test. Results: The pooled results showed that coffee/caffeine consumption is associated with a significantly increased risk of SAB for 300 mg caffeine/day (relative risk [RR]: 1.37, 95% confidence interval [95% CI]: 1.19; 1.57) and for 600 mg caffeine/day (RR: 2.32, 95% CI: 1.62; 3.31). No association was found between coffee/caffeine consumption and outcomes of fertility treatment (based on two studies). No clear association was found between exposure to coffee/caffeine and natural fertility as measured by fecundability odds ratio (based on three studies) or waiting TTP (based on two studies). Conclusion: Results from this meta-analysis support the growing evidence of an association between coffee/caffeine intake and the risk of SAB. However, viewing the reproductive capacity in a broader perspective, there seems to be little, if any, association between coffee/caffeine consumption and fecundity. In general, results from this study are supportive of a precautionary principle advised by health organizations such as European Food Safety Authority (EFSA) and World Health Organization (WHO), although the advised limit of a maximum of two to three cups of coffee/200–300 mg caffeine per day may be too high. Keywords: coffee, caffeine, fecundity, fertility, spontaneous abortion, assisted reproduction

Background: Assisted reproductive techniques are associated with an increased risk of adverse pregnancy outcomes, including low birthweight and intrauterine growth restriction. Yet, the long-term follow-up on the growth of these children is limited.Objective: To systematically review the literature on post-neonatal height and weight among children conceived following in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) treatment, compared with that of children born after spontaneous conception.Search Strategy: A systematic computerised literature search using the online databases PubMed, Embase, and Scopus.Selection Criteria: Cohort or case-control studies with an exposed group of singletons conceived following IVF or ICSI along with a control group of spontaneously conceived singletons.Data Collection and Analysis: Studies were reviewed by at least two authors. Meta-analyses were conducted using Cochrane Review Manager. The quality of the studies was assessed with the Newcastle-Ottawa Scale.Main Results: Twenty studies were included, with 13 of these eligible for meta-analyses. The meta-analyses compared 3972 children born after IVF/ICSI with 11 012 spontaneously conceived children and revealed no statistically significant difference in child weight [mean difference (MD) in weight of -160 g; 95% confidence interval (95% CI) -360, 3]. When stratifying by age of child at follow-up, we found a significant lower weight in children aged 0-4 years conceived following IVF/ICSI treatment (MD -180 g; 95% CI -320, -4), but this was no longer significant in children from 5 years of age (MD -160 g; 95% CI -580, 260). The pooled analysis revealed no statistically significant difference in childhood height.Conclusions: In vitro fertilisation/ICSI was not associated with long-term weight and height.Tweetable Abstract: Children born following IVF/ICSI do not have impaired long-term weight or height compared with spontaneously conceived children. [ABSTRACT FROM AUTHOR]

Many bacteria encode multiple toxin-antitoxin (TA) systems targeting separate, but closely related, cellular functions. The toxin of the Escherichia coli hipBA system, HipA, is a kinase that inhibits translation via phosphorylation of glutamyl-tRNA synthetase. Enteropathogenic E. coli O127:H6 encodes the hipBA -like, tripartite TA system; hipBST , in which the HipT toxin specifically targets the tryptophanyl-tRNA synthetase, TrpS. Notably, in the tripartite system, the function as antitoxin has been taken over by the third protein, HipS, but the molecular details of how activity of HipT is inhibited remain poorly understood. Here, we show that HipBST is structurally different from E. coli HipBA and that the unique HipS protein, which is homologous to the N-terminal subdomain of HipA, inhibits the kinase through insertion of a conserved Trp residue into the active site. We also show how auto-phosphorylation at two conserved sites in the kinase toxin serve different roles and affect the ability of HipS to neutralize HipT. Finally, solution structural studies show how phosphorylation affects overall TA complex flexibility., Competing Interests: RB, SN, RS, JL, FB, JP, KG, MS, DB No competing interests declared, (© 2023, Bærentsen, Nielsen et al.)

Background: Nitrate contamination is seen in drinking water worldwide. Nitrate may pass the placental barrier. Despite suggestive evidence of fetal harm, the potential association between nitrate exposure from drinking water and pregnancy loss remains to be studied. We aimed to investigate if nitrate in drinking water was associated with the risk of pregnancy loss., Methods: We conducted a nationwide cohort study of 100,410 pregnancies (enrolled around gestational week 11) in the Danish National Birth Cohort (DNBC) during 1996-2002. Spontaneous pregnancy losses before gestational week 22 were ascertained from the Danish National Patient Registry and DNBC pregnancy interviews. Using the national drinking water quality-monitoring database Jupiter, we estimated the individual and time-specific nitrate exposure by linking geocoded maternal residential addresses with water supply areas. The nitrate exposure was analyzed in spline models using a log-transformed continuous level or classified into five categories. We used Cox proportional hazards models to estimate associations between nitrate and pregnancy loss and used gestational age (days) as the time scale, adjusting for demographic, health, and lifestyle variables., Results: No consistent associations were found when investigating the exposure as a categorical variable and null findings were also found in trimester specific analyses. In the spline model using the continuous exposure variable, a modestly increased hazard of pregnancy loss was observed for the first trimester at nitrate exposures between 1 and 10 mg/L, with the highest. adjusted hazard ratio at 5 mg/L of nitrate of 1.16 (95% CI: 1.01, 1.34). This trend was attenuated in the higher exposure ranges., Conclusion: No association was seen between drinking water nitrate and the risk of pregnancy loss when investigating the exposure as a categorical variable. When we modelled the exposure as a continuous variable, a dose-dependent association was found between drinking water nitrate exposure in the first trimester and the risk of pregnancy loss. Very early pregnancy losses were not considered in this study, and whether survival bias influenced the results should be further explored., (© 2022. The Author(s).)

The self-assembly in mixtures of the anionic bile salt surfactant sodium deoxycholate (NaDC) and the zwitterionic phospholipid 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) in physiological saline solution has been investigated using light scattering, small-angle X-ray scattering and cryo-transmission electron microscopy. Rather small tri-axial ellipsoidal NaDC-DMPC mixed micelles form at a high content of bile salt in the mixture, which increase in size as an increasing amount of DMPC is incorporated into the micelles. Eventually, the micelles begin to grow substantially in length to form long wormlike micelles. At higher mole fractions of DMPC, the samples become turbid and cryo-TEM measurements reveal the existence of large perforated vesicles (stomatosomes), coexisting with geometrically open disks. To our knowledge, stomatosomes have not been observed before for any bile salt-phospholipid system. Mixed micelles are found to be the sole aggregate structure in a very wide regime of bile salt-phospholipid compositions, i.e. up to about 77 mol% phospholipid in the micelles. This is much higher than the corresponding value of 25 mol% observed for the conventional surfactant hexadecyltrimethylammonium bromide (CTAB) mixed with DMPC in the same solvent. The enhanced ability of bile salt surfactants to solubilize phospholipid bilayers and form mixed micelles is rationalized using bending elasticity theory. From our theoretical analysis, we are able to conclude that amphiphilic molecules rank in the following order of increasing spontaneous curvature: phospholipids < conventional surfactants < bile salts. The bending rigidity of the different amphiphilic molecules increases according to the following sequence: bile salts < conventional surfactants < phospholipids.

Fabry disease is a rare lysosomal storage disorder characterized by a deficiency of α-galactosidase A (GLA), a lysosomal hydrolase. The enzyme replacement therapy administering naked GLA shows several drawbacks including poor biodistribution, limited efficacy, and relatively high immunogenicity in Fabry patients. An attractive strategy to overcome these problems is the use of nanocarriers for encapsulating the enzyme. Nanoliposomes functionalized with RGD peptide have already emerged as a good platform to protect and deliver GLA to endothelial cells. However, low colloidal stability and limited enzyme entrapment efficiency could hinder the further pharmaceutical development and the clinical translation of these nanoformulations. Herein, the incorporation of the cationic miristalkonium chloride (MKC) surfactant to RGD nanovesicles is explored, comparing two different nanosystems-quatsomes and hybrid liposomes. In both systems, the positive surface charge introduced by MKC promotes electrostatic interactions between the enzyme and the nanovesicles, improving the loading capacity and colloidal stability. The presence of high MKC content in quatsomes practically abolishes GLA enzymatic activity, while low concentrations of the surfactant in hybrid liposomes stabilize the enzyme without compromising its activity. Moreover, hybrid liposomes show improved efficacy in cell cultures and a good in vitro/in vivo safety profile, ensuring their future preclinical and clinical development.

Introduction: Despite smoking being a well-established risk factor for adverse pregnancy and neonatal outcomes, a substantial proportion of women of reproductive age smoke. Previously, meta-analyses have indicated a significantly negative impact of female smoking on outcomes of assisted reproduction, yet most of the included studies have several, essential methodological limitations. We aimed to investigate whether female cigarette smoking may affect the chance of achieving a clinical pregnancy and live birth among women and couples receiving medically assisted reproduction treatment., Material and Methods: A cohort study with longitudinally and repeatedly collected exposure information from 1 January 2010 to 31 August 2015, including data on 1708 women and potential partners initiating either intrauterine insemination, in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) or frozen embryo transfer treatment cycles at the public Fertility Clinic, Aarhus University Hospital, Denmark. Smoking was assessed from self-reported questionnaires completed before treatment. Outcomes were a clinical pregnancy and a live birth. Information on these was obtained from the Danish national health registries, allowing complete follow-up. To evaluate associations between female occasional/daily cigarette smoking and successful medically assisted reproduction treatments, a modified Poisson regression with robust standard errors was used., Results: Female occasional/daily cigarette smoking was not associated with the chance of achieving a clinical pregnancy or a live birth in all intrauterine insemination or IVF/ICSI treatment cycles. When compared with nonsmokers, the adjusted relative risk for obtaining a live birth for those reporting smoking was 1.22 (0.70-2.12) among women initiating 1456 intrauterine insemination treatment cycles. Among women initiating 2788 IVF/ICSI treatment cycles, those reporting occasional/daily smoking had a relative risk for obtaining a live birth of 1.15 (0.82-1.60) when compared with nonsmokers., Conclusions: Occasionally/daily cigarette smoking women had similar chance of achieving a clinical pregnancy or a live birth as the nonsmokers when receiving medically assisted reproduction treatments. However, tobacco use before and during pregnancy remains a major cause of reduced fertility as well as maternal, fetal, and infant morbidity and mortality, and should strongly be discouraged., (© 2020 Nordic Federation of Societies of Obstetrics and Gynecology (NFOG). Published by John Wiley & Sons Ltd.)

Atherosclerosis is the main killer in the west and therefore a major health challenge today. Total serum cholesterol and lipoprotein concentrations, used as clinical markers, fail to predict the majority of cases, especially between the risk scale extremes, due to the high complexity in lipoprotein structure and composition. In particular, low-density lipoprotein (LDL) plays a key role in atherosclerosis development, with LDL size being a parameter considered for determining the risk for cardiovascular diseases. Determining LDL size and structural parameters is challenging to address experimentally under physiological-like conditions. This article describes the biochemistry and ultrastructure of normolipidemic and hypertriglyceridemic LDL fractions and subfractions using small-angle X-ray scattering. Our results conclude that LDL particles of hypertriglyceridemic compared to healthy individuals 1) have lower LDL core melting temperature, 2) have lower cholesteryl ester ordering in their core, 3) are smaller, rounder and more spherical below melting temperature, and 4) their protein-containing shell is thinner above melting temperature., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Human α 2 -macroglobulin (A2M) is the most characterized protease inhibitor in the alpha-macroglobulin (αM) superfamily, but the structure of its native conformation has not been determined. Here, we combined negative stain electron microscopy (EM), small-angle X-ray scattering (SAXS), and cross-linking-mass spectrometry (XL-MS) to investigate native A2M and its collapsed conformations that are obtained through aminolysis of its thiol ester by methylamine or cleavage of its bait region by trypsin. The combined interpretation of these data resulted in a model of the native A2M tetramer and its conformational changes. Native A2M consists of two crescent-shaped disulfide-bridged subunit dimers, which face toward each other and surround a central hollow space. In native A2M, interactions across the disulfide-bridged dimers are minimal, with a single major interface between the linker (LNK) regions of oppositely positioned subunits. Bait region cleavage induces both intrasubunit domain repositioning and an altered configuration of the disulfide-bridged dimer. These changes collapse the tetramer into a more compact conformation, which encloses an interior protease-trapping cavity. A recombinant A2M with a modified bait region was used to map the bait region's position in native A2M by XL-MS. A second recombinant A2M introduced an intersubunit disulfide into the LNK region, demonstrating the predicted interactions between these regions in native A2M. Altogether, our native A2M model provides a structural foundation for understanding A2M's protease-trapping mechanism, its conformation-dependent receptor interactions, and the dissociation of native A2M into dimers due to inflammatory oxidative stress., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Leucocyte common antigen-related protein (LAR) is a post-synaptic type I transmembrane receptor protein that is important for neuronal functionality and is genetically coupled to neuronal disorders such as attention deficit hyperactivity disorder (ADHD). To understand the molecular function of LAR, structural and biochemical studies of protein fragments derived from the ectodomain of human LAR have been performed. The crystal structure of a fragment encompassing the first four FNIII domains (LAR FN1-4 ) showed a characteristic L shape. SAXS data suggested limited flexibility within LAR FN1-4 , while rigid-body refinement of the SAXS data using the X-ray-derived atomic model showed a smaller angle between the domains defining the L shape compared with the crystal structure. The capabilities of the individual LAR fragments to interact with heparin was examined using microscale thermophoresis and heparin-affinity chromatography. The results showed that the three N-terminal immunoglobulin domains (LAR Ig1-3 ) and the four C-terminal FNIII domains (LAR FN5-8 ) both bound heparin, while LAR FN1-4 did not. The low-molecular-weight heparin drug Innohep induced a shift in hydrodynamic volume as assessed by size-exclusion chromatography of LAR Ig1-3 and LAR FN5-8 , while the chemically defined pentameric heparin drug Arixtra did not. Together, the presented results suggest the presence of an additional heparin-binding site in human LAR.

Protein fibrillation is traditionally associated with misfolding, loss of functional phenotype, and gain of toxicity in neurodegenerative diseases. However, many organisms exploit fibrils in the form of functional amyloids (FA), as seen in bacteria, such as E. coli, Salmonella, Bacillus, and Pseudomonas. Here, we provide structural information and mechanistic data for fibrillation of the smallest amyloidogenic truncation unit along with the full-length version (FL) of the major amyloid protein FapC from Pseudomonas, predicted to consist of three β-hairpin-forming imperfect repeats separated by disordered regions. Using a series of truncation mutants, we establish that the putative loops (linkers) increase the rate of aggregation. The minimal aggregation unit consisting of a single repeat with flanking disordered regions (R3C) aggregates in a pathway dominated by secondary nucleation, in contrast to the primary nucleation favored by full-length (FL) FapC. SAXS on FapC FL, R3C, and remaining truncation constructs resolves two major coexisting species in the fibrillation process, namely pre-fibrillar loosely aggregated monomers, and cylindrical, elliptical cross-section fibrils. Solid-state NMR spectra identified rigid parts of the FapC fibril. We assigned Cα-Cβ chemical shifts, indicative of a predominant β-sheet topology with some α-helix or loop chemical shifts. Our work emphasizes the complex nature of FapC fibrillation. In addition, we are able to deduce the importance of non-repeat regions (i.e., predicted loops), which enhance the amyloid protein aggregation and their influence on the polymorphism of the fibril architecture., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Anionic surfactants denature proteins at low millimolar concentrations, yet little is known about the underlying molecular mechanisms. Here, we undertake 1-μs-long atomistic molecular dynamics simulations of the denaturation of acyl coenzyme A binding protein (ACBP) and compare our results with previously published and new experimental data. Since increasing surfactant chain length is known to lead to more rapid denaturation, we studied denaturation using both the medium-length alkyl chain surfactant sodium dodecyl sulfate (SDS) and the long alkyl chain surfactant sodium hexadecyl sulfate (SHS). In silico denaturation on the microsecond timescale was not achieved using preformed surfactant micelles but required ACBP to be exposed to monomeric surfactant molecules. Micellar self-assembly occurred together with protein denaturation. To validate our analyses, we calculated small-angle X-ray scattering spectra of snapshots from the simulations. These agreed well with experimental equilibrium spectra recorded on ACBP-SDS mixtures with similar compositions. Protein denaturation occurs through the binding of partial micelles to multiple preferred binding sites followed by the accretion of surfactant monomers until these partial micelles merge to form a mature micelle and the protein chain is left disordered on the surface of the micelle. While the two surfactants attack in a similar fashion, SHS's longer alkyl chain leads to a more efficient denaturation through the formation of larger clusters that attack ACBP, a more rapid drop in native contacts, a greater expansion in size, as well as a more thorough rearrangement of hydrogen bonds and disruption of helices., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Interactions between proteins and surfactants are of relevance in many applications including food, washing powder formulations, and drug formulation. The anionic surfactant sodium dodecyl sulfate (SDS) is known to unfold globular proteins, while the non-ionic surfactant octaethyleneglycol monododecyl ether (C 12 E 8 ) can be used to refold proteins from their SDS-denatured state. While unfolding have been studied in detail at the protein level, a complete picture of the interplay between protein and surfactant in these processes is lacking. This gap in our knowledge is addressed in the current work, using the β-sheet-rich globular protein β-lactoglobulin (bLG). We combined stopped-flow time-resolved SAXS, fluorescence, and circular dichroism, respectively, to provide an unprecedented in-depth picture of the different steps involved in both protein unfolding and refolding in the presence of SDS and C 12 E 8 . During unfolding, core-shell bLG-SDS complexes were formed within ∼10 ms. This involved an initial rapid process where protein and SDS formed aggregates, followed by two slower processes, where the complexes first disaggregated into single protein structures situated asymmetrically on the SDS micelles, followed by isotropic redistribution of the protein. Refolding kinetics (>100 s) were slower than unfolding (<30 s), and involved rearrangements within the mixing deadtime (∼5 ms) and transient accumulation of unfolded monomeric protein, differing in structure from the original bLG-SDS structure. Refolding of bLG involved two steps: extraction of most of the SDS from the complexes followed by protein refolding. These results reveal that surfactant-mediated unfolding and refolding of proteins are complex processes with rearrangements occurring on time scales from sub-milliseconds to minutes., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Rheumatoid arthritis (RA) is a common autoimmune disease, which is characterized by painful chronic inflammation in the joints, and novel safe and efficacious treatments are urgently needed. RNA interference (RNAi) therapy based on small interfering RNA (siRNA) is a promising approach for silencing specific genes involved in inflammation. However, delivery of siRNA to the target site, i.e. the cytosol of immune cells, is a challenge. Here, we designed lipid-polymer hybrid nanoparticles (LPNs) composed of lipidoid and poly(DL-lactic-co-glycolic acid) loaded with a therapeutic cargo siRNA directed against the proinflammatory cytokine tumor necrosis factor (TNF), which plays a key role in the progression of RA. We compared their efficacy and safety with reference lipidoid-based stable nucleic acid lipid particles (SNALPs) in vitro and in vivo. Cryogenic transmission electron microscopy, atomic force microscopy and small-angle X-ray scattering revealed that the mode of loading of siRNA in lamellar structures differs between the two formulations. Thus, siRNA was tightly packed in LPNs, while LPNs displayed lower adhesion than SNALPs. The LPNs mediated a higher TNF silencing effect in vitro than SNALPs in the RAW 264.7 macrophage cell line activated with lipopolysaccharide. For both types of delivery systems, macropinocytosis was involved in cellular uptake. In addition, clathrin-mediated endocytosis contributed to uptake of SNALPs. LPNs loaded with TNF siRNA mediated sequence-specific suppression of inflammation in a murine experimental arthritis model upon intra-articular administration. Hence, the present study demonstrates that LPN-mediated TNF knockdown constitutes a promising approach for arthritis therapy of TNF-mediated chronic inflammatory conditions., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objective: To investigate whether female coffee consumption affects the chance of achieving a clinical pregnancy and a live birth among women and couples receiving medically assisted reproduction (MAR) treatment., Design: Cohort study with prospectively collected exposure data., Setting: Public fertility clinic., Patient(s): A total of 1,708 women and potential partners undergoing fertility treatment, contributing with 1,511 intrauterine insemination (IUI) cycles, 2,870 in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) cycles, and 1,355 frozen embryo transfer cycles., Intervention(s): None., Main Outcome Measure(s): Clinical pregnancy and live birth in consecutive treatment cycles in the Danish national health registries, enabling complete follow-up, and estimation of the cumulative chance of live birth for three consecutive treatment cycles., Result(s): Among women receiving IVF or ICSI treatment, coffee consumption did not seem to affect the chance of achieving a clinical pregnancy and a live birth. Women treated with IUI who had a daily coffee consumption of 1-5 cups were more likely to achieve a clinical pregnancy (adjusted relative risk 1.49; 95% confidence interval, 1.05-2.11) and live birth (adjusted relative risk 1.53; 95% confidence interval, 1.06-2.21) compared with the reference group of coffee abstainers., Conclusion(s): Women consuming 1-5 cups versus none had a 1.5-fold higher probability of achieving a pregnancy or a live birth when receiving IUI. No associations were found, however, between women's daily coffee consumption and achieving a pregnancy or a live birth from IVF/ICSI., (Copyright © 2019 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Treatment of polymicrobial infections requires combination therapy with drugs that have different antimicrobial spectra and possibly work in synergy. However, the different pharmacokinetics and adverse side effects challenge the simultaneous delivery of multiple drugs at the appropriate concentrations to the site of infection. Formulation of multiple drugs in nano-carrier systems may improve therapeutic efficacy by increasing the local concentration and lowering the systemic concentration, leading to fewer side effects. In this study, we loaded polymyxin B and vancomycin on bare and carboxyl-modified mesoporous silica nanoparticles (B-MSNs and C-MSNs, respectively) to achieve simulataneous local delivery of antibiotics against Gram-positive and -negative bacteria. Polymyxin B adsorbed preferentially to nanoparticles compared to vancomycin. The total antibiotic loading was 563 μg and 453 μg per mg B-MSNs or C-MSNs, respectively. Both B-MSNs and C-MSNs loaded with antibiotics were effective against Gram-negative and Gram-positive bacteria. The antibiotics had synergistic interactions against Gram-negative bacteria, and the antimicrobial efficacy was higher for antibiotic-loaded C-MSNs compared to free antibiotics at the same concentration even though the cytotoxicity was lower. Our study shows that formulations of existing antibiotics in nanocarrier systems can improve their therapeutic efficiency, indicating that combination therapy with drug-loaded silica nanoparticles may provide a better treatment outcome for infections that require high concentrations of multiple drugs., (Copyright © 2018 Elsevier B.V. All rights reserved.)

RipA plays a vital role during cell division of Mycobacterium tuberculosis by degrading the cell wall peptidoglycan at the septum, allowing daughter cell separation. The peptidoglycan degrading activity relies on the NlpC/P60 domain, and as it is potentially harmful when deregulated, spatial and temporal control is necessary in this process. The N-terminal domain of RipA has been proposed to play an inhibitory role blocking the C-terminal NlpC/P60 domain. Accessibility of the active site cysteine residue is however not limited by the presence of the N-terminal domain, but by the lid-module of the inter-domain linker, which is situated in the peptide binding groove of the crystal structures of the catalytic domain. The 2.2 Å resolution structure of the N-terminal domain, determined by Se-SAD phasing, reveals an all-α-fold with 2 long α-helices, and shows similarity to bacterial periplasmic protein domains with scaffold-building role. Size exclusion chromatography and SAXS experiments are consistent with dimer formation of this domain in solution. The SAXS data from the periplasmic two-domain RipA construct suggest a rigid baton-like structure of the N-terminal module, with the catalytic domain connected by a 24 residue long flexible linker. This flexible linker allows for a catalytic zone, which is part of the spatiotemporal control of peptidoglycan degradation., (© 2018 Wiley Periodicals, Inc.)

A variety of different tiles for the construction of DNA lattices have been developed since the structural DNA nanotechnology field was born. The majority of these are designed for the realization of close-packed structures, where DNA helices are arranged in parallel and tiles are connected through sticky ends. Assembly of such structures requires the use of cation-rich buffers to minimize repulsion between parallel helices, which poses limits to the application of DNA nanostructures. Wireframe structures, on the other hand, are less susceptible to salt concentration, but the assembly of wireframe lattices is limited by the availability of tiles and motifs. Herein, we report the construction of a polyhedral 12-arm junction for the self-assembly of wireframe DNA lattices. Our approach differs from traditional assembly of DNA tiles through hybridization of sticky ends. Instead, the assembly approach presented here uses small polyhedral shapes as connecting points and branch points of wires in a lattice structure. Using this design principle and characterization techniques, such as transmission electron microscopy, single-particle reconstruction, patterning of gold nanoparticles, dynamic light scattering, UV melting analyses, and small-angle X-ray scattering among others, we demonstrated formation of finite 12-way junction structures, as well as 1D and 2D short assemblies, demonstrating an alternative way of designing polyhedral structures and lattices.

Bioinspired in vitro collagen mineralization experiments have been performed in the presence of citrate and the combined role of the two bone organic matrix components in controlling mineral formation was investigated for the first time. Mineralized and non-mineralized collagen fibrils have been in depth characterized by combining small- and wide-angle X-ray scattering (SAXS/WAXS) techniques with Atomic Force Microscopy (AFM) imaging. A synergic effect of collagen and citrate in driving the formation of long-term stable amorphous calcium phosphate (ACP) nanoparticles with platy morphology was found. AFM images on mineralized collagen fibrils revealed that some of the ACP nanoparticles were deposited on the intramolecular nanoscopic holes of collagen fibrils., Statement of Significance: Citrate is an important component of the bone organic matrix but its specific role in bone mineralization is presently unclear. In this work, bioinspired in vitro collagen mineralization experiments in the presence of citrate have been carried out and the combined role of collagen and citrate in controlling mineral formation has been addressed for the first time. Through X-ray scattering and Atomic Force Microscopy characterizations on mineralized and non-mineralized collagen fibrils, we have found that citrate in synergy with collagen stabilizes an amorphous calcium phosphate (ACP) phase with platy morphology over one week and controls its deposition on collagen fibrils., (Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Uterine artery pseudoaneurysm is a rare complication after caesarean section and may be the cause of acute and potentially life-threatening secondary post-partum haemorrhage. In this case report, we present the clinical presentation and management of a case of uterine artery pseudoaneurysm manifesting as delayed post-partum bleeding after caesarean section. The case report emphasizes the importance of performing Doppler-flow when diagnosing the existence of a uterine artery pseudoaneurysm ultrasonically.