PEG Branched Polymer for Functionalization of Nanomaterials with Ultralong Blood Circulation

Nanomaterials have been actively pursued for biological and medical applications in recent years. Here, we report the synthesis of several new poly(ethylene glycol) grafted branched-polymers for functionalization of various nanomaterials including carbon nanotubes, gold nanoparticles (NP) and gold nanorods (NRs), affording high aqueous solubility and stability for these materials. We synthesize different surfactant polymers based upon poly-(g-glutamic acid) (gPGA) and poly(maleic anhydride-alt-1-octadecene) (PMHC18). We use the abundant free carboxylic acid groups of gPGA for attaching lipophilic species such as pyrene or phospholipid, which bind to nanomaterials via robust physisorption. Additionally, the remaining carboxylic acids on gPGA or the amine-reactive anhydrides of PMHC18 are then PEGylated, providing extended hydrophilic groups, affording polymeric amphiphiles. We show that single-walled carbon nanotubes (SWNTs), Au NPs and NRs functionalized by the polymers exhibit high stability in aqueous solutions at different pHs, at elevated temperatures and in serum. Morever, the polymer-coated SWNTs exhibit remarkably long blood circulation (t1/2 22.1 h) upon intravenous injection into mice, far exceeding the previous record of 5.4 h. The ultra-long blood circulation time suggests greatly delayed clearance of nanomaterials by the reticuloendothelial system (RES) of mice, a highly desired property for in vivo applications of nanomaterials, including imaging and drug delivery

Maternal neurofascin-specific autoantibodies bind to structures of the fetal nervous system during pregnancy, but have no long term effect on development in the rat

Neurofascin was recently reported as a target for axopathic autoantibodies in patients with multiple sclerosis (MS), a response that will exacerbate axonal pathology and disease severity in an animal model of multiple sclerosis. As transplacental transfer of maternal autoantibodies can permanently damage the developing nervous system we investigated whether intrauterine exposure to this neurofascin-specific response had any detrimental effect on white matter tract development. To address this question we intravenously injected pregnant rats with either a pathogenic anti-neurofascin monoclonal antibody or an appropriate isotype control on days 15 and 18 of pregnancy, respectively, to mimic the physiological concentration of maternal antibodies in the circulation of the fetus towards the end of pregnancy. Pups were monitored daily with respect to litter size, birth weight, growth and motor development. Histological studies were performed on E20 embryos and pups sacrificed on days 2, 10, 21, 32 and 45 days post partum. Results: Immunohistochemistry for light and confocal microscopy confirmed passively transferred anti-neurofascin antibody had crossed the placenta to bind to distinct structures in the developing cortex and cerebellum. However, this did not result in any significant differences in litter size, birth weight, or general physical development between litters from control mothers or those treated with the neurofascin-specific antibody. Histological analysis also failed to identify any neuronal or white matter tract abnormalities induced by the neurofascin-specific antibody. Conclusions: We show that transplacental transfer of circulating anti-neurofascin antibodies can occur and targets specific structures in the CNS of the developing fetus. However, this did not result in any pre- or post-natal abnormalities in the offspring of the treated mothers. These results assure that even if anti-neurofascin responses are detected in pregnant women with multiple sclerosis these are unlikely to have a negative effect on their children

Estradiol alters the immune-responsiveness of cervical epithelial cells stimulated with ligands of Toll-like receptors 2 and 4.

The mucosa of the female reproductive tract plays a pivotal role in host defence. Pregnancy must alter immunological mechanisms at this interface to protect the conceptus. We sought to determine how estradiol (E2) alters the immune-responsiveness of cervical epithelial cells to ligand stimulation of Toll-like receptor (TLR)-2 and -4. Human ectocervical epithelial cells (HECECs) were cultured and co-incubated with two concentrations of E2 and peptidoglycan (PGN) or lipopolysaccharide (LPS) over durations that ranged between 10 minutes and 18 hours. Cytometric Bead Array was performed to quantify eight cytokines in the supernatant fluid. In response to PGN, HECECs co-incubated with E2 released lesser quantities of IL-1ß and IFNγ, higher levels of RANTES, and variable levels of IL-6 and IL-8 than those not exposed to E2. In contrast, HECECs co-incubated with LPS and E2 secreted increased levels of IL-1ß, IL-6, IL-8, and IFNγ at 2 and 18 hours than HECECs not exposed to E2, and reduced levels of RANTES at same study time-points. Estradiol alters the immune-responsiveness of cultured HECECs to TLR2 and TLR4 ligands in a complex fashion that appears to vary with bacterial ligand, TLR subtype, and duration of exposure. Our observations are consistent with the functional complexity that this mucosal interface requires for its immunological roles

Beyond gene-disease validity: capturing structured data on inheritance, allelic-requirement, disease-relevant variant classes, and disease mechanism for inherited cardiac conditions

Background: As the availability of genomic testing grows, variant interpretation will increasingly be performed by genomic generalists, rather than domain-specific experts. Demand is rising for laboratories to accurately classify variants in inherited cardiac condition (ICC) genes, including secondary findings. Methods: We analyse evidence for inheritance patterns, allelic requirement, disease mechanism and disease-relevant variant classes for 65 ClinGen-curated ICC gene-disease pairs. We present this information for the first time in a structured dataset, CardiacG2P, and assess application in genomic variant filtering. Results: For 36/65 gene-disease pairs, loss of function is not an established disease mechanism, and protein truncating variants are not known to be pathogenic. Using the CardiacG2P dataset as an initial variant filter allows for efficient variant prioritisation whilst maintaining a high sensitivity for retaining pathogenic variants compared with two other variant filtering approaches. Conclusions: Access to evidence-based structured data representing disease mechanism and allelic requirement aids variant filtering and analysis and is a pre-requisite for scalable genomic testing

Beyond gene-disease validity: capturing structured data on inheritance, allelic requirement, disease-relevant variant classes, and disease mechanism for inherited cardiac conditions

Background: As the availability of genomic testing grows, variant interpretation will increasingly be performed by genomic generalists, rather than domain-specific experts. Demand is rising for laboratories to accurately classify variants in inherited cardiac condition (ICC) genes, including secondary findings. // Methods: We analyse evidence for inheritance patterns, allelic requirement, disease mechanism and disease-relevant variant classes for 65 ClinGen-curated ICC gene-disease pairs. We present this information for the first time in a structured dataset, CardiacG2P, and assess application in genomic variant filtering. // Results: For 36/65 gene-disease pairs, loss of function is not an established disease mechanism, and protein truncating variants are not known to be pathogenic. Using the CardiacG2P dataset as an initial variant filter allows for efficient variant prioritisation whilst maintaining a high sensitivity for retaining pathogenic variants compared with two other variant filtering approaches. // Conclusions: Access to evidence-based structured data representing disease mechanism and allelic requirement aids variant filtering and analysis and is a pre-requisite for scalable genomic testing

A novel application of capnography during controlled human exposure to air pollution

BACKGROUND: The objective was to determine the repeatability and stability of capnography interfaced with human exposure facility. METHODS: Capnographic wave signals were obtained from five healthy volunteers exposed to particle-free, filtered air during two consecutive 5 min intervals, 10 min apart, within the open and then the sealed and operational human exposure facility (HEF). Using a customized setup comprised of the Oridion Microcap(® )portable capnograph, DA converter and AD card, the signal was acquired and saved as an ASCII file for subsequent processing. The minute ventilation (VE), respiratory rate (RR) and expiratory tidal volume (V(TE)) were recorded before and after capnographic recording and then averaged. Each capnographic tracing was analyzed for acceptable waves. From each recorded interval, 8 to 19 acceptable waves were selected and measured. The following wave parameters were obtained: total length and length of phase II and III, slope of phase II and III, area under the curve and area under phase III. In addition, we recorded signal measures including the mean, standard deviation, mode, minimum, maximum – which equals end-tidal CO(2 )(EtCO(2)), zero-corrected maximum and true RMS. RESULTS: Statistical analysis using a paired t-test for means showed no statistically significant changes of any wave parameters and wave signal measures, corrected for RR and V(TE), comparing the measures when the HEF was open vs. sealed and operational. The coefficients of variation of the zero-corrected and uncorrected EtCO(2), phase II absolute difference, signal mean, standard deviation and RMS were less than 10% despite a sub-atmospheric barometric pressure, and slightly higher temperature and relative humidity within the HEF when operational. CONCLUSION: We showed that a customized setup for the acquisition and processing of the capnographic wave signal, interfaced with HEF was stable and repeatable. Thus, we expect that analysis of capnographic waves in controlled human air pollution exposure studies is a feasible tool for characterization of cardio-pulmonary effects of such exposures

Molecular definition of group 1 innate lymphoid cells in the mouse uterus

Determining the function of uterine lymphocytes is challenging because of the rapidly changing nature of the organ in response to sex hormones and, during pregnancy, to the invading fetal trophoblast cells. Here we provide the first genome-wide transcriptome atlas of mouse uterine group 1 innate lymphoid cells (g1 ILCs) at mid-gestation. The composition of g1 ILCs fluctuates throughout reproductive life, with Eomes-veCD49a+ ILC1s dominating before puberty and specifically expanding in second pregnancies, when the expression of CXCR6, a marker of memory cells, is upregulated. Tissue-resident Eomes+CD49a+ NK cells (trNK), which resemble human uterine NK cells, are most abundant during early pregnancy, and showcase gene signatures of responsiveness to TGF-β, connections with trophoblast, epithelial, endothelial and smooth muscle cells, leucocytes, as well as extracellular matrix. Unexpectedly, trNK cells express genes involved in anaerobic glycolysis, lipid metabolism, iron transport, protein ubiquitination, and recognition of microbial molecular patterns. Conventional NK cells expand late in gestation and may engage in crosstalk with trNK cells involving IL-18 and IFN-γ. These results identify trNK cells as the cellular hub of uterine g1 ILCs at mid-gestation and mark CXCR6+ ILC1s as potential memory cells of pregnancy.This work was funded by a Wellcome Trust Investigator Award 200841/Z/16/Z, the Centre for Trophoblast Research (CTR), and the Cambridge NIHR BRC Cell Phenotyping Hub to FC, the Associazione Italiana Ricerca per la Ricerca sul Cancro (AIRC) - Special Project 5x1000 no. 9962, AIRC IG 2017 Id.19920 and AIRC 2014 Id. 15283 to LM, and Ministero della Salute RF-2013, GR-2013-02356568 to PV. IF was funded by a CTR PhD fellowship