Reprogramming human T cell function and specificity with non-viral genome targeting.

Decades of work have aimed to genetically reprogram T cells for therapeutic purposes1,2 using recombinant viral vectors, which do not target transgenes to specific genomic sites3,4. The need for viral vectors has slowed down research and clinical use as their manufacturing and testing is lengthy and expensive. Genome editing brought the promise of specific and efficient insertion of large transgenes into target cells using homology-directed repair5,6. Here we developed a CRISPR-Cas9 genome-targeting system that does not require viral vectors, allowing rapid and efficient insertion of large DNA sequences (greater than one kilobase) at specific sites in the genomes of primary human T cells, while preserving cell viability and function. This permits individual or multiplexed modification of endogenous genes. First, we applied this strategy to correct a pathogenic IL2RA mutation in cells from patients with monogenic autoimmune disease, and demonstrate improved signalling function. Second, we replaced the endogenous T cell receptor (TCR) locus with a new TCR that redirected T cells to a cancer antigen. The resulting TCR-engineered T cells specifically recognized tumour antigens and mounted productive anti-tumour cell responses in vitro and in vivo. Together, these studies provide preclinical evidence that non-viral genome targeting can enable rapid and flexible experimental manipulation and therapeutic engineering of primary human immune cells

Particle interactions in liquid magnetic colloids by zero field cooled measurements: effects on heating efficiency

The influence of magnetic interactions in assemblies formed by either aggregated or disaggregated uniform gamma-Fe_2O_3 particles are investigated as a function of particle size, concentration, and applied field. Hyperthermia and magnetization measurements are performed in the liquid phase of colloids consisting of 8 and 13 nm uniform gamma-Fe_2O_3 particles dispersed in water and hexane. Although hexane allows the disagglomerated obtaining particle system; aggregation is observed in the case of water colloids. The zero field cooled (ZFC) curves show a discontinuity in the magnetization values associated with the melting points of water and hexane. Additionally, for 13 nm gamma-Fe_2O_3 dispersed in hexane, a second magnetization jump is observed that depends on particle concentration and shifts toward lower temperature by increasing applied field. This second jump is related to the strength of the magnetic interactions as it is only present in disagglomerated particle systems with the largest size, i.e., is not observed for 8 nm superparamagnetic particles, and surface effects can be discarded. The specific absorption rate (SAR) decreases with increasing concentration only for the hexane colloid, whereas for aqueous colloids, the SAR is almost independent of particle concentration. Our results suggest that, as a consequence of the magnetic interactions, the dipolar field acting on large particles increases with concentration, leading to a decrease of the SAR

Reduced Plasma Levels of 25-Hydroxycholesterol and Increased Cerebrospinal Fluid Levels of Bile Acid Precursors in Multiple Sclerosis Patients

Multiple sclerosis (MS) is an autoimmune, inflammatory disease of the central nervous system (CNS). We have measured the levels of over 20 non-esterified sterols in plasma and cerebrospinal fluid (CSF) from patients suffering from MS, inflammatory CNS disease, neurodegenerative disease and control patients. Analysis was performed following enzyme-assisted derivatisation by liquid chromatography-mass spectrometry (LC-MS) exploiting multistage fragmentation (MS n ). We found increased concentrations of bile acid precursors in CSF from each of the disease states and that patients with inflammatory CNS disease classified as suspected autoimmune disease or of unknown aetiology also showed elevated concentrations of 25-hydroxycholestertol (25-HC, P < 0.05) in CSF. Cholesterol concentrations in CSF were not changed except for patients diagnosed with amyotrophic lateral sclerosis (P < 0.01) or pathogen-based infections of the CNS (P < 0.05) where they were elevated. In plasma, we found that 25-HC (P < 0.01), (25R)26-hydroxycholesterol ((25R)26-HC, P < 0.05) and 7α-hydroxy-3-oxocholest-4-enoic acid (7αH,3O-CA, P < 0.05) were reduced in relapsing-remitting MS (RRMS) patients compared to controls. The pattern of reduced plasma levels of 25-HC, (25R)26-HC and 7αH,3O-CA was unique to RRMS. In summary, in plasma, we find that the concentration of 25-HC in RRMS patients is significantly lower than in controls. This is consistent with the hypothesis that a lower propensity of macrophages to synthesise 25-HC will result in reduced negative feedback by 25-HC on IL-1 family cytokine production and exacerbated MS. In CSF, we find that the dominating metabolites reflect the acidic pathway of bile acid biosynthesis and the elevated levels of these in CNS disease is likely to reflect cholesterol release as a result of demyelination or neuronal death. 25-HC is elevated in patients with inflammatory CNS disease probably as a consequence of up-regulation of the type 1 interferon-stimulated gene cholesterol 25-hydroxylase in macrophage

Early Eocene clinoenstatite boninite and boninite-like dikes of the ophiolite of New Caledonia; a witness of slab-derived enrichment of the mantle wedge in a nascent volcanic arc

Clinoenstatite-bearing boninites (CE-boninite) from the serpentinite sole of the Cenozoic ophiolite of New Caledonia near Nepoui have been dated by the 40Ar/39Ar method, yielding two plateau ages of 47.4 ± 0.9 Ma and 50.4 ± 1.3 Ma. Coarser grained, geochemically similar boninite-series felsic dikes consistently yielded U–Pb zircon ages of ca. 54 Ma. Nepoui CE-boninites display whole rock geochemical features similar to that of Cape Vogel boninites (Papua-New Guinea). They similarly have been generated by low degree hydrous melting of depleted peridotite. High contents in LILE and LREE, and some elemental ratios suggest source enrichment by subduction-derived fluids and melts. However, unlike the Cape Vogel boninite, moderately depleted MORB-like isotopic signatures (εNd50 = 7.9) rule out the role of OIB-like, or E-MORB component that might account for the relatively high LREE and LILE contents measured in the rocks. Nd isotopic ratios and positive anomalies in Zr and Hf are closely similar to that of the slightly older felsic dikes (55–50 Ma) that crosscut the peridotite from the ophiolite in New Caledonia. Most of these magmas have been generated by slab melting during the early stages of intra-oceanic subduction. The Early Eocene subduction started at or near the “oceanic” ridge and involved young and hot lithosphere; therefore, slab-derived melts may have reacted locally with hot depleted peridotites. Finally, water influx into the mantle wedge during the subduction of slightly older (cooler and hydrated) lithosphere initiated a low degree partial melting event in the mantle wedge and generated the CE-boninite magma. Geochemical modeling of hydrous melting of a depleted mantle re-enriched by slab melts suggest that the additional slab melt component was derived from the partial melting of a BABB-like barroisite-bearing eclogite, similar to some elements of the Eocene HP–LT Pouebo terrane. This potential magma source is similar to the BABB-like HT amphibolites of the metamorphic sole of the ophiolite, which have the same origin. Geochemical modeling also suggests that CE-boninite magma may have been in equilibrium with the enstatite-bearing gabbro cumulates that crop out in several places of the Massif du Sud. However, modeling fails in establishing that harzburgite of the same massif simply corresponds to the melting residue of this process. It appears that ultra-depleted supra-subduction peridotites of the Massif du Sud are probably not directly related to the overlying gabbro cumulates

Learning form Nature to improve the heat generation of iron-oxide nanoparticles for magnetic hyperthermia applications.

The performance of magnetic nanoparticles is intimately entwined with their structure, mean size and magnetic anisotropy. Besides, ensembles offer a unique way of engineering the magnetic response by modifying the strength of the dipolar interactions between particles. Here we report on an experimental and theoretical analysis of magnetic hyperthermia, a rapidly developing technique in medical research and oncology. Experimentally, we demonstrate that single-domain cubic iron oxide particles resembling bacterial magnetosomes have superior magnetic heating efficiency compared to spherical particles of similar sizes. Monte Carlo simulations at the atomic level corroborate the larger anisotropy of the cubic particles in comparison with the spherical ones, thus evidencing the beneficial role of surface anisotropy in the improved heating power. Moreover we establish a quantitative link between the particle assembling, the interactions and the heating properties. This knowledge opens new perspectives for improved hyperthermia, an alternative to conventional cancer therapies

CD40 ligand and MHC class II expression are essential for human peripheral B cell tolerance

Hyper-IgM (HIGM) syndromes are primary immunodeficiencies characterized by defects of class switch recombination and somatic hypermutation. HIGM patients who carry mutations in the CD40-ligand (CD40L) gene expressed by CD4+ T cells suffer from recurrent infections and often develop autoimmune disorders. To investigate the impact of CD40L–CD40 interactions on human B cell tolerance, we tested by ELISA the reactivity of recombinant antibodies isolated from single B cells from three CD40L-deficient patients. Antibody characteristics and reactivity from CD40L-deficient new emigrant B cells were similar to those from healthy donors, suggesting that CD40L–CD40 interactions do not regulate central B cell tolerance. In contrast, mature naive B cells from CD40L-deficient patients expressed a high proportion of autoreactive antibodies, including antinuclear antibodies. Thus, CD40L–CD40 interactions are essential for peripheral B cell tolerance. In addition, a patient with the bare lymphocyte syndrome who could not express MHC class II molecules failed to counterselect autoreactive mature naive B cells, suggesting that peripheral B cell tolerance also depends on major histocompatibility complex (MHC) class II–T cell receptor (TCR) interactions. The decreased frequency of MHC class II–restricted CD4+ regulatory T cells in CD40L-deficient patients suggests that these T cells may mediate peripheral B cell tolerance through CD40L–CD40 and MHC class II–TCR interactions

Geodynamic Significance of the Mesoproterozoic Magmatism of the Udzha Paleo-Rift (Northern Siberian Craton) Based on U-Pb Geochronology and Paleomagnetic Data

The emplacement age of the Great Udzha Dyke (northern Siberian Craton) was determined by the U-Pb dating of apatite using laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS). This produced an age of 1386 +/- 30 Ma. This dyke along with two other adjacent intrusions, which cross-cut the sedimentary units of the Udzha paleo-rift, were subjected to paleomagnetic investigation. The paleomagnetic poles for the Udzha paleo-rift intrusions are consistent with previous results published for the Chieress dyke in the Anabar shield of the Siberian Craton (1384 +/- 2 Ma). Our results suggest that there was a period of intense volcanism in the northern Siberian Craton, as well as allow us to reconstruct the apparent migration of the Siberian Craton during the Mesoproterozoic.Peer reviewe

The inflammatory APRIL (a proliferation-inducing ligand) antagonizes chondroitin sulphate proteoglycans to promote axonal growth and myelination.

Lesions in the CNS are frequently associated to a detrimental inflammatory reaction. In autoimmune neurodegenerative diseases, a proliferation-inducing ligand (APRIL) produced by CNS-infiltrating inflammatory cells binds to chondroitin sulphate proteoglycans (CSPGs). The latter are well-established obstacles to neural regeneration and remyelination in the CNS by interacting with receptor protein tyrosine phosphatase (RPTP) and Nogo receptor (NgR) families. Here, we are showing that APRIL blocks the interactions of RPTP and NgR with all types of chondroitin sulphate (CS). Functionally, APRIL neutralized the inhibitory effects of CS on mouse and human neuronal process growth. APRIL also blocked the inhibition of CS on mouse and human oligodendrocyte differentiation. Finally, APRIL increased myelination in an ex vivo organotypic model of demyelination in the presence of endogenous CSPG upregulation. Our data demonstrate the potential value for a recombinant form of soluble APRIL to achieve repair in the CNS