Heme Exporter FLVCR1a Regulates Heme Synthesis and Degradation and Controls Activity of Cytochromes P450

Background & AimsThe liver has one of the highest rates of heme synthesis of any organ. More than 50% of the heme synthesized in the liver is used for synthesis of P450 enzymes, which metabolize exogenous and endogenous compounds that include natural products, hormones, drugs, and carcinogens. Feline leukemia virus subgroup C cellular receptor 1a (FLVCR1a) is plasma membrane heme exporter that is ubiquitously expressed and controls intracellular heme content in hematopoietic lineages. We investigated the role of Flvcr1a in liver function in mice.MethodsWe created mice with conditional disruption of Mfsd7b, which encodes Flvcr1a, in hepatocytes (Flvcr1a fl/fl;alb-cre mice). Mice were analyzed under basal conditions, after phenylhydrazine-induced hemolysis, and after induction of cytochromes P450 synthesis. Livers were collected and analyzed by histologic, quantitative real-time polymerase chain reaction, and immunoblot analyses. Hepatic P450 enzymatic activities were measured.ResultsFlvcr1afl/fl;alb-cre mice accumulated heme and iron in liver despite up-regulation of heme oxygenase 1, ferroportin, and ferritins. Hepatic heme export activity of Flvcr1a was closely associated with heme biosynthesis, which is required to sustain cytochrome induction. Upon cytochromes P450 stimulation, Flvcr1afl/fl;alb-cre mice had reduced cytochrome activity, associated with accumulation of heme in hepatocytes. The expansion of the cytosolic heme pool in these mice was likely responsible for the early inhibition of heme synthesis and increased degradation of heme, which reduced expression and activity of cytochromes P450.ConclusionsIn livers of mice, Flvcr1a maintains a free heme pool that regulates heme synthesis and degradation as well as cytochromes P450 expression and activity. These findings have important implications for drug metabolism

Data demonstrating the anti-oxidant role of hemopexin in the heart

The data presented in this article are related to the research article entitled Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress (G. Ingoglia, C. M. Sag, N. Rex, L. De Franceschi, F. Vinchi, J. Cimino, S. Petrillo, S. Wagner, K. Kreitmeier, L. Silengo, F. Altruda, L. S. Maier, E. Hirsch, A. Ghigo and E. Tolosano, 2017) [1]. Data show that heme induces reactive oxygen species (ROS) production in primary cardiomyocytes. H9c2 myoblastic cells treated with heme bound to human Hemopexin (Hx) are protected from heme accumulation and oxidative stress. Similarly, the heme-driven oxidative response is reduced in primary cardiomyocytes treated with Hx-heme compared to heme alone. Our in vivo data show that mouse models of hemolytic disorders, β-thalassemic mice and phenylhydrazine-treated mice, have low serum Hx associated to enhanced expression of heme- and oxidative stress responsive genes in the heart. Hx-/- mice do not show signs of heart fibrosis or overt inflammation. For interpretation and discussion of these data, refer to the research article referenced above. Keywords: Heme, Hemopexin, Heart, Oxidative stres

AI is a viable alternative to high throughput screening: a 318-target study

: High throughput screening (HTS) is routinely used to identify bioactive small molecules. This requires physical compounds, which limits coverage of accessible chemical space. Computational approaches combined with vast on-demand chemical libraries can access far greater chemical space, provided that the predictive accuracy is sufficient to identify useful molecules. Through the largest and most diverse virtual HTS campaign reported to date, comprising 318 individual projects, we demonstrate that our AtomNet® convolutional neural network successfully finds novel hits across every major therapeutic area and protein class. We address historical limitations of computational screening by demonstrating success for target proteins without known binders, high-quality X-ray crystal structures, or manual cherry-picking of compounds. We show that the molecules selected by the AtomNet® model are novel drug-like scaffolds rather than minor modifications to known bioactive compounds. Our empirical results suggest that computational methods can substantially replace HTS as the first step of small-molecule drug discovery

Cosmological probes from weak lensing analysis on galaxy clusters

The Universe is composed of matter distributed on the large scale as a web-like structure. Galaxy clusters are located at the densest regions of the cosmic structure, the dark matter halos. Therefore, studying this class of objects gives crucial insights into the evolution of the matter distribution in the Universe and precious clues on the nature of the enigmatic dark matter. The dark matter, largely dominant in the halos, is invisible with direct observations. However, we can determine the dark matter distribution around galaxy clusters by means of weak gravitational lensing. This method takes advantage of the deflection of light induced by massive objects, namely clusters of galaxies, to derive their mass density profiles on scales reaching the halo boundaries and beyond, extending into the regime of the large-scale structure. Galaxy clusters and their hosts, the halos, are biased tracers of the underlying matter density field. This effect is characterized by the so-called halo bias, a parameter scaling the density profiles driven by the correlated matter distribution around galaxy clusters. During my thesis, I investigated the relation between the halo mass and the halo bias derived from stacked weak lensing profiles of about 7000 AMICO galaxy clusters. This catalog is assessed from the third data release of KiDS, the ESO public survey. Stacking the profiles is a process that reduces the statistical noise of the lensing signal and increases the quality of the measured parameters. We thus split the cluster sample into 14 redshift-richness bins and derived the halo bias and the virial mass in each bin by means of a standard Bayesian inference. It is carried out by a fiducial density model broken in a one-halo term, identified with the galaxy cluster halo and its physical characteristics (mass, concentration, etc), and a two-halo term, associated with matter distributed in distinct pairs of halos and directly proportional to the halo bias. The two terms of the halo profile correlate in such a way that the halo bias follows an increasing function of mass. This relation has been shown and modeled in several theoretical studies based on \textit{N}-body numerical simulations, in the framework of the $\Lambda$CDM standard cosmological model. The results of our study show an agreement within $2\sigma$ between our estimation of the halo bias and theoretical predictions. The measurements of the average mass and bias over the stacked density profile of the full cluster catalog give $M_{200c} = (4.9 \pm 0.3) \times 10^{13} M_{\odot}/\textit{h}$ and $b_h \sigma_8^2 = 1.2 \pm 0.1$. Considering the degenerated form of the halo bias and the additional prior of a bias-mass relation from numerical simulations, we constrained the normalization of the matter power spectrum. We found $\sigma_8 = 0.63 \pm 0.10$ with the matter density of the Universe set at $\Omega_m = 0.3$. Even if a fixed cosmology does not allow to complete a fully independent cosmological inference, this result agrees with other studies based on CMB data, cluster clustering, cluster counts, and cosmic shear analyses within $2\sigma$. In the upcoming years, the next generation of sky surveys will provide deeper and wider catalogs of data for cosmologists to answer modern inquiries. As part of my thesis, I have been involved in the development of a numerical tool in the context of the Science Ground Segment data processing pipeline of the Euclid consortium. COMB-CL is a python module (at the moment, in a state of prototype) that aims to measure the weak lensing mass of galaxy clusters. The code is built in such a way that catalogs of cluster and galaxy properties (position, redshift, shear, color, etc) are input and, given a fiducial cosmology and a model for the halo density profile, catalogs of weak lensing profiles and related masses are output. This toolkit has been accepted and will be reviewed in a paper of the key project LE3-CL-2 regarding the characterization of the properties of detected galaxy clusters

A New Configuration for Helmet Continuous Positive Airway Pressure Allowing Tidal Volume Monitoring

We performed a bench and human study to test the hypothesis that a ventilator can accurately estimate Vt when a helmet is used in CPAP mode in a single limb configuration with an intentional leak port placed at the helmet expiratory port. We have recently demonstrated the effectiveness of this setup in bilevel mod

AMICO galaxy clusters in KiDS-DR3

Context. The large-scale mass distribution around dark matter haloes hosting galaxy clusters provides sensitive cosmological information. Aims. In this work we make use of a large photometric galaxy cluster sample, constructed from the public Third Data Release of the Kilo-Degree Survey, and the corresponding shear signal, to assess cluster masses and test the concordance Λ-cold dark matter (ΛCDM) model. In particular, we study the weak gravitational lensing effects on scales beyond the cluster virial radius, where the signal is dominated by correlated and uncorrelated matter density distributions along the line of sight. The analysed catalogue consists of 6962 galaxy clusters, in the redshift range 0.1 ≤ z ≤ 0.6 and with signal-to-noise ratios higher than 3.5. Methods. We perform a full Bayesian analysis to model the stacked shear profiles of these clusters. The adopted likelihood function considers both the small-scale one-halo term, used primarily to constrain the cluster structural properties, and the two-halo term, that can be used to constrain cosmological parameters. Results. We find that the adopted modelling is successful in assessing both the cluster masses and the total matter density parameter, ΩM, when fitting shear profiles up to the largest available scales of 35 Mpc h−1. Moreover, our results provide a strong observational evidence of the two-halo signal in the stacked gravitational lensing of galaxy clusters, further demonstrating the reliability of this probe for cosmological studies. The main result of this work is a robust constraint on ΩM, assuming a flat ΛCDM cosmology. We get ΩM = 0.29 ± 0.02, estimated from the full posterior probability distribution, consistent with the estimates from cosmic microwave background experiments

Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress

Heart failure is a leading cause of morbidity and mortality in patients affected by different disorders associated to intravascular hemolysis. The leading factor is the presence of pathologic amount of pro-oxidant free heme in the bloodstream, due to the exhaustion of the natural heme scavenger Hemopexin (Hx). Here, we evaluated whether free heme directly affects cardiac function, and tested the therapeutic potential of replenishing serum Hx for increasing serum heme buffering capacity. The effect of heme on cardiac function was assessed in vitro, on primary cardiomyocytes and H9c2 myoblast cell line, and in vivo, in Hx(-/-) mice and in genetic and acquired mouse models of intravascular hemolysis. Purified Hx or anti-oxidants N-Acetyl-L-cysteine and a-tocopherol were used to counteract heme cardiotoxicity. In mice, Hx loss/depletion resulted in heme accumulation and enhanced reactive oxygen species (ROS) production in the heart, which ultimately led to severe systolic dysfunction. Similarly, high ROS reduced systolic Ca2+ transient amplitudes and fractional shortening in primary cardiomyocytes exposed to free heme. In keeping with these Ca2+ handling alterations, oxidation and CaMKII-dependent phosphorylation of Ryanodine Receptor 2 were higher in Hx(-/-) hearts than in controls. Administration of anti-oxidants prevented systolic failure both in vitro and in vivo. Intriguingly, Hx rescued contraction defects of heme-treated cardiomyocytes and preserved cardiac function in hemolytic mice. We show that heme-mediated oxidative stress perturbs cardiac Ca2+ homeostasis and promotes contractile dysfunction. Scavenging heme, Hx counteracts cardiac heme toxicity and preserves left ventricular function. Our data generate the rationale to consider the therapeutic use of Hx to limit the cardiotoxicity of free heme in hemolytic disorders