Biology subjects

Mora, J.

Publications and source records attributed to Mora, J..

27 records · Page 2Linked to original sources

bioRxiv2021

A multimodal imaging and analysis pipeline for creating a cellular census of the human cerebral cortex

Cells are not uniformly distributed in the human cerebral cortex. Rather, they are arranged in a regional and laminar fashion that span a range of scales. Here we demonstrate an innovative imaging and analysis pipeline to construct a reliable cell census across the human cerebral cortex. Magnetic resonance imaging (MRI) is used to establish a macroscopic reference coordinate system of laminar and cytoarchitectural boundaries. Cell counting is obtained with both traditional immunohistochemistry, to provide a stereological gold-standard, and with a custom-made inverted confocal light-sheet fluorescence microscope (LSFM) for 3D imaging at cellular resolution. Finally, mesoscale optical coherence tomography (OCT) enables the registration of the distorted histological cell typing obtained with LSFM to the MRI-based atlas coordinate system.

neuroscience↗

bioRxiv2021

Low dose AKT inhibitor miransertib cures PI3K-related vascular malformations in preclinical models of human disease

Low-flow vascular malformations are congenital overgrowths composed by abnormal blood vessels potentially causing pain, bleeding, and obstruction of different organs. These diseases are caused by oncogenic mutations in the endothelium which result in overactivation of the PI3K/AKT pathway. Lack of robust in vivo preclinical data has prevented the development and translation into clinical trials of specific molecular therapies for these diseases. Here, we describe a new reproducible preclinical in vivo model of PI3K-driven vascular malformations using the postnatal mouse retina. This model reproduces human disease with Pik3ca activating mutations expressed in a mosaic pattern and vascular malformations formed in veins and capillaries. We show that active angiogenesis is required for the pathogenesis of vascular malformations caused by activating Pik3ca mutations. Using this model, we demonstrate that low doses of the AKT inhibitor miransertib both prevents and induces the regression of PI3K-driven vascular malformations. We confirmed miransertib efficacy in isolated human endothelial cells with genotypes spanning most of human low-flow vascular malformations. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=69 SRC="FIGDIR/small/452617v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@15b29b6org.highwire.dtl.DTLVardef@fa9f8org.highwire.dtl.DTLVardef@6001d8org.highwire.dtl.DTLVardef@11926c5_HPS_FORMAT_FIGEXP M_FIG C_FIG Low-flow vascular malformations are caused by PI3K signalling overactivation in endothelial cells. We have generated an optimised and robust preclinical system of PI3K-driven vascular malformations by inducing the mosaic expression of Pik3caH1047R in the retinal angiogenic endothelium. This preclinical model displays traits constituting the main hallmarks of the pathogenesis of low-flow blood vascular malformations: overactivation of PI3K signalling (high phospho-S6), vascular compartment specificity, loss of pericyte coverage, and endothelial cell hyperproliferation. Using this preclinical model we report that low dose AKT inhibitor miransertib prevents and regress PI3K-driven vascular malformations.

molecular biology↗

bioRxiv2021

Persistence of SARS CoV-2 S1 Protein in CD16+ Monocytes in Post-Acute Sequelae of COVID-19 (PASC) Up to 15 Months Post-Infection

The recent COVID-19 pandemic is a treatment challenge in the acute infection stage but the recognition of chronic COVID-19 symptoms termed post-acute sequelae SARS-CoV-2 infection (PASC) may affect up to 30% of all infected individuals. The underlying mechanism and source of this distinct immunologic condition three months or more after initial infection remains elusive. Here, we investigated the presence of SARS-CoV-2 S1 protein in 46 individuals. We analyzed T-cell, B-cell, and monocytic subsets in both severe COVID-19 patients and in patients with post-acute sequelae of COVID-19 (PASC). The levels of both intermediate (CD14+, CD16+) and non-classical monocyte (CD14Lo, CD16+) were significantly elevated in PASC patients up to 15 months post-acute infection compared to healthy controls (P=0.002 and P=0.01, respectively). A statistically significant number of non-classical monocytes contained SARS-CoV-2 S1 protein in both severe (P=0.004) and PASC patients (P=0.02) out to 15 months post-infection. Non-classical monocytes were sorted from PASC patients using flow cytometric sorting and the SARS-CoV-2 S1 protein was confirmed by mass spectrometry. Cells from 4 out of 11 severe COVID-19 patients and 1 out of 26 PASC patients contained ddPCR+ peripheral blood mononuclear cells, however, only fragmented SARS-CoV-2 RNA was found in PASC patients. No full length sequences were identified, and no sequences that could account for the observed S1 protein were identified in any patient. Non-classical monocytes are capable of causing inflammation throughout the body in response to fractalkine/CX3CL1 and RANTES/CCR5.

immunology↗

bioRxiv2021

High efficacy of therapeutic equine hyperimmune antibodies against SARS CoV-2 variants of concern

SARS-CoV-2 variants of concern (VoC) show reduced neutralization by vaccine-induced and therapeutic monoclonal antibodies. We tested therapeutic equine polyclonal antibodies (pAbs) against four VoC (alpha, beta, epsilon and gamma). We show that equine pAbs efficiently neutralize VoC, suggesting they are an effective, broad coverage, low-cost and a scalable COVID-19 treatment.

microbiology↗

bioRxiv2021

HDAC6 regulates expression of the oncogenic driver EWSR1-FLI1 through the EWSR1 promoter in Ewing sarcoma

Ewing sarcoma (EWS) is an aggressive developmental sarcoma driven by a fusion gene, EWSR1-FLI1. However, little is known about the regulation of EWSR1-FLI1 chimeric fusion gene expression. Here, we demonstrate that active nuclear HDAC6 in EWS modulates acetylation status of specificity protein 1 (SP1), consequently regulating SP1/P300 activator complex binding to EWSR1 and EWSR1-FLI1 promoters. Selective inhibition of HDAC6 impairs binding of activator complex SP1/P300, thereby inducing EWSR1-FLI1 downregulation and significantly reducing its oncogenic functions. In addition, sensitivity of EWS cell lines to HDAC6 inhibition is higher than other tumor or non-tumor cell lines. Overexpression of HDAC6 in primary EWS tumor clinical samples correlates with a poor prognosis. Notably, a combination treatment of a selective HDAC6 inhibitor and doxorubicin (standard of care in EWS) dramatically inhibits tumor growth in two EWS murine xenograft models. These results could lead to suitable and promising therapeutic alternatives for EWS patients.

cancer biology↗

bioRxiv2020

Immune-Based Prediction of COVID-19 Severity and Chronicity Decoded Using Machine Learning

Individuals with systemic symptoms long after COVID-19 has cleared represent approximately ~10% of all COVID-19 infected individuals. Here we present a bioinformatics approach to predict and model the phases of COVID so that effective treatment strategies can be devised and monitored. We investigated 144 individuals including normal individuals and patients spanning the COVID-19 disease continuum. We collected plasma and isolated PBMCs from 29 normal individuals, 26 individuals with mild-moderate COVID-19, 25 individuals with severe COVID-19, and 64 individuals with Chronic COVID-19 symptoms. Immune subset profiling and a 14-plex cytokine panel were run on all patients. Data was analyzed using machine learning methods to predict and distinguish the groups from each other.Using a multi-class deep neural network classifier to better fit our prediction model, we recapitulated a 100% precision, 100% recall and F1 score of 1 on the test set. Moreover, a first score specific for the chronic COVID-19 patients was defined as S1 = (IFN-{gamma} + IL-2)/ CCL4-MIP-1{beta}. Second, a score specific for the severe COVID-19 patients was defined as S2 = (10*IL-10 + IL-6) - (IL-2 + IL-8). Severe cases are characterized by excessive inflammation and dysregulated T cell activation, recruitment, and counteracting activities. While chronic patients are characterized by a profile able to induce the activation of effector T cells with pro-inflammatory properties and the capacity of generating an effective immune response to eliminate the virus but without the proper recruitment signals to attract activated T cells. SummaryImmunologic Modeling of Severity and Chronicity of COVID-19

immunology↗

bioRxiv2020

Lipoprotein distribution of Apo CIII glycoforms in healthy men is linked with low TG and increased insulin sensitivity

Glycosylation of Apo CIII modulates its function in triglyceride metabolism, and some variants are associated with a protective or pro-atherogenic lipid profile. These associations have been studied in whole plasma Apo CIII proteoforms, but the proportion of Apo CIII proteoforms in individual lipoprotein fractions has been rarely evaluated. In the present study, we aim to measure the relative content of Apo CIII proteoforms in each lipoprotein fraction (VLDL, IDL, LDL and HDL) in a group of healthy subjects as a potential biomarker for triglyceride metabolism, cardiovascular risk and diabetes. Lipoprotein fractions were separated by differential ultracentrifugation of plasma samples. The relative concentrations of seven Apo CIII variants were measured by mass spectrometric immunoassay, and the complete lipoprotein profile was determined by NMR. The results showed high interindividual variability in the distribution of Apo CIII proteoforms across the study population but a uniform proportion in all lipoprotein fractions. Two Apo CIII variants, Apo CIII0b and Apo CIII1d, were negatively correlated with plasma and VLDL triglycerides regardless of VLDL size and were associated with increased LDL size when transported in LDL particles. Apo CIII0b also showed a negative correlation with lipoprotein-insulin resistance score. Therefore, Apo CIII variants can be reliably measured in lipoprotein fractions, and our results suggest that Apo CIII0b and Apo CIII1d have a protective role in triglyceride metabolism and insulin resistance in healthy individuals.

molecular biology↗

bioRxiv2020

A multilayered post-GWAS assessment on genetic susceptibility to pancreatic cancer

Pancreatic cancer (PC) is a complex disease in which both non-genetic and genetic factors interplay. To-date, 40 GWAS hits have been associated with PC risk in individuals of European descent, explaining 4.1% of the phenotypic variance. Here, we complemented a classical new PC GWAS (1D) with spatial autocorrelation analysis (2D) and Hi-C maps (3D) to gain additional insight into the inherited basis of PC. In-silico functional analysis of public genomic information allowed prioritization of potentially relevant candidate variants. We replicated 17/40 previous PC-GWAS hits and identified novel variants with potential biological functions. The spatial autocorrelation approach prioritized low MAF variants not detected by GWAS. These were further expanded via 3D interactions to 54 target regions with high functional relevance. This multi-step strategy, combined with an in-depth in-silico functional analysis, offers a comprehensive approach to advance the study of PC genetic susceptibility and could be applied to other diseases.

genetics↗

bioRxiv2020

Sequential combinations of chemotherapeutic agents with BH3 mimetics to treat rhabdomyosarcoma and avoid resistance.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in childhood and adolescence. Refractory/relapsed RMS patients present a bad prognosis that combined with the lack of specific biomarkers difficult the development of new therapies. We here utilize dynamic BH3 Profiling (DBP), a functional predictive biomarker that measures net changes in mitochondrial apoptotic signaling, to identify anti-apoptotic adaptations upon treatment. We use this information to guide the use of BH3 mimetics to specifically inhibit BCL-2 pro-survival proteins, defeat resistance and avoid relapse to therapy. Indeed, we found that BH3 mimetics that selectively target BCL-xL and MCL-1 synergistically enhance the effect of the clinically used chemotherapeutic agents vincristine and doxorubicin in RMS cells. We validated this strategy in vivo using a RMS patient-derived xenograft (PDX) model and observed a reduction on tumor growth with a tendency to its stabilization with the sequential combination of vincristine and the MCL-1 inhibitor S63845. Finally, we identified the molecular mechanism by which RMS cells acquire resistance to vincristine: through the anti-apoptotic protein MCL-1 for which we observed an enhanced binding between MCL-1 and BID after drug exposure, which is suppressed by the sequential addition of S63845. In conclusion, our findings validate the use of DBP as a functional assay to predict treatment effectiveness in RMS and provide a rationale for BH3 mimetic combination with chemotherapeutic agents to avoid tumor resistance, improve treatment efficiency and decrease undesired secondary effects.

cancer biology↗