LOX and LOX-Like Proteins as Potential Therapeutic Target for Atrial Fibrillation

Les lysyl-oxydases (LOX) et LOX-like (LOXL-1, 2, 3 et 4) influencent le remodelage de la matrice extracellulaire (MEC) lors d’anomalies cardiaques comme l’insuffisance cardiaque (IC) ou la fibrose. L’objectif principal était d’étudier les fonctions matrice-dépendantes et -indépendantes des LOX et LOXL dans la transduction des signaux favorisant la fibrose et la fibrillation atriales (FA). Dans l’oreillette gauche (OG) de chiens IC, nous avons observé une augmentation de la régulation de : LOX et LOXL-1 dans le tissue atrial, LOX et LOXL-2 dans les fibroblastes ainsi que LOX, LOXL-1, LOXL-3 et LOXL-4 dans les myocytes. Nous avons évalué le rôle des isoformes des LOX dans la signalisation de la fibrose et de la FA dans l’OG de rats avec infarctus du myocarde (IM). Chez le chien et les cellules cardiaques de rats néonataux, nous avons exploré le rôle des LOX et LOXL sur la fonction matrice-dépendante et -indépendante des fibroblastes et myocytes cardiaques, en utilisant un traitement à l’angiotensine-II (Ang II) et un knockdown par si-ARN des isoformes de LOX. L’augmentation de l’expression des ARNm de LOX et LOXL était associée à une augmentation de l’expression des ARNm de COL 1A1, FN 1, TGF-β1, CTGF, periostin, α-SMA et MMP-2 dans la zone infarcie du ventricule gauche (VG). L’expression des ARNm de LOXL-1, LOXL-3, COL 1A1, TGF-β1 et periostin était significativement augmentée dans l’OG. L’administration de β-aminopropionitrile (BAPN) post-IM a diminué significativement l’expression des ARNm de LOXL-1,2,3. Le BAPN a aussi diminué l’expression d’ARNm de marqueurs pro-fibrotiques dans l’OG. Ces changements n’étaient pas significatifs dans le VG. Le BAPN a diminué la fibrose dans l’OG ainsi que le ratio de cross-linking du collagène, mais pas significativement dans le VG. Le BAPN a réduit les remodelages structuraux et fonctionnels de l’OG sans influencer significativement ceux-ci dans le VG. L’IM était associé à une augmentation de la durée de l’onde P ainsi que la durée et l’inductibilité des FA que le BAPN a significativement réduit. Chez des rats néonataux, des fibroblastes et des myocytes de ventricule ont été mis en culture et traités à l’Ang II. Les LOX et LOXL-2 sécrétées et le ratio de cross-linking du collagène ont été augmentés; et contribueraient au remodelage de la MEC. Chez le chien, la contractilité des myocytes de l’OG a été augmentée suivant le knockdown de LOX et LOXL-1 associé à des changements de concentrations calciques. Dans les fibroblastes, le knockdown de LOXL-3 a réduit l’expression des ARNm de LOXL-2,3,4, associé à une réduction de la prolifération cellulaire et de l’expression des ARNm de COL 1A1, COL 3A1 et CCNE 2. Ces résultats suggèrent que LOXL-2,3,4 influenceraient la prolifération des fibroblastes et la synthèse du collagène. Le knockdown de LOXL-4 a augmenté ratio de l’expression en ARNm de BAX/BLC-2, ainsi LOXL-4 pourrait avoir un effet protecteur contre l’apoptose. En conclusion, les LOX et LOXL sont des médiateurs potentiels de la fibrose et la FA dans l’OG chez le rat infarci. Les LOX et LOXL seraient alors impliqués dans la régulation des fonctions des fibroblastes et myocytes cardiaques.Lysyl oxidase (LOX) and LOX-like (LOXL-1, 2, 3 and 4) proteins have a crucial role in extracellular matrix (ECM) remodeling in several types of heart disease, such as heart failure (HF) and fibrosis. The main objective of this thesis was to address the matrix-dependent and matrix-independent functions of LOX and LOXL proteins in signal transduction, leading to atrial fibrosis and atrial fibrillation (AF). We noted upregulation of LOX and LOXL-1 in tissues, LOX and LOXL-2 in fibroblasts and LOX, LOXL-1, LOXL-3 and LOXL-4 in myocytes of the canine left atrium (LA) in congestive heart failure (CHF). Based on these findings, we studied the roles of LOX isoforms as upstream targets in the signaling pathways of LA fibrosis and AF in a rat model following myocardial infarction (MI). Additionally, we explored the physiological roles of LOX and LOXL proteins in matrix-dependent and matrix-independent functions of cardiac fibroblasts and myocytes through angiotensin II (Ang II) treatment and siRNA-mediated knockdown of LOX isoforms in canine and neonatal rat cells. Upregulation of the mRNA expression of LOX and LOXL was accompanied by an increase in mRNA expression of COL 1A1, FN 1, TGF-β1, CTGF, periostin, α-SMA and MMP-2 in the infarcted area of the left ventricle (LV). mRNA expression of LOXL-1, LOXL-3, COL 1A1, TGF-β1 and periostin were significantly increased in the LA post-MI. Administration of β-aminopropionitrile (BAPN) post-MI significantly reduced the mRNA expression of LOXL-1, LOXL-2 and LOXL-3 along with a decrease in the mRNA expression of several profibrotic markers in the LA without significant changes to those in the LV. Moreover, the administration of BAPN post-MI reduced LA fibrosis and the collagen cross-linking ratio without significantly changing those in the LV. BAPN administration post-MI reduced the adverse structural and functional remodeling of LA without significantly changing those in the LV. Furthermore, MI caused an increase in the P-wave duration, AF duration and AF inducibility, while the values of those parameters were significantly reduced upon BAPN administration post-MI. The protein expression of secreted LOX and LOXL-2 were increased in cultured neonatal rat ventricular fibroblasts and myocytes along with an increase in the collagen cross-linking ratio in fibroblasts upon treatment with Ang II. The secretion of LOX and LOXL-2 from cardiac fibroblasts and myocytes may contribute to ECM remodeling. Moreover, the contractility of canine LA myocytes was enhanced upon knockdown of LOX or LOXL-1 along with slight changes in Ca2+ transients. Upon knockdown of LOXL-3 in fibroblasts, LOXL-2, LOXL-3 and LOXL-4 mRNA expression levels were reduced along with reduced proliferation and mRNA expression of COL 1A1, COL 3A1 and CCNE 2. The results showed that LOXL-2, LOXL-3 and LOXL-4 may promote fibroblast proliferation and collagen synthesis. LOXL-4 knockdown increased the mRNA expression of the BAX/BCL-2 ratio, suggesting that LOXL-4 may protect against apoptosis in cardiac fibroblasts and myocytes. In conclusion, LOX and LOXL proteins are prominent mediators of LA fibrosis and AF post-MI. Additionally, these findings suggested that LOX and LOXL proteins are implicated in the regulation of various aspects of cardiac fibroblast and myocyte functions

NMDA Receptors in Astroglia: Chronology, Controversies, and Contradictions from a Complex Molecule

The neurocentric theory dismissed for decades the role of glia in information handling within the central nervous system (CNS). Nevertheless, almost 3 decades ago, this started to change and today astrocytes are considered relevant players for this function. Astrocytes “listen” to neuronal communication, regulate it, and respond at the cellular and synctitial level. Ionotropic glutamate NMDA receptor (NMDAR) is critical in CNS. It mediates synaptic neuronal communication and it is involved in different mechanisms. However, NMDAR is also expressed by astrocytes, but its functional role in these cells has not been deeply investigated and has been a matter of debate in the last decades. In this chapter, we briefly outline NMDAR intracellular transduction pathways initiated by Ca2+ flux. Then, we review chronologically NMDAR expression and function in astrocytes that have been a source of controversies and apparent contradictions. Finally, some insights are presented regarding NMDAR in astrocytes in the context of the tripartite synapse concept and the recently described Ca2+ flux–independent metabotropic-like NMDAR function in astrocytes. Given the complex molecular nature of NMDAR, its critical role, and the relevance of astrocytes, the study of astrocytic NMDAR promises to provide further understanding of CNS physiology and pathology

Connexins: synthesis, post-translational modifications, and trafficking in health and disease

Connexins are tetraspan transmembrane proteins that form gap junctions and facilitate direct intercellular communication, a critical feature for the development, function, and homeostasis of tissues and organs. In addition, a growing number of gap junction-independent functions are being ascribed to these proteins. The connexin gene family is under extensive regulation at the transcriptional and post-transcriptional level, and undergoes numerous modifications at the protein level, including phosphorylation, which ultimately affects their trafficking, stability, and function. Here, we summarize these key regulatory events, with emphasis on how these affect connexin multifunctionality in health and disease

Mechanisms involved in the adsorption of bioinsecticides and strategies to enhance their passage through the insect midgut.

During the past decade a number of products have emerged from the effort to develop alternative biopesticidal technologies. The most recent approach to integrated pest management is based on the detection of new genes encoding for polypeptides with potential insecticidal activity, with a particular attention to biopesticides derived from viruses, microorganisms, fungi, plants and insects. Bacteria and viruses themselves can be used for biological control purpose. In most cases, biopesticides are molecules that have haemocoelic targets and must pass undegraded the gut barrier in order to exert their activity. Therefore for an effective oral delivery of these molecules it will be crucial to develop basic information on the molecular mechanisms mediating the absorption of proteins and peptides by the insect gut and to develop strategies to facilitate their passage through the midgut barrier. These issues have been the subject of my research project during these three years. In particular, I characterized the strategies adopted by Junonia coenia densovirus (JcDNV) to cross the midgut barrier of the lepidopteran pest Spodoptera frugiperda. I also studied a possible mechanism to increase the protein uptake in insect midgut characterizing the mechanisms involved in the internalization of selected Cell Penetrating Peptides (Tat, Penetratin, L-oligoarginine and D-oligoarginine) through the plasma membrane of lepidopteran larveae midgut cells. Even though these two aspects represent the main part of my PhD project, I have also collaborated to the study devoted to the identification of strategies to increase the permeability of the paracellular route

MorphDB : prioritizing genes for specialized metabolism pathways and gene ontology categories in plants

Recent times have seen an enormous growth of "omics" data, of which high-throughput gene expression data are arguably the most important from a functional perspective. Despite huge improvements in computational techniques for the functional classification of gene sequences, common similarity-based methods often fall short of providing full and reliable functional information. Recently, the combination of comparative genomics with approaches in functional genomics has received considerable interest for gene function analysis, leveraging both gene expression based guilt-by-association methods and annotation efforts in closely related model organisms. Besides the identification of missing genes in pathways, these methods also typically enable the discovery of biological regulators (i.e., transcription factors or signaling genes). A previously built guilt-by-association method is MORPH, which was proven to be an efficient algorithm that performs particularly well in identifying and prioritizing missing genes in plant metabolic pathways. Here, we present MorphDB, a resource where MORPH-based candidate genes for large-scale functional annotations (Gene Ontology, MapMan bins) are integrated across multiple plant species. Besides a gene centric query utility, we present a comparative network approach that enables researchers to efficiently browse MORPH predictions across functional gene sets and species, facilitating efficient gene discovery and candidate gene prioritization. MorphDB is available at http://bioinformatics.psb.ugent.be/webtools/morphdb/morphDB/index/. We also provide a toolkit, named "MORPH bulk" (https://github.com/arzwa/morph-bulk), for running MORPH in bulk mode on novel data sets, enabling researchers to apply MORPH to their own species of interest