Biology subjects

Orsini, T.

Publications and source records attributed to Orsini, T..

4 recordsLinked to original sources

bioRxiv2023

Inhibition of cAMP signaling prevents congenital heart defects counteracting oxidative stress in Pde2A deficient embryos

BackgroundPhosphodiesterases (PDEs) are the enzymes that hydrolyze cyclic nucleotides (cAMP and cGMP) playing a key role in the homeostasis of these two second messengers. PDE2A is a dual-specific PDE that breaks down both cAMP and cGMP and can be activated by cGMP. It appears peculiar that the Pde2A-deficient (Pde2A-/-) mouse model is embryonically lethal, likely due to a strongly reduced size of liver and to a severe anemia. In addition, the heart of Pde2A-/- embryos shows ventricular and atrial septum defects, hypertrabeculation, heart dilatation and non-compaction defects. We recently highlighted a direct relationship between Pde2A impairment, consequent increase of cAMP and the onset of mouse congenital heart defects (CHDs), however the molecular mechanisms underlining the heart defects remain unknown. MethodsTranscriptome analysis of Pde2A-/- embryonic heart was performed by RNA sequencing and the most altered genes were also analyzed by quantitative real time PCR. In vivo treatment with drugs acting on cAMP signaling (Metoprolol and H89) and oxidative stress (N-Acetyl-Cysteine, NAC) were carried out on pregnant Pde2A+/- female. Histological, biochemical, and molecular analyses were then performed on embryonic hearts. ResultsWe found a significant modulation of more than 500 genes affecting biological processes involved in the immune system, cardiomyocyte development and contractility, angiogenesis, control of gene transcription and oxidative stress in hearts from Pde2A-/- embryos. Metoprolol and H89 administration were able to prevent heart dilatation and hypertabeculation in Pde2A-/-embryos. Metoprolol was also able to partially impede heart septum defect and oxidative stress at tissue and molecular levels. Partial rescue of cardiac defects was observed by using the antioxidant NAC, indicating oxidative stress like one of the molecular mechanisms underpinning the CHDs. ConclusionsWe identified specific biological processes, molecules and cell signaling that can be targeted by selected drugs with consequent beneficial effects for cAMP-dependent CHDs. Novelty and SignificanceO_ST_ABSWhat is Known?C_ST_ABSO_LICongenital Heart Defects are the most frequent heart birth defects including septal defects, hypertrabeculation and non-compacted myocardium. C_LIO_LIPde2A hydrolyses the cAMP and cGMP second messengers. C_LIO_LIPde2A-deficient mice are embryonic lethal and show cAMP-dependent Congenital Heart Defects. C_LI What New Information Does This Article Contribute?O_LIWe identified several novel pathways altered in hearts of Pde2A-/- embryos. C_LIO_LIWe demonstrated that drugs lowering cAMP levels rescued specific CHDs in Pde2A-/- embryos. C_LIO_LIWe discovered that antioxidants are beneficial for CHDs in Pde2A-/- embryos. C_LI The significance of this work relay in molecular discoveries and pharmacological approaches to treat CHDs by using a mouse model that recapitulate the major congenital heart defects. Among the pathways involved in specific defects associated with CHDs, the transcriptome analysis revealed an impairment of genes of the immune system, cardiomyocyte development and contractility, angiogenesis, control of gene transcription and oxidative stress in Pde2A-/- hearts. The scientific community will have open access to the RNA-seq data that can be utilized to further understand the congenital cardiac pathology and clarify the molecular implication in selected defects such as septal and ventricular wall defects. Up to date CHDs, when possible and if identified in time, are mostly treated trough surgery. The identification of drugs blunting the cAMP signaling response or reducing oxidative stress pathways will be useful for setting therapeutic approaches to alleviate CHDs.

developmental biology↗

bioRxiv2023

The scheduling of adolescence with Netrin-1 and UNC5C

Dopamine axons are the only axons known to grow during adolescence. Here, using rodent models, we examined how two proteins, Netrin-1 and its receptor, UNC5C, guide dopamine axons towards the prefrontal cortex and shape behaviour. We demonstrate in mice (Mus musculus) that dopamine axons reach the cortex through a transient gradient of Netrin-1 expressing cells - disrupting this gradient reroutes axons away from their target. Using a seasonal model (Siberian hamsters; Phodopus sungorus) we find that mesocortical dopamine development can be regulated by a natural environmental cue (daylength) in a sexually dimorphic manner - delayed in males, but advanced in females. The timings of dopamine axon growth and UNC5C expression are always phase-locked. Adolescence is an ill-defined, transitional period; we pinpoint neurodevelopmental markers underlying this period.

neuroscience↗

bioRxiv2022

Amphetamine disrupts dopamine axon growth in adolescence by a sex-specific mechanism

Initiating drug use during adolescence increases the risk of developing addiction and psychiatric disorders later in life, with long-term outcomes varying according to sex and exact timing of use. Even though most individuals begin experimenting with drugs of abuse in adolescence, to date, the cellular and molecular underpinnings explaining differential sensitivity to detrimental drug effects remain unknown. The Netrin-1/DCC guidance cue system plays a critical role in the adolescent development of mesocorticolimbic dopamine circuitry, segregating the cortical and limbic pathways. Adolescent experiences, including exposure to drugs of abuse, can regulate Dcc expression in male mice, placing Netrin-1/DCC signaling as a potential molecular link between experience and enduring changes to circuitry and behavior. Here we show that exposure to a recreational-like regimen of amphetamine (AMPH) in adolescence induces sex- and age-specific alterations in Dcc expression in the ventral tegmental area. Female mice are protected against the deleterious long-term effects of AMPH-induced Dcc regulation by compensatory changes in the expression of its binding partner, Netrin-1. AMPH induces targeting errors in mesolimbic dopamine axons and triggers their ectopic growth to the prefrontal cortex, only in early-adolescent male mice, underlying a male-specific vulnerability to its enduring cognitive effects. Upregulating DCC receptor expression in dopamine neurons in adolescent males using a neuron-optimized CRISPR/dCas9 Activation System induces female-like protection against the persistent effects of AMPH in early adolescence on inhibitory control. Netrin-1/DCC signaling is therefore a molecular switch which can be differentially regulated in response to the same experience as function of age and sex of the individual, leading to divergent long-term outcomes associated with vulnerable or resilient phenotypes.

neuroscience↗

bioRxiv2019

X-ray 3D imaging of gene expression in whole-mount murine brain by microCT, implication for functional analysis of tRNA endonuclease 54 gene mutated in pontocerebellar hypoplasia.

Acquisition of detailed structural and molecular information from intact biological samples, while preserving cellular three-dimensional structures, still represents a challenge for biological studies aiming to unravel system functions. Here we describe a novel X-ray-based methodology for analysis of gene expression pattern in intact murine brain ex vivo by microCT. The method relays on detection of bromine molecules in the products of enzymatic reaction generated by the {beta}-galactosidase (lacZ) gene reporter. To demonstrate the feasibility of the method, the analysis of the expression pattern of tRNA endonuclease 54 (Tsen54)-lacZ reporter gene in the whole-mount murine brain in semi-quantitative manner is performed. Mutations in Tsen54 gene causes pontocerebellar hypoplasia (PCH), severe neurodegenerative disorder with both mental and motor deficits. Comparing relative levels of Tsen54 gene expression, we have demonstrated that highest Tsen54 expression observed in anatomical brain substructures important for the normal motor and memory functions in mice. In the forebrain strong expression in perirhinal, retrosplenial and secondary motor areas was observed. In olfactory area Tsen54 is highly expressed in the nucleus of the lateral olfactory tract, anterior olfactory and bed nuclei, while in hypothalamus in lateral mammillary nucleus and preoptic area. In hindbrain Tsen54 is expressed in the reticular, cuneate and trigeminal nuclei of medulla, and in pontine gray of pons and in cerebellum, in the molecular and Purkinje cell layers. Delineating anatomical brain regions in which Tsen54 is strongly expressed will allow functionally address the role Tsen54 gene in normal physiology and in PCH disease. Significance StatementCharacterization of gene expression pattern in the brain of model organisms is critical for unravelling the gene function in normal physiology and disease. It is performed by optical imaging of the two-dimensional brain sections which then assembled in volume images. Here we applied microCT platform, which allows three-dimensional imaging of non transparent samples, for analysis of gene expression. This method based on detection by X-ray the bromine molecules presented in the products generated by enzymatic activity of b-galactosidase reporter gene. With this method we identify anatomical brain substructures in which Tsen54 gene, mutated in pontocerebellar hypoplasia disease, is expressed.

neuroscience↗