Decrease of Pdzrn3 is required for heart maturation and protects against heart failure

Heart failure is the final common stage of most cardiopathies. Cardiomyocytes (CM) connect with others via their extremities by intercalated disk protein complexes. This planar and directional organization of myocytes is crucial for mechanical coupling and anisotropic conduction of the electric signal in the heart. One of the hallmarks of heart failure is alterations in the contact sites between CM. Yet no factor on its own is known to coordinate CM polarized organization. We have previously shown that PDZRN3, an ubiquitine ligase E3 expressed in various tissues including the heart, mediates a branch of the Planar cell polarity (PCP) signaling involved in tissue patterning, instructing cell polarity and cell polar organization within a tissue. PDZRN3 is expressed in the embryonic mouse heart then its expression dropped significantly postnatally corresponding with heart maturation and CM polarized elongation. A moderate CM overexpression of Pdzrn3 (Pdzrn3 OE) during the first week of life, induced a severe eccentric hypertrophic phenotype with heart failure. In models of pressure-overload stress heart failure, CM-specific Pdzrn3 knockout showed complete protection against degradation of heart function. We reported that Pdzrn3 signaling induced PKC ζ expression, c-Jun nuclear translocation and a reduced nuclear ß catenin level, consistent markers of the planar non-canonical Wnt signaling in CM. We then show that subcellular localization (intercalated disk) of junction proteins as Cx43, ZO1 and Desmoglein 2 was altered in Pdzrn3 OE mice, which provides a molecular explanation for impaired CM polarization in these mice. Our results reveal a novel signaling pathway that controls a genetic program essential for heart maturation and maintenance of overall geometry, as well as the contractile function of CM, and implicates PDZRN3 as a potential therapeutic target for the prevention of human heart failure.

Gata2, Nkx2-2 and Skor2 form a transcription factor network regulating development of a midbrain GABAergic neuron subtype with characteristics of REM sleep regulatory neurons

The midbrain reticular formation is a mosaic of diverse GABAergic and glutamatergic neurons that have been associated with a variety of functions, including the regulation of sleep. However the molecular characteristics and development of the midbrain reticular formation neurons are poorly understood. As the transcription factor Gata2 is required for the development of all GABAergic neurons derived from the embryonic mouse midbrain, we hypothesized that the genes expressed downstream of Gata2 could contribute to the diversification of GABAergic neuron subtypes in this brain region. Here, we show that Gata2 is indeed required for the expression of several lineage-specific transcription factors in post-mitotic midbrain GABAergic neuron precursors. These include a homeodomain transcription factor Nkx2-2 and a SKI family transcriptional repressor Skor2, which are co-expressed in a restricted group of GABAergic precursors in the midbrain reticular formation. Both Gata2, and Nkx2-2 function is required for the expression of Skor2 in GABAergic precursors. In the adult mouse as well as rat midbrain, the Nkx2-2 and Skor2 expressing GABAergic neurons locate at the boundary of the ventrolateral periaqueductal gray and the midbrain reticular formation, an area shown to contain REM-off neurons regulating REM sleep. In addition to the characteristic localization, the Skor2 positive cells increase their activity upon REM sleep inhibition, send projections to a pontine region associated with sleep control and are responsive to orexins, consistent with the known properties of the midbrain REM-off neurons.

Activin A and ALK4 Identified as Novel Regulators of Epithelial to Mesenchymal Transition (EMT) in Human Epicardial Cells

The epicardium, the mesothelial layer covering the heart, is a crucial cell source for cardiac development and repair. It provides cells and biochemical signals to the heart to facilitate vascularization and myocardial growth. An essential element of epicardial behavior is epicardial epithelial to mesenchymal transition (epiMT), which is the initial step for epicardial cells to become motile and invade the myocardium. To identify targets to optimize epicardium-driven repair of the heart, it is vital to understand which pathways are involved in the regulation of epiMT. Therefore, we established a cell culture model for human primary adult and fetal epiMT, which allows for parallel testing of inhibitors and stimulants of specific pathways. Using this approach, we reveal Activin A and ALK4 signaling as novel regulators of epiMT, independent of the commonly accepted EMT inducer TGFβ. Importantly, Activin A was able to induce epicardial invasion in cultured embryonic mouse hearts. Our results identify Activin A/ALK4 signaling as a modulator of epicardial plasticity which may be exploitable in cardiac regenerative medicine.

Single-cell delineation of lineage and genetic identity in the mouse brain

During neurogenesis, mitotic progenitor cells lining the ventricles of the embryonic mouse brain undergo their final rounds of cell division, giving rise to a wide spectrum of postmitotic neurons and glia1,2. The link between developmental lineage and cell-type diversity remains an open question. Here we used massively parallel tagging of progenitors to track clonal relationships and transcriptomic signatures during mouse forebrain development. We quantified clonal divergence and convergence across all major cell classes postnatally, and found diverse types of GABAergic neuron that share a common lineage. Divergence of GABAergic clones occurred during embryogenesis upon cell-cycle exit, suggesting that differentiation into subtypes is initiated as a lineage-dependent process at the progenitor cell level.

Brain injury environment critically influences the connectivity of transplanted neurons

Cell transplantation is a promising approach for the reconstruction of neuronal circuits after brain damage. Transplanted neurons integrate with remarkable specificity into circuitries of the mouse cerebral cortex affected by neuronal ablation. However, it remains unclear how neurons perform in a local environment undergoing reactive gliosis, inflammation, macrophage infiltration and scar formation, as in traumatic brain injury (TBI). To elucidate this, we transplanted cells from the embryonic mouse cerebral cortex into TBI-injured, inflamed-only, or intact cortex of adult mice. Brain-wide quantitative connectomics unraveled graft inputs from correct regions across the brain in all conditions, with pronounced quantitative differences: scarce in intact and inflamed brain, versus exuberant after trauma. In the latter, excessive synapse pruning follows the initial overshoot of connectivity resulting in only a few connections left. Proteomic profiling identifies candidate molecules involved in the synaptic yield, a pivotal parameter to tailor for functional restoration of neuronal circuits.

BMP4 and WNT signaling interact to promote mouse tracheal mesenchyme morphogenesis

Tracheobronchomalacia and Complete Tracheal Rings are congenital malformations of the trachea associated with morbidity and mortality for which the etiology remains poorly understood. Epithelial expression of Wls (a cargo receptor mediating Wnt ligand secretion) by tracheal cells is essential for patterning the embryonic mouse trachea's cartilage and muscle. RNA sequencing indicated that Wls differentially modulated the expression of BMP signaling molecules. We tested whether BMP signaling, induced by epithelial Wnt ligands, mediates cartilage formation. Deletion of Bmp4 from respiratory tract mesenchyme impaired tracheal cartilage formation that was replaced by ectopic smooth muscle, recapitulating the phenotype observed after epithelial deletion of Wls in the embryonic trachea. Ectopic muscle was caused in part by anomalous differentiation and proliferation of smooth muscle progenitors rather than tracheal cartilage progenitors. Mesenchymal deletion of Bmp4 impaired expression of Wnt/β-catenin target genes, including targets of WNTsignaling: Notum, and Axin2. In vitro, rBMP4 rescued the expression of Notum in Bmp4 deficient tracheal mesenchymal cells and induced Notum promoter activity via SMAD1/5. RNA sequencing of Bmp4 deficient tracheas identified genes essential for chondrogenesis and muscle development co-regulated by BMP and WNT signaling. During tracheal morphogenesis, WNT signaling induces Bmp4 in mesenchymal progenitors to promote cartilage differentiation and restrict trachealis muscle. In turn, Bmp4 differentially regulates the expression of Wnt/β-catenin targets to attenuate mesenchymal WNT signaling and to further support chondrogenesis.

Dimethyl Fumarate is a Potential Therapeutic Option for Alzheimer’s Disease

Background: Dimethyl fumarate (DMF) has been approved for clinical treatment of multiple sclerosis based on its antioxidant and anti-inflammatory effects by activating the Nrf2 pathway. Since both oxidative stress and inflammation are involved in Alzheimer’s disease (AD), DMF is a potential therapeutic option for AD. Objective: This study aims to test the therapeutic effects of DMF on AD model mice and to reveal its underlying molecular mechanisms. Methods: Cell viability assay and in vitro immunofluorescence imaging were used to evaluate the antioxidant effect of DMF on embryonic mouse hippocampal neurons. Behavioral test and brain magnetic resonance imaging were used to assess the therapeutic effects of DMF on spatial learning and memory as well as hippocampal volume in AD model mice with and without Nrf2 knockdown. Western blotting was used to analyze the expression of antioxidant enzymes and molecules associated with AD-related pathological pathways. Results: DMF inhibits reactive oxygen species overproduction and protects neurons without Nrf2 knockdown from death. DMF reduces amyloid-β induced memory impairment and hippocampal atrophy in AD model mice rather than in Nrf2 knockdown AD mice. DMF delays the progression of AD by activating the Nrf2 pathway to enhance the expression of downstream antioxidant enzymes and inhibits lipid peroxidation, apoptosis, inflammation, mitochondrial dysfunction and amyloid-β deposition. Conclusion: These results indicate that DMF is a potential therapeutic option for AD through its antioxidant, anti-inflammatory, anti-apoptotic, and other anti-AD effects by activating the Nrf2 pathway.

Temporal and Spatial Transcriptomic Dynamics across Brain Development in Xenopus laevis tadpoles

Amphibian metamorphosis is a transitional period that involves significant changes in the cell type populations and biological processes occurring in the brain. Analysis of gene expression dynamics during this process may provide insight into the molecular events underlying these changes. We conducted differential gene expression analyses of the developing X. laevis tadpole brain during this period in two ways: first, over stages of development in the midbrain, and second, across regions of the brain at a single developmental stage. We found that genes pertaining to positive regulation of neural progenitor cell proliferation as well as known progenitor cell markers were upregulated in the midbrain prior to metamorphic climax; concurrently, expression of cell cycle timing regulators decreased across this period, supporting the notion that cell cycle lengthening contributes to a decrease in proliferation by the end of metamorphosis. We also found that at the start of metamorphosis, neural progenitor populations appeared to be similar across the fore-, mid-, and hindbrain regions. Genes pertaining to negative regulation of differentiation were upregulated in the spinal cord compared to the rest of the brain, however, suggesting that a different program may regulate neurogenesis there. Finally, we found that regulation of biological processes like cell fate commitment and synaptic signaling follow similar trajectories in the brain across early tadpole metamorphosis and mid- to late-embryonic mouse development. By comparing expression across both temporal and spatial conditions, we have been able to illuminate cell type and biological pathway dynamics in the brain during metamorphosis.

Tuberin levels during cellular differentiation in brain development

Tuberin is a member of a large protein complex, Tuberous Sclerosis Complex, and acts as a sensor for nutrient status regulating protein synthesis and cell cycle progression. Mutations in the Tuberin gene, TSC2, lead to the formation of tumors and developmental defects in many organ systems, including the central nervous system. Tuberin is expressed in the brain throughout development and levels of Tuberin have been found to decrease during neuronal differentiation in cell lines in vitro. Our current work investigates the levels of Tuberin at two stages of embryonic development in vivo, and we study the mRNA and protein levels during a time course using immortalized cell lines in vitro. Our results show that Tuberin levels remain stable in the olfactory bulb but decrease in the Purkinje cell layer during embryonic mouse brain development. We show here that Tuberin levels are higher when cells are cultured as neurospheres, and knockdown of Tuberin results in a reduction in the number of neurospheres. These data provide support for the hypothesis that Tuberin is an important regulator of stemness and the reduction of Tuberin levels might support functional differentiation in the central nervous system. Understanding how Tuberin expression is regulated throughout neural development is essential to fully comprehend the role of this protein in several developmental and neural pathologies.