PKN1 Kinase: A Key Player in Adipocyte Differentiation and Glucose Metabolism.

Adipocyte dysfunction is the driver of obesity and correlates with insulin resistance and the onset of type 2 diabetes. Protein kinase N1 (PKN1) is a serine/threonine kinase that has been shown to contribute to Glut4 translocation to the membrane and glucose transport. Here, we evaluated the role of PKN1 in glucose metabolism under insulin-resistant conditions in primary visceral adipose tissue (VAT) from 31 patients with obesity and in murine 3T3-L1 adipocytes. In addition, in vitro studies in human VAT samples and mouse adipocytes were conducted to investigate the role of PKN1 in the adipogenic maturation process and glucose homeostasis control. We show that insulin-resistant adipocytes present a decrease in PKN1 activation levels compared to nondiabetic control counterparts. We further show that PKN1 controls the adipogenesis process and glucose metabolism. PKN1-silenced adipocytes present a decrease in both differentiation process and glucose uptake, with a concomitant decrease in the expression levels of adipogenic markers, such as PPARγ, FABP4, adiponectin and CEBPα. Altogether, these results point to PKN1 as a regulator of key signaling pathways involved in adipocyte differentiation and as an emerging player of adipocyte insulin responsiveness. These findings may provide new therapeutic approaches for the management of insulin resistance in type 2 diabetes.

proNGF Involvement in the Adult Neurogenesis Dysfunction in Alzheimer's Disease.

In recent decades, neurogenesis in the adult brain has been well demonstrated in a number of animal species, including humans. Interestingly, work with rodents has shown that adult neurogenesis in the dentate gyrus (DG) of the hippocampus is vital for some cognitive aspects, as increasing neurogenesis improves memory, while its disruption triggers the opposite effect. Adult neurogenesis declines with age and has been suggested to play a role in impaired progressive learning and memory loss seen in Alzheimer's disease (AD). Therefore, therapeutic strategies designed to boost adult hippocampal neurogenesis may be beneficial for the treatment of AD. The precursor forms of neurotrophins, such as pro-NGF, display remarkable increase during AD in the hippocampus and entorhinal cortex. In contrast to mature NGF, pro-NGF exerts adverse functions in survival, proliferation, and differentiation. Hence, we hypothesized that pro-NGF and its p75 neurotrophin receptor (p75NTR) contribute to disrupting adult hippocampal neurogenesis during AD. To test this hypothesis, in this study, we took advantage of the availability of mouse models of AD (APP/PS1), which display memory impairment, and AD human samples to address the role of pro-NGF/p75NTR signaling in different aspects of adult neurogenesis. First, we observed that DG doublecortin (DCX) + progenitors express p75NTR both, in healthy humans and control animals, although the percentage of DCX+ cells are significantly reduced in AD. Interestingly, the expression of p75NTR in these progenitors is significantly decreased in AD conditions compared to controls. In order to assess the contribution of the pro-NGF/p75NTR pathway to the memory deficits of APP/PS1 mice, we injected pro-NGF neutralizing antibodies (anti-proNGF) into the DG of control and APP/PS1 mice and animals are subjected to a Morris water maze test. Intriguingly, we observed that anti-pro-NGF significantly restored memory performance of APP/PS1 animals and significantly increase the percentage of DCX+ progenitors in the DG region of these animals. In summary, our results suggest that pro-NGF is involved in disrupting spatial memory in AD, at least in part by blocking adult neurogenesis. Moreover, we propose that adult neurogenesis alteration should be taken into consideration for better understanding of AD pathology. Additionally, we provide a new molecular entry point (pro-NGF/p75NTR signaling) as a promising therapeutic target in AD.

FLRT2 and FLRT3 Cooperate in Maintaining the Tangential Migratory Streams of Cortical Interneurons during Development.

Neuron migration is a hallmark of nervous system development that allows gathering of neurons from different origins for assembling of functional neuronal circuits. Cortical inhibitory interneurons arise in the ventral telencephalon and migrate tangentially forming three transient migratory streams in the cortex before reaching the final laminar destination. Although migration defects lead to the disruption of inhibitory circuits and are linked to aspects of psychiatric disorders such as autism and schizophrenia, the molecular mechanisms controlling cortical interneuron development and final layer positioning are incompletely understood. Here, we show that mouse embryos with a double deletion of FLRT2 and FLRT3 genes encoding cell adhesion molecules exhibit an abnormal distribution of interneurons within the streams during development, which in turn, affect the layering of somatostatin+ interneurons postnatally. Mechanistically, FLRT2 and FLRT3 proteins act in a noncell-autonomous manner, possibly through a repulsive mechanism. In support of such a conclusion, double knockouts deficient in the repulsive receptors for FLRTs, Unc5B and Unc5D, also display interneuron defects during development, similar to the FLRT2/FLRT3 mutants. Moreover, FLRT proteins are chemorepellent ligands for developing interneurons in vitro, an effect that is in part dependent on FLRT-Unc5 interaction. Together, we propose that FLRTs act through Unc5 receptors to control cortical interneuron distribution in a mechanism that involves cell repulsion.SIGNIFICANCE STATEMENT Disruption of inhibitory cortical circuits is responsible for some aspects of psychiatric disorders such as schizophrenia or autism. These defects include interneuron migration during development. A crucial step during this process is the formation of three transient migratory streams within the developing cortex that determine the timing of interneuron final positioning and the formation of functional cortical circuits in the adult. We report that FLRT proteins are required for the proper distribution of interneurons within the cortical migratory streams and for the final laminar allocation in the postnatal cortex. These results expand the multifunctional role of FLRTs during nervous system development in addition to the role of FLRTs in axon guidance and the migration of excitatory cortical neurons.

Genetic ablation of the Rho GTPase Rnd3 triggers developmental defects in internal capsule and the globus pallidus formation.

The forebrain includes the cerebral cortex, the thalamus, and the striatum and globus pallidus (GP) in the subpallium. The formation of these structures and their interconnections by specific axonal tracts take place in a precise and orchestrated time and spatial-dependent manner during development. However, the knowledge of the molecular and cellular mechanisms that are involved is rather limited. Moreover, while many extracellular cues and specific receptors have been shown to play a role in different aspects of nervous system development, including neuron migration and axon guidance, examples of intracellular signaling effectors involved in these processes are sparse. In the present work, we have shown that the atypical RhoGTPase, Rnd3, is expressed very early during brain development and keeps a dynamic expression in several brain regions including the cortex, the thalamus, and the subpallium. By using a gene-trap allele (Rnd3gt ) and immunological techniques, we have shown that Rnd3gt/gt embryos display severe defects in striatal and thalamocortical axonal projections (SAs and TCAs, respectively) and defects in GP formation already at early stages. Surprisingly, the corridor, an important intermediate target for TCAs is still present in these mutants. Mechanistically, a conditional genetic deletion approach revealed that Rnd3 is primarily required for the normal development of Medial Ganglionic Eminence-derived structures, such as the GP, and therefore acts non-cell autonomously in SAs and TCAs. In conclusion, we have demonstrated the important role of Rnd3 as an early regulator of subpallium development in vivo and revealed new insights about SAs and TCAs development.

proBDNF is modified by advanced glycation end products in Alzheimer's disease and causes neuronal apoptosis by inducing p75 neurotrophin receptor processing.

Alzheimer disease (AD) is a complex pathology related to multiple causes including oxidative stress. Brain-derived neurotrophic factor (BDNF) is a neutrotrophic factor essential for the survival and differentiation of neurons and is considered a key target in the pathophysiology of various neurodegenerative diseases, as for example AD. Contrarily to BDNF, the precursor form of BDNF (proBDNF) induces apoptosis through the specific interaction with p75 and its co-receptor, Sortilin.We used hippocampal tissue and cerebrospinal fluid from AD patients and controls. to study the localization and the levels of proBDNF, p75 and Sortilin as well as the post-traduccional modifications of proBDNF induced by Radical Oxygen Species, by immunofluorescence and Western blot. Differentiation and survival were assessed on differentiated mouse hippocampal neurons derived from postnatal neural stem cells from WT animals or from the transgenic AD animal model APP/PS1∆E9, based on mutations of familiar AD. In AD patients we observe a significative increase of proBDNF and Sortilin expression and a significative increase of the ratio proBDNF/BDNF in their cerebrospinal fluid compared to controls. In addition, the proBDNF of AD patients is modified by ROS-derived advanced glycation end products, which prevent the processing of the proBDNF to the mature BDNF, leading to an increase of pathogenicity and a decrease of trophic effects. The cerebrospinal fluid from AD patients, but not from controls, induces apoptosis in differentiated hippocampal neurons mainly by the action of AGE-modified proBDNF present in the cerebrospinal fluid of the patients. This effect is triggered by the activation and processing of p75 that stimulate the internalization of the intracellular domain (ICD) within the nucleus causing apoptosis. Induction of apoptosis and p75 ICD internalization by AD patients-derived proBDNF is further enhanced in neuron cultures from the AD model expressing the APP/PS1∆E9 transgene.Our results indicate the importance of proBDNF neurotoxic signaling in AD pathology essentially by three mechanisms: i) by an increase of proBDNF stability due to ROS-induced post-traductional modifications; ii) by the increase of expression of the p75 co-receptor, Sortilin and iii) by the increase of the basal levels of p75 processing found in AD.

Oxidative stress and neurodegenerative diseases: a neurotrophic approach.

Neurotrophins are important neurotrophic factors involved in the survival, differentiation and function of a wide variety of neuron populations. A common feature for most neurotrophins is that they are synthesized as precursor proteins (pro-neurotrophins) that upon being processed by proteolysis render the mature active form responsible for most of their trophic functions. However, some of the pro-neurotrophin form of these proteins, such as the precursor form of NGF (pro-NGF), have been shown to induce opposite effects and trigger apoptosis on neurons through the p75NTR receptor. This suggests that the balance between the levels of proneurotrophin and neurotrophin must be tightly controlled. In this context, it has been shown that in conditions of oxidative stress due for instance to aging or the development of some neurodegenerative disease, neurotrophins are oxidatively modified at least by advanced glycation/lipoxidation end products (AGE/ALEs) which makes pro-NGF refractary to be processed. The lack of maturation and the imbalance in favor of the precursor form may change the pattern of active signaling pathways towards cell death, thus exacerbating the deleterious alterations, for instance during the development of neurodegenerative diseases. Besides that, AGE/ALEs also induce the processing of the pro-NGF receptor p75NTR by α- secretase which is followed by the processing by γ -secretase and the release of the intracellular domain of p75NTR (p75NTRICD). Once cleaved, p75NTRICD recruits two intracellular interactors, NRIF and TRAF6, which allows NRIF phosphorylation by JNK. The phosphorylated form of NRIF then translocates to the nucleus and induces the expression of pro-apoptotic proteins. In this chapter we will summarize the mechanisms by which ROS- induce protein modifications, which proteins are susceptible to be modified, how these modifications affect function and signaling and, finally, how they can be related to neurodegenerative diseases.