Impaired Sprouting and Axonal Atrophy in Cerebellar Climbing Fibres following In Vivo Silencing of the Growth-Associated Protein GAP-43

The adult mammalian central nervous system has a limited ability to establish new connections and to recover from traumatic or degenerative events. The olivo-cerebellar network represents an excellent model to investigate neuroprotection and repair in the brain during adulthood, due to its high plasticity and ordered synaptic organization. To shed light on the molecular mechanisms involved in these events, we focused on the growth-associated protein GAP-43 (also known as B-50 or neuromodulin). During development, this protein plays a crucial role in growth and in branch formation of neurites, while in the adult it is only expressed in a few brain regions, including the inferior olive (IO) where climbing fibres (CFs) originate. Following axotomy GAP-43 is usually up-regulated in association with regeneration. Here we describe an in vivo lentiviral-mediated gene silencing approach, used for the first time in the olivo-cerebellar system, to efficiently and specifically downregulate GAP-43 in rodents CFs. We show that lack of GAP-43 causes an atrophy of the CF in non-traumatic conditions, consisting in a decrease of its length, branching and number of synaptic boutons. We also investigated CF regenerative ability by inducing a subtotal lesion of the IO. Noteworthy, surviving CFs lacking GAP-43 were largely unable to sprout on surrounding Purkinje cells. Collectively, our results demonstrate that GAP-43 is essential both to maintain CFs structure in non-traumatic condition and to promote sprouting after partial lesion of the IO

The SARS-Unique Domain (SUD) of SARS Coronavirus Contains Two Macrodomains That Bind G-Quadruplexes

Since the outbreak of severe acute respiratory syndrome (SARS) in 2003, the three-dimensional structures of several of the replicase/transcriptase components of SARS coronavirus (SARS-CoV), the non-structural proteins (Nsps), have been determined. However, within the large Nsp3 (1922 amino-acid residues), the structure and function of the so-called SARS-unique domain (SUD) have remained elusive. SUD occurs only in SARS-CoV and the highly related viruses found in certain bats, but is absent from all other coronaviruses. Therefore, it has been speculated that it may be involved in the extreme pathogenicity of SARS-CoV, compared to other coronaviruses, most of which cause only mild infections in humans. In order to help elucidate the function of the SUD, we have determined crystal structures of fragment 389–652 (“SUDcore”) of Nsp3, which comprises 264 of the 338 residues of the domain. Both the monoclinic and triclinic crystal forms (2.2 and 2.8 Å resolution, respectively) revealed that SUDcore forms a homodimer. Each monomer consists of two subdomains, SUD-N and SUD-M, with a macrodomain fold similar to the SARS-CoV X-domain. However, in contrast to the latter, SUD fails to bind ADP-ribose, as determined by zone-interference gel electrophoresis. Instead, the entire SUDcore as well as its individual subdomains interact with oligonucleotides known to form G-quadruplexes. This includes oligodeoxy- as well as oligoribonucleotides. Mutations of selected lysine residues on the surface of the SUD-N subdomain lead to reduction of G-quadruplex binding, whereas mutations in the SUD-M subdomain abolish it. As there is no evidence for Nsp3 entering the nucleus of the host cell, the SARS-CoV genomic RNA or host-cell mRNA containing long G-stretches may be targets of SUD. The SARS-CoV genome is devoid of G-stretches longer than 5–6 nucleotides, but more extended G-stretches are found in the 3′-nontranslated regions of mRNAs coding for certain host-cell proteins involved in apoptosis or signal transduction, and have been shown to bind to SUD in vitro. Therefore, SUD may be involved in controlling the host cell's response to the viral infection. Possible interference with poly(ADP-ribose) polymerase-like domains is also discussed

PGC-1α and exercise in the control of body weight

The increasing prevalence of obesity and its comorbidities represents a major threat to human health globally. Pharmacological treatments exist to achieve weight loss, but the subsequent weight maintenance is prone to fail in the long run. Accordingly, efficient new strategies to persistently control body weight need to be elaborated. Exercise and dietary interventions constitute classical approaches to reduce and maintain body weight, yet people suffering from metabolic diseases are often unwilling or unable to move adequately. The administration of drugs that partially mimic exercise adaptation might circumvent this problem by easing and supporting physical activity. The thermogenic peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) largely mediates the adaptive response of skeletal muscle to endurance exercise and is a potential target for such interventions. Here, we review the role of PGC-1α in mediating exercise adaptation, coordinating metabolic circuits and enhancing thermogenic capacity in skeletal muscle. We suggest a combination of elevated muscle PGC-1α and exercise as a modified approach for the efficient long-term control of body weight and the treatment of the metabolic syndrome

Galantamine Attenuates Type 1 Diabetes and Inhibits Anti-Insulin Antibodies in Nonobese Diabetic Mice

Type 1 diabetes in mice is characterized by autoimmune destruction of insulin-producing pancreatic β-cells. Disease pathogenesis involves invasion of pancreatic islets by immune cells, including macrophages and T cells, and production of antibodies to self-antigens, including insulin. Activation of the inflammatory reflex, the neural circuit that inhibits inflammation, culminates on cholinergic receptor signals on immune cells to attenuate cytokine release and inhibit B-cell antibody production. Here, we show that galantamine, a centrally acting acetylcholinesterase inhibitor and an activator of the inflammatory reflex, attenuates murine experimental type 1 diabetes. Administration of galantamine to animals immunized with keyhole limpet hemocyanin (KLH) significantly suppressed splenocyte release of immunoglobulin G (IgG) and interleukin (IL)-4 and IL-6 during KLH challenge ex vivo. Administration of galantamine beginning at 1 month of age in nonobese diabetic (NOD) mice significantly delayed the onset of hyperglycemia, attenuated immune cell infiltration in pancreatic islets and decreased anti-insulin antibodies in serum. These observations indicate that galantamine attenuates experimental type 1 diabetes in mice and suggest that activation of the inflammatory reflex should be further studied as a potential therapeutic approach