Dissecting the role of sortilin receptor signaling in neurodegeneration induced by NGF deprivation.

Sortilin is a member of the family of vacuolar protein sorting 10 protein domain receptors which has emerged as a co-receptor in cell death and neurodegeneration processes mediated by proneurotrophins. Here we tested the possibility that sortilin deficiency interferes with behavioral and neuropathological endpoints in a chronic Nerve Growth factor (NGF)-deprivation model of Alzheimer's disease (AD), the AD10 anti-NGF mouse. AD10 mice show cholinergic deficit, increased APP processing and tau hyper-phosphorylation, resulting in behavioral deficits in learning and memory paradigms assessed by novel object recognition and Morris water maze tests. Sort1(-/-) mice were crossed with AD10 anti-NGF mice and the neurodegenerative phenotype was studied. We found that the loss of sortilin partially protected AD10 anti-NGF mice from neurodegeneration. A protective effect was observed on non-spatial memory as assessed by novel object recognition, and histopathologically at the level of Aß and BFCNs, while the phosphotau increase was unaltered by knocking out sortilin. We suggest that sortilin might be involved in different aspects of neurodegeneration in a complex way, supporting the view that sortilin functions in the CNS are broader than being a co-receptor in proneurotrophin and neurotrophin signaling.

SorLA deficiency dissects amyloid pathology from tau and cholinergic neurodegeneration in a mouse model of Alzheimer's disease.

Sortilin-related receptor with A-type repeats (SorLA, also known as LR11) has been implicated in Alzheimer's disease (AD). Thus, genetic studies associated SorLA gene variants with the risk of sporadic AD. Also, in vitro and in vivo studies showed that SorLA impairs processing of the amyloid-ß protein precursor (AßPP) to amyloid-ß. In particular, it has been found that loss of SorLA accelerates senile plaque deposition in mouse models overexpressing mutant forms of human AßPP and presenilin 1. Here we tested the possibility that SorLA deficiency also interferes with behavioral and neuropathological endpoints in an alternative murine AD model, the AD10 anti-nerve growth factor (NGF) mouse, in which amyloid-ß accumulation derives from the altered processing of endogenous AßPP. In addition to alterations in AßPP processing, AD10 mice also show cholinergic deficit and tau hyperphosphorylation resulting in behavioral deficits in learning and memory paradigms. We found that the loss of SorLA not only exacerbates early amyloid pathology but, at the same time, protects from cholinergic deficit and from early phospho-tau mislocalization. The results show that in the AD10 anti-NGF mouse model the AßPP processing-related aspects of neurodegeneration can be dissociated from those related to tau posttranslational processing and to cholinergic phenotypic maintenance by modulation of SorLA expression. We suggest that SorLA regulates different aspects of neurodegeneration in a complex way, supporting the hypothesis that SorLA expression might be critical not only for amyloid-related pathology but also for other cellular processes altered in AD.

Intranasal "painless" human Nerve Growth Factor [corrected] slows amyloid neurodegeneration and prevents memory deficits in App X PS1 mice.

Nerve Growth Factor (NGF) is being considered as a therapeutic candidate for Alzheimer's disease (AD) treatment but the clinical application is hindered by its potent pro-nociceptive activity. Thus, to reduce systemic exposure that would induce pain, in recent clinical studies NGF was administered through an invasive intracerebral gene-therapy approach. Our group demonstrated the feasibility of a non-invasive intranasal delivery of NGF in a mouse model of neurodegeneration. NGF therapeutic window could be further increased if its nociceptive effects could be avoided altogether. In this study we exploit forms of NGF, mutated at residue R100, inspired by the human genetic disease HSAN V (Hereditary Sensory Autonomic Neuropathy Type V), which would allow increasing the dose of NGF without triggering pain. We show that "painless" hNGF displays full neurotrophic and anti-amyloidogenic activities in neuronal cultures, and a reduced nociceptive activity in vivo. When administered intranasally to APPxPS1 mice ( n = 8), hNGFP61S/R100E prevents the progress of neurodegeneration and of behavioral deficits. These results demonstrate the in vivo neuroprotective and anti-amyloidogenic properties of hNGFR100 mutants and provide a rational basis for the development of "painless" hNGF variants as a new generation of therapeutics for neurodegenerative diseases.

Percutaneous coronary interventions with an endothelial progenitor cell capture stent (EPC) for high risk patients with no option for drug eluting stents: long term clinical outcomes of a single centre registry.

AIMS: Drug eluting stents (DES) are currently considered the gold standard for reducing restenosis of coronary artery lesions. Owing to their effect on the healing process, DES use requires mandatory prolonged dual antiplatelet therapy (DAT). The endothelial progenitor cell (EPC) capture stent, attracting circulating EPCs, promotes vascular healing and allows a short post-procedural period of DAT. The aim of the present study was to evaluate the short and long term clinical outcomes of the use of the Genos R stent™ in a selected high risk population with "no option" for DES. METHODS AND RESULTS: From December 2005 to October 2008, 61 high risk patients with clear contraindications to a prolonged period of DAT who underwent PCI with EPC capture stent implantation in our institution were prospectively selected and analysed. Technical success rate was 100%. Procedural success rate was 95.1%. After two years, major adverse cardiovascular events (MACE) free survival was 80.6%. According to the Academic Research Consortium definitions, cardiac death occurred in 1.6% of patients, and re-infarction, target lesion revascularisation (TLR), and target vessel revascularisation (TVR) occurred in 6.6%, 9.8%, and 11.5% of patients, respectively. Definite stent thrombosis occurred in one patient (specifically at 0 days). In patients who underwent surgery, no post-procedural MACE and no stent thrombosis were recorded. CONCLUSIONS: EPC capture stent implantation in high-risk patients with no option for DES seems encouraging, with satisfactory clinical outcomes both at short and at long term follow-up.

Transgenic mice with chronic NGF deprivation and Alzheimer's disease-like pathology display hippocampal region-specific impairments in short- and long-term plasticities.

The etiology of Alzheimer's disease (AD) remains elusive. The "amyloid" hypothesis states that toxic action of accumulated ß-amyloid peptide (Aß) on synaptic function causes AD cognitive decline. This hypothesis is supported by analysis of familial AD (FAD)-based transgenic mouse models, where altered amyloid precursor protein (APP) processing leads to Aß accumulation correlating with hippocampal-dependent memory deficits. Some studies report prominent dentate gyrus (DG) glutamatergic plasticity alterations in these mice, while CA1 plasticity remains relatively unaffected. The "neurotrophic unbalance" hypothesis, on the other hand, states that AD-related loss of cholinergic signaling and altered APP processing are due to alterations in nerve growth factor (NGF) trophic support. This hypothesis is supported by analysis of the AD11 mouse, which exhibits chronic NGF deprivation during adulthood and displays AD-like pathology, including Aß accumulation and hippocampal-dependent memory deficits. In this study, we analyzed CA1 and DG glutamatergic plasticity in AD11 mice to evaluate whether these mice also share with FAD models a common phenotype in hippocampal synaptic dysfunction. We report that AD11 mice display age-dependent short- and long-term DG plasticity deficits, while CA1 plasticity remains relatively spared. We also report that both structures exhibit enhanced glutamatergic transmission under lower, yet physiological, neurotransmitter release conditions, a defect that should be considered when further evaluating hippocampal synaptic deficits underlying AD pathology. We conclude that severe deficits in DG plasticity represent another common denominator between these two etiologically different types of AD mouse models, independent of the initial insult (overexpression of FAD mutation or NGF deprivation).

Dissecting the involvement of tropomyosin-related kinase A and p75 neurotrophin receptor signaling in NGF deficit-induced neurodegeneration.

NGF, the principal neurotrophic factor for basal forebrain cholinergic neurons (BFCNs), has been correlated to Alzheimer's disease (AD) because of the selective vulnerability of BFCNs in AD. These correlative links do not substantiate a comprehensive cause-effect mechanism connecting NGF deficit to overall AD neurodegeneration. A demonstration that neutralizing NGF activity could have consequences beyond a direct interference with the cholinergic system came from studies in the AD11 mouse model, in which the expression of a highly specific anti-NGF antibody determines a neurodegeneration that encompasses several features of human AD. Because the transgenic antibody binds to mature NGF much more strongly than to proNGF and prevents binding of mature NGF to the tropomyosin-related kinase A (TrkA) receptor and to p75 neurotrophin receptor (p75NTR), we postulated that neurodegeneration in AD11 mice is provoked by an imbalance of proNGF/NGF signaling and, consequently, of TrkA/p75NTR signaling. To test this hypothesis, in this study we characterize the phenotype of two lines of transgenic mice, one in which TrkA signaling is inhibited by neutralizing anti-TrkA antibodies and a second one in which anti-NGF mice were crossed to p75NTR(exonIII(-/-)) mice to abrogate p75NTR signaling. TrkA neutralization determines a strong cholinergic deficit and the appearance of beta-amyloid peptide (Abeta) but no tau-related pathology. In contrast, abrogating p75NTR signaling determines a full rescue of the cholinergic and Abeta phenotype of anti-NGF mice, but tau hyperphosphorylation is exacerbated. Thus, we demonstrate that inhibiting TrkA signaling activates Abeta accumulation and that different streams of AD neurodegeneration are related in complex ways to TrkA versus p75NTR signaling.

Peripheral neutralization of nerve growth factor induces immunosympathectomy and central neurodegeneration in transgenic mice.

We previously showed that anti-nerve growth factor (NGF) antibodies expressed in transgenic mice (AD11) elicit a progressive neurodegeneration, comprising the triad of Alzheimer's disease (AD) hallmarks: cholinergic loss, tau hyperphosphorylation, and amyloid-beta peptide formation. However, since anti-NGF antibodies are produced both in the brain and in peripheral tissues of AD11 mice, the contribution of peripheral neutralization of NGF to the onset of brain neurodegeneration was still unexplored. To address this question, we characterized a line of transgenic mice (AD10) in which anti-NGF antibodies are obligatorily produced only in lymphocytes, being initially found in blood. In AD10 mice, peripheral NGF neutralization elicits shrinkage of superior cervical ganglia (immunosympathectomy) and, as a consequence of this, peripheral anti-NGF antibodies cross the blood brain barrier (BBB) and reach the brain, generating an NGF-dependent neurodegeneration, largely superimposable to that observed in AD11 mice. This demonstrates that peripherally originated anti-NGF antibodies can generate a neurodegeneration in the central nervous system of an animal model. Consistently, peripherally-delivered NGF is effective in preventing the onset of the central cholinergic deficit. These findings could have a direct relevance for some human sporadic AD cases, highlighting the role of the BBB disruption and suggesting a causally relevant role of circulating antibodies in AD pathology.

[Current perspectives in cell therapy in cardiology: an overview of ongoing trials]. / Prospettive e difficoltà della terapia cellulare in Cardiologia: analisi critica delle sperimentazioni in corso.

BACKGROUND: Cardiac cell therapy is still restricted to experimental studies and clinical trials. This investigation is an analytic overview of worldwide ongoing trials and is aimed to show useful findings for planning new clinical trials. METHODS: Based on the online searching engine of the National Library of Medicine database for clinical trials (www.clinicaltrials.gov), all the ongoing studies registered were analyzed and classified. RESULTS: On May 30, 2009, 95 studies were registered worldwide: 40 of these were conducted in Europe and 4 in Italy. Phase 1, 2, 3 and 4 studies were 28%, 57%, 14% and 1%, respectively; observational and randomized studies were 22% and 78%, respectively. The clinical setting of enrolled patients was acute coronary syndrome in 43 studies (46%), heart failure in 29 studies (30%), chronic heart disease in 23 studies (24%). The treatment strategies investigated are cytokine-induced cell mobilization in 8% and cell implantation in 92%. The cell types used for direct implantation are mononuclear cells in 81%, endothelial precursor cells CD34+ in 11%, endothelial precursor cells CD133+ in 5%, myoblasts in 2% and adipoblasts in 1% of cases. CONCLUSIONS; The present snapshot of worldwide ongoing clinical trials provides useful information for planning an experimental clinical program on cardiac cell therapy.

In vitro receptor binding properties of a "painless" NGF mutein, linked to hereditary sensory autonomic neuropathy type V.

Nerve Growth Factor (NGF) signalling is mediated by the TrkA and p75NTR receptors. Besides its neurotrophic and survival activities, NGF displays a potent pro-nociceptive activity. Recently, a missense point mutation was found in the NGFB gene (C661T, leading to the aminoacid substitution R100W) of individuals affected by a form of hereditary loss of pain perception (hereditary sensory and autonomic neuropathy type V, HSAN V). In order to gain insights into the functional consequences of the HSAN V NGF mutation, two sets of hNGFR100 mutants were expressed in Escherichia coli and purified, as mature NGF or proNGF, for in vitro receptor binding studies. Here, we show by Surface Plasmon Resonance analysis that the R100 mutation selectively disrupts binding of hNGF to p75NTR receptor, to an extent which depends on the substituting residue at position 100, while the affinity of hNGFR100 mutants for TrkA receptor is not affected. As for unprocessed hproNGF, the binding of the R100 variants to p75NTR receptor shows only a limited impairment, showing that the impact of the R100 mutation on p75NTR receptor binding is greater in the context of mature, processed hNGF. These results provide a basis for elucidating the mechanisms underlying the clinical manifestations of HSAN V patients, and provide a basis for the development of "painless" hNGF molecules with therapeutic potential.

Delivery of NGF to the brain: intranasal versus ocular administration in anti-NGF transgenic mice.

Nerve growth factor (NGF) has a great potential for the treatment of Alzheimer's disease. However, the therapeutic administration of NGF represents a significant challenge, due to the difficulty to deliver relevant doses to the brain, in a safe and non-invasive way. We previously demonstrated the efficacy of a non-invasive delivery of NGF to the brain in animal models, by an intranasal route. Recently, topical eye application of NGF was proposed, as an option for the delivery of NGF to the brain. Here, we compare the efficacy of the two delivery routes of hNGF-61, a recombinant traceable form of human NGF, in the mouse neurodegeneration model AD11. The intranasal administration appeared to be significantly more effective than the ocular one, in rescuing the neurodegenerative phenotypic hallmarks in AD11 mice. The ocular administration of hNGF-61 showed a more limited efficacy, even at higher doses. Thus, NGF nasal drops represent a viable and effective option to successfully deliver therapeutic NGF to the brain in a non-invasive manner.