Characteristic Hallmarks of Aging and the Impact on Carcinogenesis.

Evidence shows that there is a synergistic, bidirectional association between cancer and aging with many shared traits. Age itself is a risk factor for the onset of most cancers, while evidence suggests that cancer and its treatments might accelerate aging by causing genotoxic and cytotoxic insults. Aging has been associated with a series of alterations that can be linked to cancer: i) genomic instability caused by DNA damage or epigenetic alterations coupled with repair errors, which lead to progressive accumulation of mutations; ii) telomere attrition with possible impairment of telomerase, shelterin complex, or the trimeric complex (Cdc13, Stn1 and Ten1 - CST) activities associated with abnormalities in DNA replication and repair; iii) altered proteostasis, especially when leading to an augmented proteasome, chaperon and autophagy-lysosome activity; iv) mitochondrial dysfunction causing oxidative stress; v) cellular senescence; vi) stem cells exhaustion, intercellular altered communication and deregulated nutrient sensing which are associated with microenvironmental modifications which may facilitate the subsequential role of cancer stem cells. Nowadays, anti-growth factor agents and epigenetic therapies seem to assume an increasing role in fighting aging-related diseases, especially cancer. This report aims to discuss the impact of age on cancer growth.

NGF Prevents Loss of TrkA/VEGFR2 Cells, and VEGF Isoform Dysregulation in the Retina of Adult Diabetic Rats.

Among the factors involved in diabetic retinopathy (DR), nerve growth factor (NGF) and vascular endothelial growth factor A (VEGFA) have been shown to affect both neuronal survival and vascular function, suggesting that their crosstalk might influence DR outcomes. To address this question, the administration of eye drops containing NGF (ed-NGF) to adult Sprague Dawley rats receiving streptozotocin (STZ) intraperitoneal injection was used as an experimental paradigm to investigate NGF modulation of VEGFA and its receptor VEGFR2 expression. We show that ed-NGF treatment prevents the histological and vascular alterations in STZ retina, VEGFR2 expression decreased in GCL and INL, and preserved the co-expression of VEGFR2 and NGF-tropomyosin-related kinase A (TrkA) receptor in retinal ganglion cells (RGCs). The WB analysis confirmed the NGF effect on VEGFR2 expression and activation, and showed a recovery of VEGF isoform dysregulation by suppressing STZ-induced VEGFA121 expression. Reduction in inflammatory and pro-apoptotic intracellular signals were also found in STZ+NGF retina. These findings suggest that ed-NGF administration might favor neuroretina protection, and in turn counteract the vascular impairment by regulating VEGFR2 and/or VEGFA isoform expression during the early stages of the disease. The possibility that an increase in the NGF availability might contribute to the switch from the proangiogenic/apoptotic to the neuroprotective action of VEGF is discussed.

Pilot Investigation on p75ICD Expression in Laryngeal Squamous Cell Carcinoma.

We investigated the p75 Neurotrophin Receptor (p75NTR) expression and cleavage product p75NTR Intracellular Domain (p75ICD) as potential oncogenic and metastatic markers in human Laryngeal Squamous Cell Carcinoma (LSCC). p75NTR is highly expressed in Cancer Stem Cells (CSCs) of the laryngeal epithelia and it has been proposed as a marker for stemness, cell migration, and chemo-resistance in different squamous carcinomas. To investigate the clinical significance of p75NTR cleavage products in solid tumors, full-length and cleaved p75NTR expression was analyzed in laryngeal primary tumors from different-stage LSCC patients, diagnosed at the Policlinico Umberto I Hospital. Molecular and histological techniques were used to detect the expressions of p75NTR and p75ICD, and ATP Binding Cassette Subfamily G Member 2 (ABCG2), a CSC marker. We found regulated p75NTR cleavage during squamous epithelial tumor progression and tissue invasion. Our preliminary investigation suggests p75ICD expression and localization as possible features of tumorigenesis and metastaticity. Its co-localization with ABCG2 in squamous cells in the parenchyma invaded by the tumor formation allows us to hypothesize p75NTR and p75ICD roles in tumor invasion and CSC spreading in LSCC patients. These data might represent a starting point for a comprehensive analysis of p75NTR cleavage and of its clinical relevance as a potential molecular LSCC signature, possibly helping diagnosis, and improving prognosis and personalized therapy.

hNGF Peptides Elicit the NGF-TrkA Signalling Pathway in Cholinergic Neurons and Retain Full Neurotrophic Activity in the DRG Assay.

In the last decade, Nerve Growth Factor (NGF)-based clinical approaches have lacked specific and efficient Tyrosine Kinase A (TrkA) agonists for brain delivery. Nowadays, the characterization of novel small peptidomimetic is taking centre stage in preclinical studies, in order to overcome the main size-related limitation in brain delivery of NGF holoprotein for Central Nervous System (CNS) pathologies. Here we investigated the NGF mimetic properties of the human NGF 1-14 sequence (hNGF1-14) and its derivatives, by resorting to primary cholinergic and dorsal root ganglia (DRG) neurons. Briefly, we observed that: 1) hNGF1-14 peptides engage the NGF pathway through TrkA phosphorylation at tyrosine 490 (Y490), and activation of ShcC/PI3K and Plc-γ/MAPK signalling, promoting AKT-dependent survival and CREB-driven neuronal activity, as seen by levels of the immediate early gene c-Fos, of the cholinergic marker Choline Acetyltransferase (ChAT), and of Brain Derived Neurotrophic Factor (BDNF); 2) their NGF mimetic activity is lost upon selective TrkA inhibition by means of GW441756; 3) hNGF1-14 peptides are able to sustain DRG survival and differentiation in absence of NGF. Furthermore, the acetylated derivative Ac-hNGF1-14 demonstrated an optimal NGF mimetic activity in both neuronal paradigms and an electrophysiological profile similar to NGF in cholinergic neurons. Cumulatively, the findings here reported pinpoint the hNGF1-14 peptide, and in particular its acetylated derivative, as novel, specific and low molecular weight TrkA specific agonists in both CNS and PNS primary neurons.

Cancer stem cells-driven tumor growth and immune escape: the Janus face of neurotrophins.

Cancer Stem Cells (CSCs) are self-renewing cancer cells responsible for expansion of the malignant mass in a dynamic process shaping the tumor microenvironment. CSCs may hijack the host immune surveillance resulting in typically aggressive tumors with poor prognosis.In this review, we focus on neurotrophic control of cellular substrates and molecular mechanisms involved in CSC-driven tumor growth as well as in host immune surveillance. Neurotrophins have been demonstrated to be key tumor promoting signaling platforms. Particularly, Nerve Growth Factor (NGF) and its specific receptor Tropomyosin related kinase A (TrkA) have been implicated in initiation and progression of many aggressive cancers. On the other hand, an active NGF pathway has been recently proven to be critical to oncogenic inflammation control and in promoting immune response against cancer, pinpointing possible pro-tumoral effects of NGF/TrkA-inhibitory therapy.A better understanding of the molecular mechanisms involved in the control of tumor growth/immunoediting is essential to identify new predictive and prognostic intervention and to design more effective therapies. Fine and timely modulation of CSCs-driven tumor growth and of peripheral lymph nodes activation by the immune system will possibly open the way to precision medicine in neurotrophic therapy and improve patient's prognosis in both TrkA- dependent and independent cancers.

The Copper(II)-Assisted Connection between NGF and BDNF by Means of Nerve Growth Factor-Mimicking Short Peptides.

Nerve growth factor (NGF) is a protein necessary for development and maintenance of the sympathetic and sensory nervous systems. We have previously shown that the NGF N-terminus peptide NGF(1-14) is sufficient to activate TrkA signaling pathways essential for neuronal survival and to induce an increase in brain-derived neurotrophic factor (BDNF) expression. Cu2+ ions played a critical role in the modulation of the biological activity of NGF(1-14). Using computational, spectroscopic, and biochemical techniques, here we report on the ability of a newly synthesized peptide named d-NGF(1-15), which is the dimeric form of NGF(1-14), to interact with TrkA. We found that d-NGF(1-15) interacts with the TrkA-D5 domain and induces the activation of its signaling pathways. Copper binding to d-NGF(1-15) stabilizes the secondary structure of the peptides, suggesting a strengthening of the noncovalent interactions that allow for the molecular recognition of D5 domain of TrkA and the activation of the signaling pathways. Intriguingly, the signaling cascade induced by the NGF peptides ultimately involves cAMP response element-binding protein (CREB) activation and an increase in BDNF protein level, in keeping with our previous result showing an increase of BDNF mRNA. All these promising connections can pave the way for developing interesting novel drugs for neurodegenerative diseases.

The Intersection of NGF/TrkA Signaling and Amyloid Precursor Protein Processing in Alzheimer's Disease Neuropathology.

Dysfunction of nerve growth factor (NGF) and its high-affinity Tropomyosin receptor kinase A (TrkA) receptor has been suggested to contribute to the selective degeneration of basal forebrain cholinergic neurons (BFCN) associated with the progressive cognitive decline in Alzheimer's disease (AD). The aim of this review is to describe our progress in elucidating the molecular mechanisms underlying the dynamic interplay between NGF/TrkA signaling and amyloid precursor protein (APP) metabolism within the context of AD neuropathology. This is mainly based on the finding that TrkA receptor binding to APP depends on a minimal stretch of ~20 amino acids located in the juxtamembrane/extracellular domain of APP that carries the α- and ß-secretase cleavage sites. Here, we provide evidence that: (i) NGF could be one of the "routing" proteins responsible for modulating the metabolism of APP from amyloidogenic towards non-amyloidogenic processing via binding to the TrkA receptor; (ii) the loss of NGF/TrkA signaling could be linked to sporadic AD contributing to the classical hallmarks of the neuropathology, such as synaptic loss, ß-amyloid peptide (Aß) deposition and tau abnormalities. These findings will hopefully help to design therapeutic strategies for AD treatment aimed at preserving cholinergic function and anti-amyloidogenic activity of the physiological NGF/TrkA pathway in the septo-hippocampal system.

NGF controls APP cleavage by downregulating APP phosphorylation at Thr668: relevance for Alzheimer's disease.

NGF has been implicated in forebrain neuroprotection from amyloidogenesis and Alzheimer's disease (AD). However, the underlying molecular mechanisms are still poorly understood. Here, we investigated the role of NGF signalling in the metabolism of amyloid precursor protein (APP) in forebrain neurons using primary cultures of septal neurons and acute septo-hippocampal brain slices. In this study, we show that NGF controls the basal level of APP phosphorylation at Thr668 (T668) by downregulating the activity of the Ser/Thr kinase JNK(p54) through the Tyr kinase signalling adaptor SH2-containing sequence C (ShcC). We also found that the specific NGF receptor, Tyr kinase A (TrkA), which is known to bind to APP, fails to interact with the fraction of APP molecules phosphorylated at T668 (APP(pT668) ). Accordingly, the amount of TrkA bound to APP is significantly reduced in the hippocampus of ShcC KO mice and of patients with AD in which elevated APP(pT668) levels are detected. NGF promotes TrkA binding to APP and APP trafficking to the Golgi, where APP-BACE interaction is hindered, finally resulting in reduced generation of sAPPß, CTFß and amyloid-beta (1-42). These results demonstrate that NGF signalling directly controls basal APP phosphorylation, subcellular localization and BACE cleavage, and pave the way for novel approaches specifically targeting ShcC signalling and/or the APP-TrkA interaction in AD therapy.

Loss of NGF-TrkA signaling from the CNS is not sufficient to induce cognitive impairments in young adult or intermediate-aged mice.

Many molecules expressed in the CNS contribute to cognitive functions either by modulating neuronal activity or by mediating neuronal trophic support and/or connectivity. An ongoing discussion is whether signaling of nerve growth factor (NGF) through its high-affinity receptor TrkA contributes to attention behavior and/or learning and memory, based on its expression in relevant regions of the CNS such as the hippocampus, cerebral cortex, amygdala and basal forebrain. Previous animal models carrying either a null allele or transgenic manipulation of Ngf or Trka have proved difficult in addressing this question. To overcome this problem, we conditionally deleted Ngf or Trka from the CNS. Our findings confirm that NGF-TrkA signaling supports survival of only a small proportion of cholinergic neurons during development; however, this signaling is not required for trophic support or connectivity of the remaining basal forebrain cholinergic neurons. Moreover, comprehensive behavioral analysis of young adult and intermediate-aged mice lacking NGF-TrkA signaling demonstrates that this signaling is dispensable for both attention behavior and various aspects of learning and memory.

Increased dopamine after mating impairs olfaction and prevents odor interference with pregnancy.

In rodents, social odor sensing influences female reproductive status by affecting neuroendocrine cascades. The odor of male mouse urine can induce ovulation or block pregnancy within 3 d post coitus. Females avoid the action of such olfactory stimuli after embryonic implantation. The mechanisms underlying these changes are unknown. Here we report that shortly after mating, a surge in dopamine in the mouse main olfactory bulb impairs the perception of social odors contained in male urine. Treatment of females at 6.5 d post coitus with a dopamine D2 receptor antagonist restores social odor sensing and favors disruption of pregnancy by inhibition of prolactin release, when administered in the presence of alien male urine odors. These results show that an active sensory barrier blocks social olfactory cues detrimental to pregnancy, consistent with the main olfactory bulb being a major relay through which social odor modulates reproductive status.

Nerve growth factor and brain-derived neurotrophic factor levels in patients with rheumatoid arthritis treated with TNF-alpha blockers.

Twenty consecutive rheumatoid arthritis (RA) patients (mean age 50.4 +/- 10.5 years; 17 females; mean disease duration 5.78 +/- 3.75 years) enrolled for tumor necrosis factor-alpha (TNF-alpha) blockers therapy (10 infliximab and 10 etanercept) were selected. Before starting therapy, 3 and 6 months thereafter all patients were evaluated for disease activity score (DAS), erythrocyte sedimentation rate (ESR), serum levels of C-reactive protein (CRP), interleukin-6 (IL-6), nerve growth factor (NGF), and brain-derived neurotrophic factor (BDNF). After 3 and 6 months a significant reduction in DAS, ESR, CRP, and IL-6 was observed, whereas no significant differences of NGF and BDNF serum levels were found. These preliminary results confirm that TNF-alpha blockers significantly improve disease activity and inflammation in RA; nevertheless further studies are needed to explain the mechanisms regulating NGF and BDNF release in RA patients treated with TNF-alpha blockers.

Neuronal markers expression of NGF-primed bone marrow cells (BMCs) transplanted in the brain of 6-hydroxydopamine and ibotenic acid lesioned littermate mice.

In the present study, we aim to show that non-adherent bone marrow cells (BMCs) express TrkA, the nerve growth factor (NGF) receptor, and that addition of NGF promotes the survival and neuronal commitment of BMC transplanted into the experimentally injured brain of littermates mice. Immunohistochemical analysis revealed that transplanted BMCs express tyrosine hydroxylase (TH) in proximity of the damaged dopaminergic tissues and choline acetyltransferase (ChAT) in the lesioned cholinergic regions. These results suggest that NGF supports the survival and differentiation of uncommitted BMCs and concurs with other local environmental signals to promote the expression of neuronal markers in these cells. The possible functional significance of these observations will be discussed.

Fighting in the aged male mouse increases the expression of TrkA and TrkB in the subventricular zone and in the hippocampus.

In the present study, we investigate the effect of aggressive behavior and release of nerve growth factor (NGF) on brain progenitor cells. We found that the condition of subordination enhances the level of NGF in the subventricular zone and hippocampus whereas dominance elevates brain derived neurotrophic factor (BDNF). It was also found that mRNA-TrkA is over-expressed in the subventricular zone and hippocampus of dominant and subordinate mice, whereas mRNA-TrkB is increased in the subventricular zone of both dominants and subordinates and in the hippocampus of dominant mice. Fighting was also associated with increased presence of proliferating cells in the hippocampus stained with the nuclear marker bromodeoxyuridine (BrdU). Moreover, the brain of subordinate mice displayed tyrosine hydroxylase (TH) immunoreactivity in the wall of the periventricular region of the third ventricles and a marked neuropeptide Y (NPY) presence in the hippocampus co-expressed with BrdU. These results provide additional evidence that agonistic behavior in the aged mouse alters neurotrophin levels and increases brain progenitor cells number. The functional significance of these findings is discussed.

Presence of nerve growth factor and TrkA expression in the SVZ of EAE rats: evidence for a possible functional significance.

Nerve growth factor (NGF) is a well-characterized neurotrophic factor that plays a crucial role during development in the growth, differentiation, and maintenance of brain neurons as well as in the reparative response of the adult brain to neuronal damage. Recent studies have shown that acute axonal loss occurs in multiple sclerosis (MS) and its animal model, experimental allergic encephalomyelitis (EAE), and that NGF suppresses clinical symptoms of EAE in nonhuman primates. Aim of the present study was to investigate the role of NGF in the regenerative response of the adult brain to neuronal damage occurring in EAE. Using EAE rats, we have found that exogenous NGF injection and NGF deprivation (NGF autoimmunization) can act on growth and differentiation of brain precursor cells in the subventricular zone (SVZ) of EAE rats. Moreover, NGF administration in brain of EAE rats stimulates the expression of early neuronal markers on proliferating precursor cells of the SVZ. The data obtained demonstrated that NGF and its antibody affect bromodeoxyuridine (BrdU) incorporation and NGF receptor expression by SVZ progenitor cells in the brain of EAE rats.

Structural and ultrastructural localization of NGF and NGF receptors in the thymus of subjects affected by myasthenia gravis.

We have previously reported that the thymus of patients affected by myasthenia gravis (MG) is characterized by an elevated level of nerve growth factor (NGF), an endogenous polypeptide which plays a marked role in the cell biology of nervous and immune system. A consistent number of studies has shown altered expression of NGF in diseases associated with inflammatory and/or autoimmune responses. To evaluate the biochemical and molecular mechanisms implicated in NGF action in human myasthenic thymus, it is important to identify the cellular and structural organization of NGF receptors. To address this question, we investigated, both at light and electron microscopic levels, the cellular distribution of immunoreactivity for NGF and its low-affinity receptors, (p75) and its high-affinity receptor (TrkA) in the thymus of patients with MG. The present investigation shows that NGF and NGF receptors are overexpressed in the thymic cells of patients with MG compared to control subjects.