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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.

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.

Background: Fetal alcohol spectrum disorders (FASD) are the manifestation of the damage caused by alcohol consumption during pregnancy. Children with fetal alcohol syndrome (FAS), the extreme FASD manifestation, show both facial dysmorphology and mental retardation. Alcohol consumed during gestational age prejudices brain development by reducing, among others, the synthesis and release of neurotrophic factors and neuroinflammatory markers. Alcohol drinking also induces oxidative stress. Hypothesis/Objective: The present study aimed to investigate the potential association between neurotrophins, neuroinflammation, and oxidative stress in 12 prepubertal male and female FASD children diagnosed as FAS or partial FAS (pFAS). Methods: Accordingly, we analyzed, in the serum, the level of BDNF and NGF and the oxidative stress, as free oxygen radicals test (FORT) and free oxygen radicals defense (FORD). Moreover, serum levels of inflammatory mediators (IL-1α, IL-2, IL-6, IL-10, IL-12, MCP-1, TGF-β, and TNF- α) involved in neuroinflammatory and oxidative processes have been investigated. Results: We demonstrated low serum levels of NGF and BDNF in pre-pubertal FASD children with respect to healthy controls. These changes were associated with higher serum presence of TNF- α and IL-1α. Quite interestingly, an elevation in the FORD was also found despite normal FORT levels. Moreover, we found a potentiation of IL-1α, IL-2, IL-10, and IL-1α1 in the analyzed female compared to male children. Conclusion: The present investigation shows an imbalance in the peripheral neuroimmune pathways that could be used in children as early biomarkers of the deficits observed in FASD.

Eye-drop recombinant human nerve growth factor (ed-rhNGF) has proved to recover the retina and optic nerve damage in animal models, including the unilateral optic nerve crush (ONC), and to improve visual acuity in humans. These data, associated with evidence that ed-rhNGF stimulates the brain derived neurotrophic factor (BDNF) in retina and cortex, suggests that NGF might exert retino-fugal effects by affecting BDNF and its receptor TrkB. To address these questions, their expression and relationship with the GABAergic and glutamatergic transmission markers, GAD65 and GAD67, vesicular inhibitory amino acid transporter (VGAT), and vesicular glutamate transporters 1 and 2 (VGLUT-1 and VGLUT-2) were investigated in adult ONC rats contralateral and ipsilateral visual cortex (VCx). Ed-rhNGF recovers the ONC-induced alteration of GABAergic and glutamatergic markers in contralateral VCx, induces an upregulation of TrkB, which is positively correlated with BDNF precursor (proBDNF) decrease in both VCx sides, and strongly enhances TrkB+ cell soma and neuronal endings surrounded by GAD65 immuno-reactive afferents. These findings contribute to enlarging the knowledge on the mechanism of actions and cellular targets of exogenously administrated NGF, and suggest that ed-rhNGF might act by potentiating the activity-dependent TrkB expression in GAD+ cells in VCx following retina damage and/or ONC.

The nerve growth factor (NGF) belongs to a family of proteins named neurotrophins, consisting of NGF, brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), NT-4/5 and NT-6. NGF regulates a large number of physiological mechanisms that result in neurotrophic, metabotrophic and/or immunotrophic effects. Neurodegenerative diseases, including Alzheimer disease, psychiatric disorders (e.g. depression and schizophrenia) and brain parasitic infection have in common the effect of changing the brain levels of neurotrophins, in particular NGF. The contribution of both NGF and its receptor TrkA in such events and the recent promising results of NGF based therapies are here presented and discussed.

Since 2004, vagus nerve stimulation (VNS) has been used in treatment-resistant or treatment-intolerant depressive episodes. Today, VNS is suggested as possible therapy for a larger spectrum of psychiatric disorders, including schizophrenia, obsessive compulsive disorders, and panic disorders. Despite a large body of literature supports the application of VNS in patients’ treatment, the exact mechanism of action of VNS remains not fully understood. In the present study, the major knowledges on the brain areas and neuronal pathways regulating neuroimmune and autonomic response subserving VNS effects are reviewed. Furthermore, the involvement of the neurotrophins (NTs) Nerve Growth Factor (NGF) and Brain Derived Neurotrophic Factor (BDNF) in vagus nerve (VN) physiology and stimulation is revised. The data on brain NGF/BDNF synthesis and in turn on the activity-dependent plasticity, connectivity rearrangement and neurogenesis, are presented and discussed as potential biomarkers for optimizing stimulatory parameters for VNS. A vagus nerve-neurotrophin interaction model in the brain is finally proposed as a working hypothesis for future studies addressed to understand pathophysiology of psychiatric disturbance.

Neurotrophins (NTs) are a family of polypeptides whose functions have been extensively studied in the past two decades. In particular, Nerve Growth Factor (NGF) and Brain-Derived Neurotrophic Factor (BDNF) play a major role in the development, nutrition and growth of the central and peripheral nervous system and in the pathogenesis of neurodegenerative, cardiometabolic and (auto)immune diseases. However, NGF and BDNF have subtle functions for follicular development, implantation, and placentation. This short narrative review summarizes the existing evidence, published between 2000 and 2019, about the role of NTs in many different conditions that might affect women during and after pregnancy such as preeclampsia, gestational diabetes, obesity, depression, anxiety, smoking and alcohol abuse. Literature suggests that the dysregulation of synthesis and release of NTs may lead to decisive effects on both maternal and fetal health. Some piece of evidences was found about a possible association between NGF/BDNF and breastfeeding. Additional studies on human models are necessary to further characterize the role of NTs in life-changing experiences like labor and delivery.

This study sought to evaluate the prospective role exerted by vascular endothelial growth factor (VEGF) in the modulation of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) signalling pathways in the rabbit retina. To reach this aim, the anti-VEGF agents aflibercept and ranibizumab were used as a pharmacological approach to evaluate the putative consequences elicited by VEGF inhibition on neurotrophin signalling. VEGF inhibition determined a marked imbalance in proneurotrophin expression, a significant reduction in TrkA and TrkB phosphorylation states and a decrease in the pan-neurotrophin receptor p75. Importantly, VEGF blockade also caused a strong increase in cleaved caspase-3, beclin-1 and lipidated LC3. The effects were more pronounced in the aflibercept group when compared with ranibizumab-treated rabbits, particularly 1 week after injection. This study demonstrates that VEGF exerts pivotal physiological roles in regulating NGF and BDNF pathways in the retina, as its inhibition by anti-VEGF agents deeply impacts neurotrophin homeostasis. These events are accompanied by a sustained induction of apoptotic and autophagic markers, suggesting that anti-VEGF-dependent impairments in neurotrophin signalling could be responsible for the activation of retinal cell death pathways.

Retinal ganglion cell (RGC) degeneration occurs within 2 weeks following optic nerve crush (ONC) as a consequence of reduced retro-transport of growth factors including nerve growth factor (NGF). The hypothesis that intravitreal (ivt) and eye drop (ed) administration of recombinant human NGF (rhNGF) might counteract ONC in adult rats is explored in this study. We found that both ivt- and ed-rhNGF reduced RGC loss and stimulated axonal regrowth. Chiefly, survival and regenerative effects of rhNGF were associated with a reduction of cells co-expressing Nogo-A/p75NTR at crush site borders, which contribute to glia scar formation following nerve injury, and induce further degeneration. We also found that ocular application of rhNGF reduced p75NTR and proNGF and enhanced phosphorylation of TrkA and its intracellular signals at retina level. Nogo-R and Rock2 expression was also normalized by ed-rhNGF treatment in both ONC and contralateral retina. Our findings that ocular applied NGF reaches and exerts biological actions on posterior segment of the eye give a further insight into the neurotrophin diffusion/transport through eye structures and/or their trafficking in optic nerve. In addition, the use of a highly purified NGF form in injury condition in which proNGF/p75NTR binding is favored indicates that increased availability of mature NGF restores the balance between TrkA and p75NGF, thus resulting in RGC survival and axonal growth. In conclusion, ocular applied NGF is confirmed as a good experimental paradigm to study mechanisms of neurodegeneration and regeneration, disclose biomarkers, and time windows for efficacy treatment following cell or nerve injury.

Intravitreal injection (IVT) of antivascular endothelial growth factor (anti-VEGF) agents is widely used for the treatment of retinal vascular diseases. Recently, the injection of anti-VEGF agents in the ocular anterior chamber has been proposed for the treatment of neovascular glaucoma and potential side effects on the corneal structures have been investigated with contrasting results. Increasing evidence has demonstrated that VEGF inhibition is associated with cellular apoptotic changes and that this effect may be mediated by alterations in nerve growth factor (NGF) pathway. In this study, we demonstrated that anterior chamber injection (IC), but not IVT injection of two different anti-VEGF agents, aflibercept and ranibizumab, affects rabbit corneal endothelium in terms of survival and apoptosis and is associated with changes in endothelial expression of NGF precursor (proNGF) and p75 neurotrophin receptor (p75NTR) receptor. We observed an increase in corneal endothelial cell incorporation of trypan blue and expression of cleaved-caspase 3 (c-Casp3), p75NTR, and RhoA after IC injection of both anti-VEGF drugs when compared with the vehicle. Our results showed that apoptosis induction by aflibercept was more pronounced when compared with that of ranibizumab. Aflibercept also mediated a significant increase in endothelial expression of proNGF when compared with the vehicle. In line with these data, IC administration of both anti-VEGF agents induced the activation of apoptotic signals in endothelial cells, including an increase in c-Casp3, decrease in Bad Ser 112 phosphorylation, and unbalance of AKT phosphorylation. These results demonstrated that administration of anti-VEGF in the anterior chamber of rabbit affects endothelial cell survival by inducing apoptosis through alteration of NGF pathway.

The eye is a central nervous system structure that is uniquely accessible to local treatment. Through the ocular surface, it is possible to access the retina, optic nerve, and brain. Animal models of retina degeneration or optic nerve crush could thus serve as tools to investigate whether and how factors, which are anterogradely or retrogradely transported through the optic nerve, might contribute to activate neuroprotection and eventually regeneration. Among these factors, nerve growth factor (NGF) plays a crucial role during development of the visual system, as well as during the entire life span, and in pathological conditions. The ability of NGF to exert survival and trophic actions on the retina and brain cells when applied intraocularly and topically as eye drops is critically reviewed here, together with the effects of ocular neurotrophins on neuronal pathways influencing body rhythm, cognitions, and behavioral functions. The latest data from animal models and humans are presented, and the mechanism of action of ocularly administered NGF is discussed. NGF eye drops are proposed as an experimental strategy to investigate the role and cellular targets of neurotrophins in the mechanism(s) underlying neurodegeneration/regeneration and their involvement in the regulation of neurological and behavioral dysfunctions.

The eye is a central nervous system structure that is uniquely accessible to local treatment. Through the ocular surface, it is possible to access the retina, optic nerve, and brain. Animal models of retina degeneration or optic nerve crush could thus serve as tools to investigate whether and how factors, which are anterogradely or retrogradely transported through the optic nerve, might contribute to activate neuroprotection and eventually regeneration. Among these factors, nerve growth factor (NGF) plays a crucial role during development of the visual system, as well as during the entire life span, and in pathological conditions. The ability of NGF to exert survival and trophic actions on the retina and brain cells when applied intraocularly and topically as eye drops is critically reviewed here, together with the effects of ocular neurotrophins on neuronal pathways influencing body rhythm, cognitions, and behavioral functions. The latest data from animal models and humans are presented, and the mechanism of action of ocularly administered NGF is discussed. NGF eye drops are proposed as an experimental strategy to investigate the role and cellular targets of neurotrophins in the mechanism(s) underlying neurodegeneration/regeneration and their involvement in the regulation of neurological and behavioral dysfunctions.

The present study was aimed at evaluating whether single intermittent acute cervical vagus nerve stimulation (ACVS), pro-vided at a frequency which exhibits a clinical efficacy, may influence brain neurotrophins and hippocampal plasticity. With this purpose, the brain of adult male rats undergoing ACVS was used to analyze the expression of Nerve Growth Factor (NGF) and Brain-Derived Neurotrophic Factor (BDNF) in brain areas known to synthetize these growth factors, and the expression the neural cell adhesion molecule (NCAM), the synaptophysin (SYP) and biosynthetic GABA (GAD67) in the hippocampus. The effects of ACVS on NGF and BDNF protein and mRNA in hippocampus, hypothalamus and cortex two hours after stimulation were shown to be dependent on the frequencies of ACVS stimulation. Prolonged (three days post stimulation) modifications of NGF and BDNF were also observed in the hippocampus of ACVS rats. An early enhancement of the plasticity markers NCAM, SYP and GAD67 was also found in ACVS hippocampus. Three days after stimulation, NCAM and GAD67 levels were still higher than controls. Immunohistochemistry confirms the stimulatory effects of ACVS on GABA showing an increase in GAD67-positive cells in the dentate gyrus and CA3 hippocampal areas. This study shows that ACVS affects brain NGF and BDNF synthesis in a frequency-dependent manner. Neurotrophins changes are associated with increased hippocampal plasticity, as demonstrated by the observed molecular and morphological modifications. These findings support the role of brain neurotrophins in the ACVS mechanism of action.

SummaryAims Nerve growth factor (NGF) eyedrops (ed-NGF) activate brain neurons, stimulate growth factors, including brain-derived neurotrophic factor (BDNF), and exert neuroprotection in the forebrain of streptozotocin-induced diabetic rats (STZ rats). In this study, the effects of ed-NGF on BDNF signaling in the prefrontal cortex (PFC) were explored in healthy and STZ-diabetic rats, in which cortical neuronal and axonal loss, and altered circulating BDNF associated with depressive phenotype are also described. Methods STZ and healthy (CTR) adult rats received ed-NGF twice a day for 2 weeks. Depressive phenotype was identified by force swimming test (FST). Proteins extracted from PFC were processed for ELISA and Western blot analyses to measure the expression of BDNF, proBDNF, and their receptors and intracellular signals. Results ed-NGF treatment modulates BDNF pathway in PFC and normalizes the STZ-induced BDNF alterations by stimulating TRK-mediated survival mechanism. A decreased latency in FST was also found in STZ rats, while no change was observed comparing CTR + NGF and STZ + NGF with CTR. Conclusion The present data confirm the capacity of ed-NGF treatment to affect brain neurons and lead to brain damage recovery by activating protective and remodeling pathways triggered by BDNF. We suggest that the ed-NGF-induced changes in BDNF signaling might influence the manifestation of depressive phenotype in diabetic rats.

Nerve growth factor (NGF) exerts protective actions in the healthy and diseased nervous system. Intranasal administration is a suitable and safe strategy to deliver NGF to CNS neurons. We investigated whether nasal anti-NGF-antibody (ANA) administration affects neuronal autophagy, in view of its putative regulatory role in this process. We focused on olfactory bulbs (OB), neocortex (Cx), hippocampus (HF) and septal complex (SC), known to be NGF-responsive and autophagically active. Our combined molecular/morphological results demonstrate that intranasally administered ANA reaches brain NGF-target neurons and lowers the levels of endogenous NGF and its receptors. Treatment also affects - in a brain region-dependent manner - the expression of the autophagic proteins Beclin-1 and Ambra1, as well as that of proteins belonging to the Bcl2 family, namely Bax and Bcl-2, reflecting apoptotic dysregulation. This study provides a nongenetically modified, NGF-defective animal model, representing a suitable tool to investigate novel properties of the neurotrophin, especially in relation to autophagy.

http://hdl.handle.net/20.500.11768/96884 Nerve growth factor (NGF) is suggested to be neuroprotective after nerve injury; however, retinal ganglion cells (RGC) degenerate following optic-nerve crush (ONC), even in the presence of increased levels of endogenous NGF. To further investigate this apparently paradoxical condition, a time-course study was performed to evaluate the effects of unilateral ONC on NGF expression and signaling in the adult retina. Visually evoked potential and immunofluorescence staining were used to assess axonal damage and RGC loss. The levels of NGF, proNGF, p75NTR, TrkA and GFAP and the activation of several intracellular pathways were analyzed at 1, 3, 7 and 14 days after crush (dac) by ELISA/Western Blot and PathScan intracellular signaling array. The progressive RGC loss and nerve impairment featured an early and sustained activation of apoptotic pathways; and GFAP and p75NTR enhancement. In contrast, ONC-induced reduction of TrkA, and increased proNGF were observed only at 7 and 14 dac. We propose that proNGF and p75NTR contribute to exacerbate retinal degeneration by further stimulating apoptosis during the second week after injury, and thus hamper the neuroprotective effect of the endogenous NGF. These findings might aid in identifying effective treatment windows for NGF-based strategies to counteract retinal and/or optic-nerve degeneration.