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Cancer stem cells (CSCs), are thought to be at the origin of tumor development and resistance to therapies. Thus, a better understanding of the molecular mechanisms involved in the control of CSC stemness is essential to the design of more effective therapies for cancer patients. Cancer cell stemness and the subsequent expansion of CSCs are regulated by micro-environmental signals including neurotrophins. Over the years, the roles of neurotrophins in tumor development have been well established and regularly reviewed. Especially, nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are reported to stimulate tumor cell proliferation, survival, migration and/or invasion, and favors tumor angiogenesis. More recently, neurotrophins have been reported to regulate CSCs. This review briefly presents neurotrophins and their receptors, summarizes their roles in different cancers, and discusses the emerging evidence of neurotrophins-induced enrichment of CSCs as well as the involved signaling pathways. [ABSTRACT FROM AUTHOR]

Inflammation, demyelination, oligodendrocyte (OLG) death, and axonal degeneration are primary characteristics of multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). OLGs generate myelin sheaths that surround axons, while damage to OLGs leads to demyelination and neurological functional deficit. Matrine (MAT), a quinolizidine alkaloid derived from the herb Radix Sophorae Flave, has been recently found to effectively ameliorate clinical signs in EAE. Its therapeutic mechanism has, however, not been completely elucidated. In the present study, we found that MAT retarded the disease process, attenuated the clinical severity of EAE rats, ameliorated inflammation and demyelination, and suppressed the apoptosis of OLGs in the central nervous system (CNS) of EAE rats. In addition, MAT markedly blocked increased expression of the proNGF-p75 NTR death signaling complex, which is known to mediate OLG death in EAE animals. At the same time, MAT also prevented a decrease in the levels of NGF and its receptor TrkA, which together mediate the cell survival pathway and differentiation of OLGs. ProNGF, NGF, and the downstream effector proteins play an important role in the growth, differentiation, and apoptosis of OLGs as well as the reparative response to neuronal damage. These findings thus indicate that MAT improves clinical severity of EAE in part by reducing OLG apoptosis via restoring the ratios of proNGF:NGF and the respective receptors p75 NTR :TrkA in vivo. Taken together, these results suggest that MAT may be a promising agent for MS treatment based on its protective effect on OLGs. [ABSTRACT FROM AUTHOR]

Macrophage and microglial activation by HIV in the central nervous system (CNS) triggers the secretion of soluble factors which damage neurons. Therapeutic approaches designed to restore cognitive function by suppressing this inflammatory activity have not yet been successful. Recent studies have indicated that the phenotype of macrophages is differentially controlled by the mature and pro form of nerve growth factor. These cells therefore may be highly responsive to the imbalance in pro versus mature neurotrophins often associated with neurodegenerative diseases. In this study we evaluated the interactions between neurotrophins and HIV induced macrophage activation. HIV stimulation of macrophages induced a neurotoxic phenotype characterized by the expression of podosomes, suppression of calcium spiking and increased neurotoxin production. The secretome of the activated macrophages revealed a bias toward anti-angiogenic like activity and increased secretion of MMP-9. Co-stimulation with NGF and HIV suppressed neurotoxin secretion, increased calcium spiking, suppressed podosome expression and reversed 86 % of the proteins secreted in response to HIV, including MMP-9 and many growth factors. In contrast, co-stimulation of macrophages with proNGF not only failed to reverse the effects of HIV but increased the neurotoxic phenotype. These differential effects of proNGF and NGF on HIV activation provide a potential novel therapeutic avenue for controlling macrophage activation in response to HIV. [ABSTRACT FROM AUTHOR]

Nerve growth factor ( NGF) was first discovered approximately 60 years ago by Rita Levi-Montalcini as a protein that induces the growth of nerves. It is now known that NGF is also associated with Alzheimer's disease and intractable pain, and hence, it, along with its high-affinity receptor, tropomyosin receptor kinase (Trk) A, is considered to be 1 of the new targets for therapies being developed to treat these diseases. Anti- NGF antibody and TrkA inhibitors are known drugs that suppress NGF/TrkA signaling, and many drugs of these classes have been developed thus far. Interestingly, local anesthetics also possess TrkA inhibitory effects. This manuscript describes the development of an analgesic that suppresses NGF/TrkA signaling, which is anticipated to be 1 of the new methods to treat intractable pain. [ABSTRACT FROM AUTHOR]

Nerve growth factor (NGF) is critical for the proliferation, differentiation, and survival of neurons through its binding to the p75 NTR and TrkA receptors. Dysregulation of NGF has been implicated in several pathologies, including neurodegeneration (i.e., Parkinson's and Alzheimer's diseases) and both inflammatory and neuropathic pain states. Therefore, small molecule inhibitors that block NGF–receptor interactions have significant therapeutic potential. Small molecule antagonists ALE-0540, PD90780, Ro 08-2750, and PQC 083 have all been reported to inhibit NGF from binding the TrkA receptor. Interestingly, the characterization of the ability of these molecules to block NGF–p75 NTR interactions has not been performed. In addition, the inhibitory action of these molecules has never been evaluated using surface plasmon resonance (SPR) spectroscopy, which has been proven to be highly useful in drug discovery applications. In the current study, we used SPR biosensors to characterize the binding of NGF to the p75 NTR receptor in addition to characterizing the inhibitory potential of the known NGF antagonists. The results of this study provide the first evaluation of the ability of these compounds to block NGF binding to p75 NTR receptor. In addition, only PD90780 was effective at inhibiting the interaction of NGF with p75 NTR , suggesting receptor selectivity between known NGF inhibitors. [ABSTRACT FROM AUTHOR]

Neurotrophins, essential regulators of many aspects of neuronal differentiation and function, signal via four receptors, p75, TrkA, TrkB and TrkC. The three Trk paralogs are members of the LIG superfamily of membrane proteins, which share extracellular domains consisting of leucine-rich repeat and C2 Ig domains. Another LIG protein, LINGO-1 has been reported to bind and influence signaling of p75 as well as TrkA, TrkB and TrkC. Here we examine the manner in which LINGO-1 influences the function of TrkA, TrkB and TrkC. We report that Trk activation promotes Trk association with LINGO-1, and that this association promotes Trk degradation by a lysosomal mechanism. This mechanism resembles the mechanism by which another LIG protein, LRIG1, promotes lysosomal degradation of receptor tyrosine kinases such as the EGF receptor. We present evidence indicating that the Trk/LINGO-1 interaction occurs, in part, within recycling endosomes. We show that a mutant form of LINGO-1, with much of the extracellular domain deleted, has the capacity to enhance TrkA signaling in PC12 cells, possibly by acting as an inhibitor of Trk down-regulation by full length LINGO-1. We propose that LINGO-1 functions as a negative feedback regulator of signaling by cognate receptor tyrosine kinases including TrkA, TrkB and TrkC. [ABSTRACT FROM AUTHOR]

Chronic renal disease is known to alter olfactory function, but the specific changes induced in olfactory organs during this process remain unclear. Of the uraemic toxins generated during renal disease, high levels of urea are known to induce hyposmic conditions. In this study, the effects of environmental exposure to elevated concentrations of urea (7, 13.5 and 20 g L-1) on the sensory mucosa of zebrafish in acute toxicity and chronic toxicity tests were described. It was observed that lamellae maintained structural integrity and epithelial thickness was slightly reduced, but only following exposure to the highest concentrations of urea. Panneuronal labelling with anti-Hu revealed a negative correlation with levels of urea, leading to investigation of whether distinct neuronal subtypes were equally sensitive. Using densitometric analysis of immunolabelled tissues, numbers of Gα olf-, TRPC2- and TrkA-expressing cells were compared, representing ciliated, microvillous and crypt neurons, respectively. The three neuronal subpopulations responded differently to increasing levels of urea. In particular, crypt cells were more severely affected than the other cell types, and Gα olf-immunoreactivity was found to increase when fish were exposed to low doses of urea. It can be concluded that exposure to moderate levels of urea leads to sensory toxicity directly affecting olfactory organs, in accordance with the functional olfactometric measurements previously reported in the literature. [ABSTRACT FROM AUTHOR]

The distribution of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and their high-affinity tyrosine kinase receptors TrkA and TrkB was investigated by immunohistochemical method in the mammary gland of ewes from prepubertal stage to involution. NGF and BDNF protein expressions were strong during development of glands at prepubertal stage and during pregnancy and decreased during lactation and involution. The expressions localized in both stromal and parenchymal cells of developing gland were mainly arranged in the apical side of secretory cells during lactation. These observations were also confirmed at transcriptional level by RT-PCR analyses. The highest expression of all genes significantly occurred at prepubertal stage. NGF was then down-regulated from pregnancy to involution, and no statistical differences were observed among these stages. The receptor TrkA was also under-expressed from pregnancy to involution, and its expression significantly differed between pregnancy and 30 days of lactation and also between 30 and 60 days of lactation. BDNF was significantly down-regulated at 60 days of lactation in comparison with prepubertal stage and again between pregnancy and 30 days of lactation. The relative abundance of its receptor, TrkB, showed also a significant down-regulation at 60 days of lactation in comparison with pregnancy and involution. Among the myriad of other molecular signals involved in the mammary gland cycle, the local production of neuropeptides and their receptors could be of interest in understanding their potential role in mammary biology. [ABSTRACT FROM AUTHOR]

Our recent transcriptome profiling studies suggest that presenilin 1 (PS1) regulates expression of neural cell adhesion molecule (Ncam1) through p75 neurotrophin receptor. To better understand regulation of Ncam1 transcript and protein levels by p75, we performed a series of in vitro and in vivo experiments. The combined results suggest that p75 receptor is required for both resting and NGF-induced Ncam1 expression. Activation of TrkA receptors alone does not upregulate Ncam1. The normal Ncam1 expression depends on the relative ratio of TrkA and p75 receptors, and p75 extracellular domain is necessary for baseline Ncam1 expression. NGF-induced Ncam1 expression is dependent on the presence of an intact palmitoylation site within p75 receptor. Finally, we show that the expression of Ncam1 is altered in brains of two transgenic mouse lines that express familial Alzheimer's disease (FAD)-linked PS1 variants, suggesting that expression of dominantly inherited mutant PS1 genes interferes with the normal Ncam1 expression via the p75 signaling pathway.

Nerve growth factor (NGF) promotes the survival and differentiation of neurons. NGF is initially synthesized as a precursor, proNGF, which is the predominant form in the central nervous system. NGF and proNGF bind to TrkA/p75NTR to mediate cell survival and to sortilin/p75NTR to promote apoptosis. The ratio of TrkA to p75NTR affects whether proNGF and mature NGF signal cell survival or apoptosis. The purpose of this study was to determine whether the loss of TrkA influences p75NTR or sortilin expression levels, and to establish whether proNGF and mature NGF have a similar ability to switch between cell survival and cell death. We systematically altered TrkA receptor levels by priming cells with NGF, using small interfering RNA, and using the mutagenized PC12nnr5 cell line. We found that both NGF and proNGF can support cell survival in cells expressing TrkA, even in the presence of p75NTR and sortilin. However, when TrkA is reduced, proNGF signals cell death, while NGF exhibits no activity. In the absence of TrkA, proNGF-induced cell death occurs, even when p75NTR and sortilin levels are reduced. These results show that proNGF can switch between neurotrophic and apoptotic activity in response to changes in TrkA receptor levels, whereas mature NGF cannot. These results also support the model that proNGF is neurotrophic under normal circumstances, but that a loss in TrkA in the presence of p75NTR and sortilin, as occurs in neurodegenerative disease or injury, shifts proNGF, but not NGF, signalling from cell survival to cell death.

[Display omitted] • Novel phenyl thiazole/triazole derivatives were designed and synthesized. • 19c showed excellent inhibitory potency and selectivity on TrkA (IC 50 = 1.6 nM). • The binding mode of 19c with TrkA ideally elucidated its excellent enzymatic potency. • 19c suppressed migration and significantly inhibited KM-12 cell colony formation. • 19c weakly induced apoptosis of KM-12 cell in immunofluorescent staining analysis. Aiming to develop novel tropomyosin receptor kinase A (TrkA) inhibitors, a scaffold hopping strategy was utilized by transforming the fused indazole of Entrectinib to phenyl triazole/thiazole skeleton to obtain compounds 7a - 7 h and 13a - 13 h. In the light of MTT assay, phenyl triazole derivatives 7a-7 h exhibited moderate anti-proliferative activities against KM-12 cells with the IC 50 values of 1.78–17.51 μM, while phenyl thiazole derivatives 13a-13 h showed the weaker efficacy. Further structure-guided optimizations by combining the phenyl triazole skeleton with 3,5‑difluorophenyl and 3-carbamoyl-4-piperazinylaniline moiety led to compounds 19a - 19d and 20. Eventually, 19c bearing (2-(4-methylpiperazin-1-yl)phenyl)(morpholino)methanone moiety exhibited excellent anti-proliferative activity on TrkA-positive KM-12 cells with IC 50 value of 0.17 μM. Meanwhile, compound 19c showed the inhibitory potency on TrkA with IC 50 value of 1.6 nM, and displayed higher selectivity on TrkA over TrkB (IC 50 = 12.3 nM) and TrkC (IC 50 = 18.4 nM). The dedicated wound healing and colony formation assay indicated that the optimal compound 19c could suppress migration and significantly inhibit KM-12 cell colony formation in a dose-dependent manner. In addition, 19c could weakly induce apoptosis of KM-12 cell in immunofluorescent staining analysis. Taken together, the above results suggest 19c as a novel TrkA inhibitor worthy of further profiling. [ABSTRACT FROM AUTHOR]

• Neurotrophins have been linked to several disorders, including AD and depression. • Complex biology has led to difficulties in identifying suitable drug candidates. • Novel compounds targeting this biological system are now in clinical testing. • ACD856 is a clinical drug candidate targeting cognitive disorders, e.g. AD. Neurotrophins, such as brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF), are small proteins expressed in the brain and peripheral tissues, which regulate several key aspects of neuronal function, including neurogenesis, synaptic plasticity and neuroprotection, but also programmed cell death. This broad range of effects is a result of a complex downstream signaling pathway, with differential spatial and temporal activation patterns further diversifying their physiological effects. Alterations in neurotrophin levels, or known polymorphisms in neurotrophin genes, have been linked to a variety of disorders, including depression and Alzheimer's disease (AD). Historically, their therapeutic potential in these disorders has been hampered by the lack of suitable tool molecules for clinical studies. However, recent advancements have led to the development of new therapeutic candidates, which are now in clinical testing. [ABSTRACT FROM AUTHOR]

Brown adipogenesis and thermogenesis in brown and beige adipose tissue (AT) involve vascular remodeling and sympathetic neuronal guidance. Here, we investigated the molecular mechanism coordinating these processes. We used mouse models to identify the molecular target of a peptide CPATAERPC homing to the endothelium of brown and beige AT. We demonstrate that CPATAERPC mimics nerve growth factor (NGF) and identify a low molecular weight isoform of NGF receptor, TrkA, as the CPATAERPC cell surface target. We show that the expression of truncated endothelial TrkA is selective for brown and subcutaneous AT. Analysis of mice with endothelium-specific TrkA knockout revealed the role of TrkA in neuro-vascular coordination supporting the thermogenic function of brown adipocytes. A hunter-killer peptide D-BAT, composed of CPATAERPC and a pro-apoptotic domain, induced cell death in the endothelium and adipocytes. This resulted in thermogenesis impairment, and predisposed mice to obesity and glucose intolerance. We also tested if this treatment can inhibit the tumor recruitment of lipids mobilized from adipocytes from adjacent AT. Indeed, in a mouse model of breast cancer D-BAT suppressed tumor-associated AT lipolysis, which resulted in reduced fatty acid utilization by cancer cells. Our study demonstrates that TrkA signaling in the endothelium supports neuro-vascular coordination enabling beige adipogenesis. • Endothelial TrkA isoform expression is selective for brown and beige adipose tissue. • Endothelial TrkA isoform coordinates neuro-vasculature promoting thermogenesis. • Endothelial TrkA isoform TrkA coordinates neuro-vasculature promoting thermogenesis. • Peptide D-BAT mimicking NGF targets Endothelial TrkA in brown/beige adipose vasculature. • D-BAT causes brown/beige adipose tissue reorganization and suppresses its function. • Cancer-induced adipose tissue lipolysis and beiging is suppressed by D-BAT. [ABSTRACT FROM AUTHOR]

Post-therapeutic relapse, poor survival rates and increasing incidence justify the search for novel therapeutic targets and strategies in cutaneous malignant melanoma (CMM). Within this context, a potential oncogenic role for TrkA in CMM is suggested by reports of NTRK1 amplification, enhanced TrkA expression and intracellular TrkA activation associated with poor prognosis. TrkA, however, exhibits tumour-suppressing properties in melanoma cell lines and has recently been reported not to be associated with CMM progression. To better understand these contradictions, we present the first analysis of potential oncogenic alternative TrkA mRNA splicing, associated with TrkA immunoreactivity, in CMMs, and compare the behaviour of fully spliced TrkA and the alternative TrkAIII splice variant in BRAF(V600E) -mutated A375 melanoma cells. Alternative TrkA splicing in CMMs was associated with unfolded protein response (UPR) activation. Of the several alternative TrkA mRNA splice variants detected, TrkAIII was the only variant with an open reading frame and, therefore, oncogenic potential. TrkAIII expression was more frequent in metastatic CMMs, predominated over fully spliced TrkA mRNA expression in ≈50% and was invariably linked to intracellular phosphorylated TrkA immunoreactivity. Phosphorylated TrkA species resembling TrkAIII were also detected in metastatic CMM extracts. In A375 cells, reductive stress induced UPR activation and promoted TrkAIII expression and, in transient transfectants, promoted TrkAIII and Akt phosphorylation, enhancing resistance to reductive stress-induced death, which was prevented by lestaurtinib and entrectinib. In contrast, fully spliced TrkA was dysfunctional in A375 cells. The data identify fully spliced TrkA dysfunction as a novel mechanism for reducing melanoma suppression, support a causal relationship between reductive stress, UPR activation, alternative TrkAIII splicing and TrkAIII activation and characterise a targetable oncogenic pro-survival role for TrkAIII in CMM.

Competing Interests: The authors declare no conflicts of interest.

Background: Tropomyosin-related kinase A (TRKA) is a nerve growth factor (NGF) receptor that belongs to the tyrosine kinase receptor family. It is critical for the correct development of many types of neurons including pain-mediating sensory neurons and also controls proliferation, differentiation and survival of many neuronal and non-neuronal cells. TRKA (also known as NTRK1) gene is a target of alternative splicing which can result in several different protein isoforms. Presently, three human isoforms (TRKAI, TRKAII and TRKAIII) and two rat isoforms (TRKA L0 and TRKA L1) have been described.Results: We show here that human TRKA gene is overlapped by two genes and spans 67 kb--almost three times the size that has been previously described. Numerous transcription initiation sites from eight different 5' exons and a sophisticated splicing pattern among exons encoding the extracellular part of TRKA receptor indicate that there might be a large variety of alternative protein isoforms. TrkA genes in rat and mouse appear to be considerably shorter, are not overlapped by other genes and display more straightforward splicing patterns. We describe the expression profile of alternatively spliced TRKA transcripts in different tissues of human, rat and mouse, as well as analyze putative endogenous TRKA protein isoforms in human SH-SY5Y and rat PC12 cells. We also characterize a selection of novel putative protein isoforms by portraying their phosphorylation, glycosylation and intracellular localization patterns. Our findings show that an isoform comprising mainly of TRKA kinase domain is capable of entering the nucleus.Conclusions: Results obtained in this study refer to the existence of a multitude of TRKA mRNA and protein isoforms, with some putative proteins possessing very distinct properties. [ABSTRACT FROM AUTHOR]

Pulmonary arterial hypertension (PAH) is characterized by excessive proliferation, resistance to apoptosis, and increased migration of pulmonary artery smooth muscle cells (PASMCs). We hypothesized that quercetin exerts protective effects against this disease; thus, a chronic hypoxia model of PAH was generated using male Sprague-Dawley rats, which were treated with quercetin. In this model, quercetin prevented the development of PAH, right ventricular hypertrophy, and vascular remodeling after exposure to hypoxia. Quercetin inhibited PASMC proliferation and increased the apoptosis of PASMCs in vivo. In vitro, quercetin significantly inhibited hypoxia-induced PASMC proliferation, arrested cells in G1/G0 and inhibited cell migration in a dose-dependent manner. Moreover, our results showed that quercetin increased cyclin D1 protein levels and decreased the protein expression of cyclin B1 and Cdc2. Additionally, quercetin altered the Bax/Bcl-2 ratio and reduced MMP2, MMP9, CXCR4, integrin β1, and integrin α5 expression. Using genome-wide microarray analysis, we found that factors regulating proliferation, apoptosis, cell cycle, and migration were related to the tyrosine receptor kinase A (TrkA) pathway. In addition, activation of the TrkA/AKT signaling cascade during hypoxia was inhibited by quercetin in a dose-dependent manner. Moreover, quercetin alone inhibited the TrkA/AKT signaling pathway, resulting in decreased PASMC migration, cell cycle arrest and the induction of apoptosis. Our data suggest that quercetin is a potential candidate for the treatment of hypoxia-induced PAH. [ABSTRACT FROM AUTHOR]

Correct axonal growth during nervous system development is critical for synaptic transduction and nervous system function. Proper axon outgrowth relies on a suitable growing environment and the expression of a series of endogenous neuronal factors. However, the mechanisms of these neuronal proteins involved in neuronal development remain unknown. ZDHHC17 is a member of the DHHC (Asp-His-His-Cys)-containing family, a family of highly homologous proteins. Here, we show that loss of function of ZDHHC17 in zebrafish leads to motor dysfunction in 3-day post-fertilization (dpf) larvae. We performed immunolabeling analysis to reveal that mobility dysfunction was due to a significant defect in the axonal outgrowth of spinal motor neurons (SMNs) without affecting neuron generation. In addition, we found a similar phenotype in zdhhc17 siRNA-treated neural stem cells (NSCs) and PC12 cells. Inhibition of zdhhc17 limited neurite outgrowth and branching in both NSCs and PC12. Furthermore, we discovered that the level of phosphorylation of extracellular-regulated kinase (ERK) 1/2, a major downstream effector of tyrosine kinase (TrkA), was largely upregulated in ZDHHC17 overexpressing PC12 cells by a mechanism independent on its palmitoyltransferase (PAT) activity. Specifically, ZDHHC17 is necessary for proper TrkA–tubulin module formation in PC12 cells. These results strongly indicate that ZDHHC17 is essential for correct axon outgrowth in vivo and in vitro. Our findings identify ZDHHC17 as an important upstream factor of ERK1/2 to regulate the interaction between TrkA and tubulin during neuronal development. [ABSTRACT FROM AUTHOR]

We recently reported that increased NGF and p75 as well as decreased trkA characterized the Reelin-deprived ( E- Reeler) retina, prospecting a potential contribution of NGF during E- Reeler retinogenesis. Herein, retinal ganglion cells (RGCs), glial cells and rod bipolar cells (RBCs) were isolated from E- Reeler retinas, and NGF, trkA/p75 expression and apoptosis were investigated. E- Reeler ( n = 28) and E-control ( n = 34) retinas were digested, and RGCs, glial cells and RBCs were isolated by the magnetic bead separation. Expression of NGF, trkA, p75, Annexin V/PI and Bcl2/Bax was quantified by flow cytometry and validated by real-time PCR or WB. In E- Reeler retinas, NGF was significantly increased in RGCs and glial cells, p75 was increased in both RBCs and RGCs, and trkA was unchanged. In E-control retinas, NGF and p75 were expressed mainly in RBCs and RGCs and faintly in glial cells, while trkA was weakly expressed by RBCs and RGCs. In RBCs and RGCs, Annexin V expression was unchanged, while Bcl2 increased and Bax decreased selectively in E- Reeler RGCs. The data indicate that E- Reeler RBCs and RGCs overexpress NGF and p75 as a protective endogenous response to Reelin deprivation. The observation is strongly supported by the absence of apoptosis in both cell types. [ABSTRACT FROM AUTHOR]

Chromosomal rearrangements of the NTRK1 gene, which encodes the high affinity nerve growth factor receptor (tropomyosin related kinase, TRKA), have been observed in several epithelial cancers, such as colon cancer, papillary thyroid carcinoma or non small cell lung cancer. The various NTRK1 fusions described so far lead to constitutive activation of TRKA kinase activity and are oncogenic. We further investigated here the existence and the frequency of NTRK1 gene rearrangements in colorectal cancer. Using immunohistochemistry and quantitative reverse transcriptase PCR, we analyzed a series of human colorectal cancers. We identified two TRKA positive cases over 408, with NTRK1 chromosomal rearrangements. One of these rearrangements is a TPM3–NTRK1 fusion already observed in colon cancer, while the second one is a TPR–NTRK1 fusion never described in this type of cancer. These findings further confirm that translocations in the NTRK1 gene are recurring events in colorectal cancer, although occurring at a low frequency (around 0.5%). [ABSTRACT FROM AUTHOR]

TWIK-related acid-sensitive K+ (TASK) channels produce background K+ currents. We elucidated that TASK1 channels in rat adrenal medullary cells and PC12 cells are internalized in a clathrin-dependent manner in response to nerve growth factor (NGF). Here, the molecular mechanism for this internalization in PC12 cells was explored. The combination of enzyme inhibitors with tropomyosin receptor kinase A mutants revealed that the internalization was mediated by both phospholipase C and phosphatidylinositol 3-kinase pathways that converge on protein kinase C with the consequent activation of Src, a nonreceptor tyrosine kinase. The NGF-induced endocytosis of TASK1 channels did not occur in the presence of the Src inhibitor or with the expression of a kinase-dead Src mutant. Additionally, NGF induced a transient colocalization of Src with the TASK1 channel, but not the TASK1 mutant, in which tyrosine at 370 was replaced with phenylalanine. This TASK1 mutant showed no increase in tyrosine phosphorylation and markedly diminished internalization in response to NGF. We concluded that NGF induces endocytosis of TASK1 channels via tyrosine phosphorylation in its carboxyl terminus. [ABSTRACT FROM AUTHOR]

OBJECTIVE: We sought to investigate the intervention effect of nicotine on β-amyloid (Aβ)25-35 protein cytotoxicity in the rat basal forebrain neurons primary cultures. MATERIALS AND METHODS: For this purpose, freshly isolated rat basal forebrain neurons were cultured for 7 days and then exposed to either Aβ(25-35) or the combination of Aβ(25-35) and nicotine for 48 hours. The effects of Aβ(25-35) and nicotine on neurons morphology, growth status and TrkA expression were evaluated through microscopy, MTT assay, RT-PCR and immunocytochemistry. RESULTS: We found that the exposure of cultured neurons to Aβ(25-35) resulted in remarkable morphological changes. The average process number and length as well as the maximum process length of neurons were significantly decreased as compared with those of control. MTT assay showed that Aβ(25-35) impaired the growth of neurons. Aβ(25-35) also inhibited the expression of TrkA at both mRNA and protein levels. However, the addition of nicotine significantly attenuated these changes, indicating that nicotine could protect the neurons from the cytotoxicity of Aβ(25-35). CONCLUSIONS: Nicotine could be useful for the treatment of Alzheimer's disease through its ability to rescue the neurons from Aβ(25-35) cytotoxicity and the protective effect involved upregulated expression of TrkA receptors. [ABSTRACT FROM AUTHOR]

Introduction: Nerve growth factor (NGF) induces profound hyperalgesia. In this study we explored patterns of NGF sensitization in muscle and fascia of distal and paraspinal sites. Methods: We injected 1 µg of NGF into human (n?=?8) tibialis anterior and erector spinae muscles and their fasciae. The spatial extent of pressure sensitization, pressure pain threshold, and mechanical hyperalgesia (150 kPa, 10 s) was assessed at days 0.25, 1, 3, 7, 14, and 21. Chemical sensitization was explored by acidic buffer injections (pH 4, 100 µl) at days 7 and 14. Results: The mechanical hyperalgesia area was larger in tibial fascia than in muscle. Pressure pain thresholds were lower, tonic pressure pain ratings, and citrate buffer evoked pain higher in fascia than in muscle. Conclusions: Spatial mechanical sensitization differs between muscle and fascia. Thoracolumbar fasciae appear more sensitive than tibial fasciae and may be major contributors to low back pain, but the temporal sensitization profile is similar between paraspinal and distal sites. Muscle Nerve 52: 265-272, 2015. [ABSTRACT FROM AUTHOR]

Background: Chromodomain-helicase DNA binding protein 5 (CHD5) is an important tumor suppressor gene deleted from 1p36.31 in neuroblastomas (NBs). High CHD5 expression is associated with a favorable prognosis, but deletion or low expression is frequent in high-risk tumors. We explored the role of CHD5 expression in the neuronal differentiation of NB cell lines. Methods: NB cell lines SH-SY5Y (SY5Y), NGP, SK-N-DZ, IMR5, LAN5, SK-N-FI, NB69 and SH-EP were treated with 1–10 μM 13-cis-retinoic acid (13cRA) for 3–12 days. qRT-PCR and Western blot analyses were performed to measure mRNA and protein expression levels, respectively. Morphological differences were examined by both phase contrast and immunofluorescence studies. Results: Treatment of SY5Y cells with 13cRA caused upregulation of CHD5 expression in a time- and dose-dependent manner (1, 5, or 10 μM for 7 or 12 days) and also induced neuronal differentiation. Furthermore, both NGP and SK-N-DZ cells showed CHD5 upregulation and neuronal differentiation after 13cRA treatment. In contrast, 13cRA treatment of IMR5, LAN5, or SK-N-FI induced neither CHD5 expression nor neuronal differentiation. NB69 cells showed two different morphologies (neuronal and substrate adherent) after 12 days treatment with 10 μM of 13cRA. CHD5 expression was high in the neuronal cells, but low/absent in the flat, substrate adherent cells. Finally, NGF treatment caused upregulation of CHD5 expression and neuronal differentiation in SY5Y cells transfected to express TrkA (SY5Y-TrkA) but not in TrkA-null parental SY5Y cells, and both changes were blocked by a pan-TRK inhibitor. Conclusions: Treatment with 13cRA induces neuronal differentiation only in NB cells that upregulate CHD5. In addition, NGF induced CHD5 upregulation and neuronal differentiation only in TrkA expressing cells. Together, these results suggest that CHD5 is downstream of TrkA, and CHD5 expression may be crucial for neuronal differentiation induced by either 13cRA or TrkA/NGF signaling. [ABSTRACT FROM AUTHOR]

Acteoside, also known as verbascoside or orobanchin, is a common compound found in many important medicinal plants including the Chinese herb Cistanche deserticola Y. C. Ma, which is used for its neuroprotective and memory enhancement properties. We have investigated the effects of acteoside using a senescent mouse model induced by a combination of chronic intraperitoneal administration of d-gal (60 mg/kg/day) and oral administration AlCl3 (5 mg/kg/day) once daily for 90 days. After 60 days, acteoside (30, 60, and 120 mg/kg/day) was orally administered once daily for 30 days. The memory enhancing effects of acteoside were evaluated using the Morris water maze test. The results showed that 30-120 mg/kg/day of acteoside reduced the escape latency in finding the platform, and increased the number of crossings of the platform. A 30-120 mg/kg/day of acteoside increased significantly the expression of nerve growth factor and tropomycin receptor kinase A mRNA and protein in the hippocampus, measured using real-time RT-PCR, immunohistochemical analysis, and western blotting. These results support the use of C. deserticola for memory enhancement and indicate that the effects of acteoside are induced via promotion of nerve growth factor and tropomycin receptor kinase A expression. Copyright © 2015 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Background: Chronic psychological distress can cause neuroinflammation, but the involvement of leukocytes in this inflammatory response remains unclear. The area postrema (AP) is considered a neural-immune interface because it lacks a blood-brain barrier and a site for leukocyte recruitment in neuroinflammatory conditions induced by immunological insults, but its role in chronic psychological distress has not been explored. Objective: To determine leukocyte recruitment to the AP after chronic psychological distress. Methods: Rats were exposed to cat odor for 5 consecutive days to induce distress, and, on the 6th day, their brains were dissected to perform immunohistofluorescence studies of the AP. Immune cells were identified and quantified with CD45 and CD11b markers. The distribution of neurons and immune cells was determined using TrkA and CD45 markers, respectively. Results: Distress induced a significant increase in CD45+ and CD11b+ cells in the AP. Three immunophenotypes were determined in the control and distress groups: CD45+/CD11b-, CD45+/CD11b+ and CD45-/CD11b+. CD expression, morphology and fluorescence intensity enabled the identification of different immune cell types: starting from longitudinal ramified microglia (mainly in the control group) to amoeboid microglia, monocytes and lymphocytes (mostly in the distressed group). TrkA and CD45 expression in the AP revealed the proximity between soma neurons and leukocytes. Interestingly, some CD45+ cells expressed TrkA, with increased expression in the distressed group. Conclusions: The identification of microglial activation, leukocyte recruitment and the close proximity between neurons and leukocytes in the AP after chronic psychological distress exposure suggests the AP as a site for distress-induced immune responses and engraftment of leukocytes infiltrating the CNS. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Objective: The molecular mechanisms underlying perineural invasion (PNI)-a characteristic pathological feature of adenoid cystic carcinoma (ACC)-remain largely unclear. Recently, nerve growth factor (NGF) has been implicated in perineural invasion in certain malignancies. Overexpression of Myb related to the MYB-NFIB fusion gene in ACC has also been correlated with perineural invasion and survival. However, this concept is controversial. The aim of this study was to examine the expression of NGF together with its receptors, tropomyosin receptor kinase A (TrkA) and p75NRT, and Myb in ACC and assess their relationship with perineural invasion and survival. Study Design: Case series with chart review. Setting: The University of Tokyo Hospital. Subjects and Methods: We retrospectively analyzed 37 patients with ACC surgically treated from 1991 to 2011. We examined expression levels of NGF, TrkA, p75NRT, and Myb in the ACC specimens and their correlations with PNI and prognosis. Results: NGF, TrkA, p75NRT, and Myb overexpression rates were 65%, 65%, 30%, and 62%, respectively. Pearson product-moment correlation coefficient revealed a strong correlation between NGF/TrkA immunostaining and PNI (NGF: r = 0.68, P < .0001; TrkA: r = 0.53, P = .0007). Moreover, NGF overexpression was significantly associated with worse 8-year local control rate (27% vs 80%, P = .005). However, p75NRT and Myb expression was related to neither perineural invasion nor survival. Conclusion: Our findings demonstrated that NGF and TrkA overexpression, but not Myb and p75NRT overexpression, may contribute to PNI and thus cause local recurrence in patients with ACC. [ABSTRACT FROM AUTHOR]

The development of the nervous system is a temporally and spatially coordinated process that relies on the proper regulation of the genes involved. Neurotrophins and their receptors are directly responsible for the survival and differentiation of sensory and sympathetic neurons; however, it is not fully understood how genes encoding Trk neurotrophin receptors are regulated. Here, we show that rat Bex3 protein specifically regulates TrkA expression by acting at the trkA gene promoter level. Bex3 dimerization and shuttling to the nucleus regulate the transcription of the trkA promoter under basal conditions and also enhance nerve growth factor (NGF)-mediated trkA promoter activation. Moreover, qChIP assays indicate that Bex3 associates with the trkA promoter within a 150 bp sequence, immediately upstream from the transcription start site, which is sufficient to mediate the effects of Bex3. Consequently, the downregulation of Bex3 using shRNA increases neuronal apoptosis in NGF-dependent sensory neurons deprived of NGF and compromises PC12 cell differentiation in response to NGF. Our results support an important role for Bex3 in the regulation of TrkA expression and in NGF-mediated functions through modulation of the trkA promoter. [ABSTRACT FROM AUTHOR]

Background Perineural invasion (PNI) in desmoplastic melanoma is associated with increased local recurrence and reduced disease-free survival. The biological mechanisms underlying PNI remain unclear although several lines of evidence implicate neurotrophins and their receptors. Objectives We investigated the expression of p75NGFR and TrkA, and the presence of functional RET polymorphism (RETp) as they relate to PNI in desmoplastic melanoma. Methods In all, 43 cases of desmoplastic melanoma were immunohistochemically evaluated for TrkA and p75NGFR expression and RETp was detected by direct DNA sequencing. Results PNI was present in 67% of cases. On univariate analysis, p75NGFR was associated with PNI (expression detected in 79% of PNI-positive cases compared with 36% of PNI-negative cases, P = .005), increased Breslow depth ( P = .007), and greater Clark level ( P = .01). RETp was noted in 28% of cases but was not significantly associated with PNI ( P = .27) or other histopathologic variables. TrkA expression was absent in all cases. PNI was associated with increased Breslow depth and Clark level ( P = .01 and P = .009, respectively). Controlling for the association between p75NGFR and depth, p75NGFR remained associated with an increased propensity for PNI (odds ratio 4.68, P = .04). Limitations The sample size was limited. Conclusion In desmoplastic melanoma, p75NGFR expression is significantly associated with PNI and a more locally aggressive phenotype. [ABSTRACT FROM AUTHOR]

The tropomyosin-related kinase (Trk) family of receptor tyrosine kinases controls synaptic function, plasticity and sustains differentiation, morphology, and neuronal cell survival. Understanding Trk receptors down-regulation and recycling is a crucial step to point out sympathetic and sensory neuron function and survival. PC12 cells derived from pheochromocytoma of the rat adrenal medulla have been widely used as a model system for studies of neuronal differentiation as they respond to nerve growth factor (NGF) with a dramatic change in phenotype and acquire a number of properties characteristic of sympathetic neurons. In this study we demonstrated that in PC12 cells the TrkA receptor interacts with the deubiquitinating enzyme USP8/UBPy in a NGF-dependent manner and that it is deubiquitinated in vivo and in vitro by USP8. USP8 overexpression blocked NGF-induced neurites outgrowth while the overexpression of the catalytically inactive mutant USP8/UBPy C748A caused a marked increase of cell differentiation. Localization and biochemical experiments have point out that USP8 and TrkA partially co-localize in endosomes after NGF stimulation. Finally we have studied the role played by USP8 on TrkA turnover; using specific siRNA for USP8 we found that USP8 knockdown increases TrkA half-life, suggesting that the deubiquitinating activity of USP8 promotes TrkA degradation. [ABSTRACT FROM AUTHOR]

Development of a functional peripheral nervous system requires axons to rapidly innervate and arborize into final target organs and then slow but not halt their growth to establish stable connections while keeping pace with organ growth. Here we examine the role of the NGF-TrkA effector protein, Coronin-1, on postganglionic sympathetic neuron final target innervation. In the absence of Coronin-1 we find that NGF-TrkA-PI3K signaling drives robust axon growth and branching in part by suppressing GSK3β. In contrast, the presence of Coronin-1 (wild-type neurons) suppresses but does not halt NGF-TrkA-dependent growth and branching. This relative suppression in axon growth behaviors is due to Coronin-1-dependent calcium release via PLC-γ1 signaling, which releases PI3K-dependent suppression of GSK3β. Finally, we demonstrate that Coro1a-/- mice display sympathetic axon overgrowth and overbranching phenotypes in the developing heart. Together with previous work demonstrating the Coronin-1 expression is NGF dependent, this work suggests that periods before and after NGF-TrkA-induced Coronin-1 expression (and likely other factors) defines two distinct axon growth states, which are critical for proper circuit formation in the sympathetic nervous system. [ABSTRACT FROM AUTHOR]

Human podocytes are highly specialized cells with a key role in kidney physiology. Alteration of their structure as a consequence of injury or developmental failure leads to severe renal diseases. Although several studies have tried to elucidate the molecular framework of this cellular system, the functional bases for the maintenance of podocytes in their specialized state to sustain kidney barrier filtration are not completely understood. In this study, the capability of podocytes to produce and secrete the nerve growth factor (NGF) has been demonstrated via a validated in vitro model. During the process of cell differentiation, NGF and its receptors are modulated in human podocytes just as NGF-responsive neurons. Blockade of NGF biological activity results in severe changes of cell morphology. Collectively, our results outline a novel function of the neurotrophin and add a new cellular target in the complex biological framework of NGF. [ABSTRACT FROM AUTHOR]

We studied the changes in the expression of high-affinity TrkA and TrkB receptors of the nerve-growth factor and brain-derived neurotrophic factor in the rat hippocampus at different time points after the administration of Aβ(25-35) into the lateral cerebral ventricles. Studies on TrkA and TrkB expression were performed using immunohistochemistry and the method of the real-time polymerase chain reaction. We found that intracerebroventricular administration of Aβ(25-35) to rats resulted in long-term alterations in the system of neurotrophic signaling in the hippocampus. Expression of the TrkA receptor increased 28 days after treatment, whereas expression of the TrkB receptor increased 12 days and then decreased by 28 days after the treatment. Thus, alterations in neurotrophin signaling may be involved in the mechanisms that are responsible for Aβ(25-35)-induced impairment of hippocampal functions in rats. [ABSTRACT FROM AUTHOR]